Research Papers of Antrodia camphorata Search from PubMed http://www.ncbi.nlm.nih.

gov/sites/entrez keyword: camphorata or Taiwanofungus as of 2-4-2013 Table of Contents

1: Chemical Characterization and Anti-inflammatory Effect of Polysaccharides Fractionated from Submerge-Cultured Antrodia camphorata Mycelia. ................................................................................. 8 2: Unique Formosan Mushroom Antrodia camphorata Differentially Inhibits Androgen-Responsive LNCaP and -Independent PC-3 Prostate Cancer Cells. ............................................................................ 8 3: Production of a COX-2 inhibitor, 2,4,5-trimethoxybenzaldehyde, with submerged cultured Antrodia camphorata. ............................................................................................................................................... 8 4: Inhibition of cyclooxygenase-2 and induction of apoptosis in estrogen-nonresponsive breast cancer cells by Antrodia camphorata. ................................................................................................................... 9 5: Modulation of inflammation-related genes of polysaccharides fractionated from mycelia of medicinal basidiomycete Antrodia camphorata. ....................................................................................... 9 6: Antihypertensive activities of a solid-state culture of Taiwanofungus camphoratus (Chang-chih) in spontaneously hypertensive rats.............................................................................................................. 10 7: Biochemical characterization of 1-Cys peroxiredoxin from Antrodia camphorata. ........................... 10 8: Protective effects of mycelia of Antrodia camphorata and Armillariella tabescens in submerged culture against ethanol-induced hepatic toxicity in rats. ......................................................................... 10 9: Phylogenetic analysis of Antrodia species and Antrodia camphorata inferred from internal transcribed spacer region. ........................................................................................................................11 10: Biochemical characterization of a novel 2-Cys peroxiredoxin from Antrodia camphorata. .............11 11: Antrodia camphorata extract induces replicative senescence in superficial TCC, and inhibits the absolute migration capability in invasive bladder carcinoma cells. .........................................................11 12: Filtrate of fermented mycelia from Antrodia camphorata reduces liver fibrosis induced by carbon tetrachloride in rats.................................................................................................................................. 12 13: Neuroprotective diterpenes from the fruiting body of Antrodia camphorata. .................................. 12 14: Antrodia camphorata in submerged culture protects low density lipoproteins against oxidative modification. ........................................................................................................................................... 12 15: Human urinary bladder cancer T24 cells are susceptible to the Antrodia camphorata extracts. ...... 13 16: Adenosine as an active component of Antrodia cinnamomea that prevents rat PC12 cells from serum deprivation-induced apoptosis through the activation of adenosine A(2A) receptors. ................ 13 17: Proteomic analysis of the effect of Antrodia camphorata extract on human lung cancer A549 cell. 14 18: Growth inhibition and induction of apoptosis in MCF-7 breast cancer cells by Antrodia camphorata. ................................................................................................................................................................. 14 19: Anti-inflammatory potential of Antrodia Camphorata through inhibition of iNOS, COX-2 and cytokines via the NF-kappaB pathway. ................................................................................................... 14 20: Promotion of hyphal growth and underlying chemical changes in Antrodia camphorata by host factors from Cinnamomum camphora. ................................................................................................... 15 21: Mycelia from Antrodia camphorata in Submerged culture induce apoptosis of human hepatoma
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HepG2 cells possibly through regulation of Fas pathway. ...................................................................... 15 22: Induction of apoptosis in human hepatoma cells by mycelia of Antrodia camphorata in submerged culture...................................................................................................................................................... 15 23: Apoptotic effects of extract from Antrodia camphorata fruiting bodies in human hepatocellular carcinoma cell lines. ................................................................................................................................ 16 24: Antrodia camphorata prevents rat pheochromocytoma cells from serum deprivation-induced apoptosis.................................................................................................................................................. 16 25: Characterization and functional study of Antrodia camphorata lipopolysaccharide. ....................... 16 26: Antitumor effects of the partially purified polysaccharides from Antrodia camphorata and the mechanism of its action........................................................................................................................... 17 27: Induction of apoptosis by Antrodia camphorata in human premyelocytic leukemia HL-60 cells.... 17 28: Evaluation of the anti-inflammatory activity of zhankuic acids isolated from the fruiting bodies of Antrodia camphorata. .............................................................................................................................. 18 29: Anti-inflammatory activity of the extracts from mycelia of Antrodia camphorata cultured with water-soluble fractions from five different Cinnamomum species. ........................................................ 18 30: Five new maleic and succinic acid derivatives from the mycelium of Antrodia camphorata and their cytotoxic effects on LLC tumor cell line. ............................................................................................... 19 31: The vasorelaxation of Antrodia camphorata mycelia: involvement of endothelial Ca(2+)-NO-cGMP pathway. .................................................................................................................................................. 19 32: Antioxidative and hepatoprotective effects of Antrodia camphorata extract. ................................... 19 33: Protective effects of fermented filtrate from Antrodia camphorata in submerged culture against CCl4-induced hepatic toxicity in rats...................................................................................................... 20 34: Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes. ................................................................................................................................ 20 35: Antioxidant properties of Antrodia camphorata in submerged culture. ............................................ 20 36: Antrodia camphorata polysaccharides exhibit anti-hepatitis B virus effects. ................................... 21 37: Anti-inflammatory Benzenoids from Antrodia camphorata. ............................................................ 21 38. Comparative anti-inflammatory characterization of wild fruiting body, liquid-state fermentation, and solid-state culture of Taiwanofungus camphoratus in microglia and the mechanism of its action .. 21 39. A highly stable cambialistic-superoxide dismutase from Antrodia camphorata: Expression in yeast and enzyme properties ............................................................................................................................ 22 40: A New Cytotoxic Agent from Solid-State Fermented Mycelium of Antrodia camphorata. ............. 22 41: Evaluation of the anti-inflammatory and anti-proliferation tumoral cells activities of Antrodia camphorata, Cordyceps sinensis, and Cinnamomum osmophloeum bark extracts. ............................... 23 42: In vivo immunomodulatory effects of Antrodia camphorata polysaccharides in a T1/T2 doubly transgenic mouse model for inhibiting infection of Schistosoma mansoni. ........................................... 23 43. Fermented Antrodia cinnamomea Extract Protects Rat PC12 Cells from Serum Deprivation-Induced Apoptosis: The Role of the MAPK Family. ............................................................................................ 24 44. Isolation and analysis of genes specifically expressed during basidiomatal development in Antrodia cinnamomea by subtractive PCR and cDNA microarray. ....................................................................... 24
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45. DPPH radical scavenging and semicarbazide-sensitive amine oxidase inhibitory and cytotoxic activities of Taiwanofungus camphoratus (Chang-chih)......................................................................... 24 46. Effects on Tyrosinase Activity by the Extracts of Ganoderma lucidum and Related Mushrooms. .. 24 47. Polysaccharides from Antrodia camphorata mycelia extracts possess immunomodulatory activity and inhibits infection of Schistosoma mansoni....................................................................................... 25 48: The adjuvant effects of Antrodia Camphorata extracts combined with anti-tumor agents on multidrug resistant human hepatoma cells. ............................................................................................. 25 49: Anti-inflammatory Activities of New Succinic and Maleic Derivatives from the Fruiting Body of Antrodia camphorata. .............................................................................................................................. 26 50. Antioxidant activity of Antrodia camphorata on free radical-induced endothelial cell damage. ...... 26 51. Effects on Tyrosinase Activity by the Extracts of Ganoderma lucidum and Related Mushrooms. .. 26 52 Antrodia camphorata inhibits proliferation of human breast cancer cells in vitro and in vivo. ......... 27 53. Active extracts of wild fruiting bodies of Antrodia camphorata (EEAC) induce leukemia HL 60 cells apoptosis partially through histone hypoacetylation and synergistically promote anticancer effect of trichostatin A. ...................................................................................................................................... 27 54: Effects of antrodia camphorata on viability, apoptosis, and [Ca2+]i in PC3 human prostate cancer cells. ........................................................................................................................................................ 27 55. Immunomodulatory effect of Antrodia camphorata mycelia and culture filtrate. ............................ 28 56. Probing Inhibitory Effects of Antrodia camphorata Isolates Using Insect Cell-Based Impedance Spectroscopy: Inhibition vs Chemical Structure. .................................................................................... 28 57. An Extract of Antrodia camphorata Mycelia Attenuates the Progression of Nephritis in Systemic Lupus Erythematosus-Prone NZB/W F1 Mice ....................................................................................... 28 58. Niuchangchih (Antrodia camphorata) and its potential in treating liver diseases. ........................... 29 59. New Constituents with iNOS Inhibitory Activity from Mycelium of Antrodia camphorata. ........... 29 60. Purification, Cloning, and Functional Characterization of a Novel Immunomodulatory Protein from Antrodia camphorata (Bitter Mushroom) That Exhibits TLR2-Dependent NF-kappaB Activation and M1 Polarization within Murine Macrophages. ....................................................................................... 29 61. Cytotoxic triterpenes from Antrodia camphorata and their mode of action in HT-29 human colon cancer cells. ............................................................................................................................................. 30 62. Cyclodextrin-modified capillary electrophoresis for achiral and chiral separation of ergostane and lanostane compounds extracted from the fruiting body of Antrodia camphorata. .................................. 30 63. Review of Pharmacological Effects of Antrodia camphorata and its Bioactive Compounds. .......... 31 64. Preferential blockade of dioxin-induced activation of the Aryl hydrocarbon receptor by Antrodia camphorata. ............................................................................................................................................. 31 65. Rapid Evaluation of Antrodia camphorata Natural Products and Derivatives in Tumourigenic Liver Progenitor Cells with a Novel Cell Proliferation Assay. ......................................................................... 31 66. Effects of Antrodia camphorata extracts on the viability, apoptosis, [Ca2+]i, and MAPKs phosphorylation of OC2 human oral cancer cells. .................................................................................. 32 67. Protective effect of Antrodia Camphorata on bladder ischemia/reperfusion injury. ......................... 32 68. Taiwanofungus camphoratus activates peroxisome proliferator-activated receptors and induces
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hypotriglyceride in hypercholesterolemic rats. ....................................................................................... 33 69. Cloning, expression, and characterization of an enzyme possessing both glutaredoxin and dehydroascorbate reductase activity from Taiwanofungus camphorata. ................................................ 33 70. Inhibition of Anchorage-Independent Proliferation and G0/G1 Cell-Cycle Regulation in Human Colorectal Carcinoma Cells by 4,7-Dimethoxy-5-methyl-l,3-benzodioxole Isolated from the Fruiting Body of Antrodia camphorate. ................................................................................................................ 33 71. The augmented anti-tumor effects of Antrodia camphorata co-fermented with Chinese medicinal herb in human hepatoma cells. ................................................................................................................ 34 72. Antroquinonol displays anticancer potential against human hepatocellular carcinoma cells: a crucial role of AMPK and mTOR pathways. ...................................................................................................... 34 73. Dehydroeburicoic acid induces calcium- and calpain-dependent necrosis in human U87MG glioblastomas. ......................................................................................................................................... 35 74. Administration of polysaccharides from Antrodia camphorata modulates dendritic cell function and alleviates allergen-induced T helper type 2 responses in a mouse model of asthma. ............................. 35 75. Apoptotic effects of a high performance liquid chromatography (HPLC) fraction of Antrodia camphorata mycelia are mediated by down-regulation of the expressions of four tumor-related genes in human non-small cell lung carcinoma A549 cell. ................................................................................... 36 76. Antrocamphin A, an Anti-inflammatory Principal from the Fruiting Body of Taiwanofungus camphoratus , and Its Mechanisms. ........................................................................................................ 36 77. Fruiting Body of Niuchangchih (Antrodia camphorata) Protects Livers against Chronic Alcohol Consumption Damage. ............................................................................................................................ 36 78. Quality evaluation of mycelial Antrodia camphorata using high-performance liquid chromatography (HPLC) coupled with diode array detector and mass spectrometry (DAD-MS). ........ 37 79. Cloning, expression, and characterization of a thioredoxin reductase cDNA from Taiwanofungus camphorata .............................................................................................................................................. 37 80. Promoting effect of Antrodia camphorata as an immunomodulating adjuvant on the antitumor efficacy of HER-2/neu DNA vaccine. ..................................................................................................... 37 81. Analgesic Effects and the Mechanisms of Anti-inflammation of Ergostatrien-3beta-ol from Antrodia camphorata Submerged Whole Broth in Mice. ....................................................................................... 38 82. Development of a LC-MS/MS method for the determination of antrodin B and antrodin C from Antrodia camphorata extract in rat plasma for pharmacokinetic study................................................... 38 83. Methyl Antcinate A from Antrodia camphorata Induces Apoptosis in Human Liver Cancer Cells through Oxidant-Mediated Cofilin- and Bax-Triggered Mitochondrial Pathway. .................................. 39 84. Dietary effect of Antrodia Camphorate extracts on immune responses in WEHI-3 leukemia BALB/c mice. ........................................................................................................................................................ 39 85. Further studies on the hepatoprotective effect of Antrodia camphorata in submerged culture on ethanol-induced acute liver injury in rats. ............................................................................................... 40 86. Antrodia camphorata suppresses lipopolysaccharide-induced nuclear factor-kappaB activation in transgenic mice evaluated by bioluminescence imaging. ....................................................................... 40 87. Structure and functions of gamma-dodecalactone isolated from Antrodia camphorata for NK cell
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activation. ................................................................................................................................................ 40 88. Methylantcinate A induces tumor specific growth inhibition in oral cancer cells via Bax-mediated mitochondrial apoptotic pathway. ........................................................................................................... 41 89. Anti-metastatic activities of Antrodia camphorata against human breast cancer cells mediated through suppression of the MAPK signaling pathway. ........................................................................... 41 90. First total synthesis of antrocamphin A and its analogs as anti-inflammatory and anti-platelet aggregation agents................................................................................................................................... 41 91. Biologically active constituents from the fruiting body of Taiwanofungus camphoratus. ............... 42 92. Identification of Antrocin from Antrodia camphorata as a Selective and Novel Class of Small Molecule Inhibitor of Akt/mTOR Signaling in Metastatic Breast Cancer MDA-MB-231 Cells. .......... 42 93. Antroquinonol inhibits NSCLC proliferation by altering PI3K/mTOR proteins and miRNA expression profiles. ................................................................................................................................. 42 94. Antroquinonol reduces oxidative stress through enhancing Nrf2 signaling pathway and inhibits inflammation and sclerosis in focal segmental glomerulosclerosis mice. .............................................. 43 95. Monothiol Glutaredoxin cDNA from Taiwanofungus camphorata : A Novel CGFS-type Glutaredoxin Possessing Glutathione Reductase Activity. ..................................................................... 43 96. Effects of Antrodia camphorata on Alcohol Clearance and Antifibrosis in Livers of Rats Continuously Fed Alcohol. ..................................................................................................................... 44 97. Is 2,3,4,5-tetramethoxybenzoyl chloride a natural product? ............................................................. 44 98. Apoptotic Cell Death and Inhibition of Wnt/-Catenin Signaling Pathway in Human Colon Cancer Cells by an Active Fraction (HS7) from Taiwanofungus camphoratus. ................................................. 44 99. Macrophage Mediated Anti-Proliferation Effects of Anthodia camphorata Non-Polysaccharide Based Extracts on Human Hepatoma Cells. ........................................................................................... 45 100. Antcin A, a steroid-like compound from Antrodia camphorata, exerts anti-inflammatory effect via mimicking glucocorticoids. ..................................................................................................................... 45 101. Study of the Anti-Proliferative Activity of 5-Substituted 4,7-Dimethoxy-1,3-Benzodioxole Derivatives of SY-1 from Antrodia camphorata on Human COLO 205 Colon Cancer Cells. ............... 45 102. A Preclinical Evaluation of Antrodia camphorata Alcohol Extracts in the Treatment of Non-Small Cell Lung Cancer Using Non-Invasive Molecular Imaging. .................................................................. 46 103. Antcin A contributs to anti-inflammatory effect of Niuchangchih (Antrodia camphorata). ........... 46 104. Changes in volatile compound composition of Antrodia camphorata during solid state fermentation. ........................................................................................................................................... 46 105. Antroquinonol, a natural ubiquinone derivative, induces a cross talk between apoptosis, autophagy and senescence in human pancreatic carcinoma cells. ............................................................................ 47 106. Optimization of fermentation medium for triterpenoid production from Antrodiacamphorata ATCC 200183 using artificial intelligence-based techniques. ........................................................................... 47 107. Antrodia camphorata Induces Apoptosis and Enhances the Cytotoxic Effect of Paclitaxel in Human Ovarian Cancer Cells. ................................................................................................................ 48 108. Pretreatment with an ethanolic extract of Taiwanofungus camphoratus (Antrodiacamphorata) enhances the cytotoxic effects of Amphotericin B. ................................................................................. 48
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109. Antroquinonol differentially modulates T cells activity, reduces IL-18 production, but enhances Nrf2 activation in accelerated severe lupus nephritis. ............................................................................ 49 110. Antcin B and its Ester Derivative from Antrodia camphorata Induce Apoptosis in Hepatocellular Carcinoma Cells Involves Enhancing Oxidative Stress Coincident with Activation of Intrinsic and Extrinsic Apoptotic Pathway. .................................................................................................................. 49 111. Chemical profiling of the cytotoxic triterpenoid-concentrating fraction and characterization of ergostane stereo-isomer ingredients from Antrodiacamphorata. ............................................................ 50 112. Production of bioactive exopolysaccharides from bitter medicinal mushroom, Antrodia camphorata (M. Zang et C.H. Su) Sh.H. Wu et al. (Aphyllophoromycetideae) in submerged cultivation................................................................................................................................................ 50 113. In vitro and in vivo comparisons of the effects of the fruiting body and mycelium of Antrodia camphorata against amyloid -protein-induced neurotoxicity and memory impairment. ...................... 50 114. In Vivo Antitumor Effects of 4,7-Dimethoxy-5-methyl-1,3-benzodioxole Isolated from the Fruiting Body of Antrodia camphorata through Activation of the p53-Mediated p27/Kip1 Signaling Pathway. .................................................................................................................................................. 51 115. [Analyze on volatile compounds of Antrodia camphorata using HS-SPME-GC-MS]. .................. 51 116. [Medium optimization for mycelia production of Antrodia camphorata based on artificial neural network-genetic algorithm]. .................................................................................................................... 51 117. Antileukemia component, dehydroeburicoic acid from Antrodia camphoratainduces DNA damage and apoptosis in vitro and in vivo models............................................................................................... 52 118. Polysaccharides from extracts of Antrodia camphorata mycelia and fruiting bodies modulate inflammatory mediator expression in mice with polymicrobial sepsis. .................................................. 52 119. Review of Biological and Pharmacological Activities of the Endemic Taiwanese Bitter Medicinal Mushroom, Antrodia camphorata (M. Zang et C. H. Su) Sh. H. Wu et al. (Higher Basidiomycetes). ... 53 120. Taiwanofungus camphoratus (Syn Antrodia camphorata) Extract and Amphotericin B Exert Adjuvant Effects via Mitochondrial Apoptotic Pathway. ....................................................................... 53 121. Anticancer Effects of Eleven Triterpenoids Derived from Antrodia camphorata. .......................... 54 122. Inhibition of Cell Growth and Induction of Apoptosis by Antrodia camphorata in HER-2/neu-Overexpressing Breast Cancer Cells through the Induction of ROS, Depletion of HER-2/neu, and Disruption of the PI3K/Akt Signaling Pathway. .......................................................... 54 123. Triterpenoid-Rich Extract from Antrodia camphorata Improves Physical Fatigue and Exercise Performance in Mice. .............................................................................................................................. 54 124. Inhibition of Na(+)/K(+) -ATPase by Antcins, Unique Steroid-Like Compounds in Antrodia camphorate. ............................................................................................................................................. 55 125. An efficient total synthesis of a potent anti-inflammatory agent, benzocamphorin f, and its anti-inflammatory activity. ...................................................................................................................... 55 126. Taiwanofungus camphorata nitroreductase: cDNA cloning and biochemical characterisation. ..... 55 127. Lanostane triterpenoids and sterols from Antrodia camphorata. .................................................... 56 128. The Antitumor Activity of Antrodia camphorata in Melanoma Cells: Modulation of Wnt/-Catenin Signaling Pathways. ................................................................................................................................ 56
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129. Lanostanoids from Fungi: A Group of Potential Anticancer Compounds. ..................................... 56 130. Antrodia camphorata ATCC 200183 sporulates asexually in submerged culture. .......................... 57 131. Quality of bread supplemented with mushroom mycelia. .............................................................. 57 132. The in vivo antitumor effects on human COLO 205 cancer cells of the 4,7-dimethoxy-5-(2-propen-1-yl)-1,3-benzodioxole (apiole) derivative of 5-substituted 4,7-dimethoxy-5-methyl-l,3-benzodioxole (SY-1) isolated from the fruiting body of Antrodia camphorate. ............................................................................................................................................. 57 133. Inhibition of Helicobacter pylori CagA-Induced Pathogenesis by Methylantcinate B from Antrodia camphorata. ............................................................................................................................................. 58 134. New Anti-Inflammatory Aromatic Components from Antrodia camphorata. ................................ 58

1: Chemical Characterization and Anti-inflammatory Effect of Polysaccharides Fractionated from Submerge-Cultured Antrodia camphorata Mycelia. J Agric Food Chem. 2007 May 31; [Epub ahead of print] Chen CC, Liu YW, Ker YB, Wu YY, Lai EY, Chyau CC, Hseu TH, Peng RY. Institute of Biotechnology, National Tsing Hua University, No. 101, Section 2 Kuang-Fu Road, Hsinchu, Taiwan, Republic of China; Graduate Institute of Biopharmaceutics, Life Science College, National Chiayi University, 300 University Road, Chiayi, Taiwan, Republic of China; and Department of Food and Nutrition and Research Institute of Biotechnology, Hung Kuang University, No. 34 Chung-Chie Road, Shalu County, Taichung Hsien, Taiwan, Republic of China. Five polysaccharide fractions (AC-1, AC-2, AC-3, AC-4, and AC-5) were obtained after systemic solvent extractions and precipitations from Antrodia camphorata mycelia with yields of 2.92, 10.38, 1.65, 0.34, and 1.64%, respectively. Gel permeation chromatography (GPC) analysis showed that the distribution of mean molecular mass of the fractionated polysaccharides was in the range of 394-940 kDa. The proximate compositions from each polysaccharide fraction revealed that all fractions belonged to the category of glycoprotein, having ratios of carbohydrate/protein ranging from 0.29 to 10.79 (w/w). Glucose or galactose was the major monosaccharide in all fractions except fraction AC-2, which has a mean molecular mass of 394 kDa with lyxose as the most prominent constituent. In the evaluation of the DPPH* radical scavenging capability, fraction AC-1 and AC-2 polysaccharides showed the better capabilities, around 74.5 and 50.5%, respectively, compared to the reference control of Trolox (87.5%) at a concentration of 1 muM. In testing with macrophage RAW264.7 cells, fraction AC-2 demonstrated a rather potent anti-inflammatory capability. Furthermore, the lipopolysaccharide-induced NO production and the protein expression by the inducible nitric oxide synthase (iNOS) gene were inhibited, respectively, in a dose-dependent (50-200 mug/mL) manner by fraction AC-2 polysaccharide. Keywords: Antrodia camphorata; mycelia; polysaccharides; DPPH radicals; macrophage; nitric oxide; inducible nitric oxide synthase (iNOS). PMID: 17536816 [PubMed - as supplied by publisher] 2: Unique Formosan Mushroom Antrodia camphorata Differentially Inhibits Androgen-Responsive LNCaP and -Independent PC-3 Prostate Cancer Cells. Nutr Cancer. 2007;57(1):111-21. Chen KC, Peng CC, Peng RY, Su CH, Chiang HS, Yan JH, Hsieh-Li HM. Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University Hospital, Taipei Medical University, Taipei, Taiwan. Antrodia camphorata (AC), a precious and unique folkloric medicinal mushroom enriched in polyphenolics, isoflavonoids, triterpenoids, and polysaccharides, has been diversely used in Formosa (Taiwan) since the 18th century. In this study, prostate cancer (PCa) cell lines PC-3 (androgen independent) and LNCaP (androgen responsive) were treated with AC crude extract (ACCE) at 50-200 mug/mL, respectively, for 48 h. At the minimum effective dose 150 mug/mL, LNCaP showed a G1/S phase arrest with significant apoptosis. Such dose-dependent behavior of LNCaP cells in response to ACCE was confirmed to proceed as Akt --> p53 --> p21 -->CDK4/cyclin D1 --> G1/S-phase arrest --> apoptosis, which involved inhibiting cyclin D1 activity and preventing pRb phosphorylation. In contrast, being without p53, PC-3 cells showed a G2/M-phase arrest mediated through pathway p21 --> cyclin B1/Cdc2 --> G2/M-phase arrest, however, with limited degree of apoptosis, implicating that ACCE is able to differentially inhibit the growth of different PCa cells by modulating different cell cycle signaling pathways. We conclude that this unique Formosan mushroom, A. camphorata, due to its nontoxicity, might be used as a good adjuvant anticancer therapy for prostate cancers despite its androgen-responsive behaviors, which has long been a serious drawback often encountered clinically in hormonal refractory cases treated by antihormonal therapies and chemotherapeutics. PMID: 17516868 [PubMed - in process] 3: Production of a COX-2 inhibitor, 2,4,5-trimethoxybenzaldehyde, with submerged cultured Antrodia camphorata. Lett Appl Microbiol. 2007 Apr;44(4):387-92. Chen CC, Chyau CC, Hseu TH. Institute of Biotechnology, National Tsing Hua University, Hsinchu, Taiwan. gkbioeng@mars.seed.net.tw AIMS: To investigate the active ingredient in fruiting bodies and to produce it with cultured mycelium in Antrodia camphorata (BCRC 35398). METHODS AND RESULTS: The volatile components from the fruiting bodies, the liquid 8

cultured broth of A. camphorata and Cinnamomum kanehirae wood were separately isolated by steam distillation-solvent extraction and identified by gas chromatography-mass spectrometry. In the fruiting bodies, a COX-2 inhibitor 2,4,5-trimethoxybenzaldehyde (TMBA) was found to be the most abundant constituent, but was totally absent in its cultured broth and its natural host, C. kanehirae wood. On feeding with the acid-digested sawdust of C. kanehirae wood or vanillin to the broth for culture, TMBA was produced in both cultured broths. CONCLUSION: The TMBA identified in fruiting bodies was an active ingredient whose functions consisted with the reported experiences of this mushroom. Feeding vanillin to culture broth could produce TMBA containing mycelium product like its fruiting bodies did. SIGNIFICANCE AND IMPACT OF THE STUDY: This study found an active ingredient in fruiting bodies of A. camphorata and elucidated this compound derived from digested sawdust of C. kanehirae wood. A feasible method was also developed to produce TMBA containing mycelium by feeding vanillin. PMID: 17397476 [PubMed - in process] 4: Inhibition of cyclooxygenase-2 and induction of apoptosis in estrogen-nonresponsive breast cancer cells by Antrodia camphorata. Food Chem Toxicol. 2007 Jul;45(7):1107-15. Epub 2006 Dec 27. Hseu YC, Chen SC, Tsai PC, Chen CS, Lu FJ, Chang NW, Yang HL. Department of Cosmeceutics, China Medical University, Taichung, Taiwan. The objective of this study was to investigate the fermented culture broth of Antrodia camphorata (A. camphorata) to induce apoptosis and inhibit cyclooxygenase-2 (COX-2) in estrogen-nonresponsive (MDA-MB-231) human breast cancer cells. Treatment of the highly invasive MDA-MB-231 cells with A. camphorata (40-240mug/ml) resulted in dose and time-dependent sequences of events marked by apoptosis, as evidenced by loss of cell viability, chromatin condensation, and internucleosomal DNA fragmentation. Apoptosis in the MDA-MB-231 cells was accompanied by release of cytochrome c, activation of caspase-3, -8, and -9, and specific proteolytic cleavage of poly (ADP-ribose) polymerase (PARP). Although the A. camphorata-induced apoptosis was associated with a reduction in Bcl-2 protein levels, negligible Bax increase was observed. Furthermore, A. camphorata treatment inhibited COX-2 protein expression and prostaglandin E2 (PGE2) production in MDA-MB-231 cells. Analysis of the study data suggests that A. camphorata exerts growth inhibition on (highly invasive) estrogen-nonresponsive human breast cancer cells through apoptosis induction associated with COX-2 inhibition, and that it may possess anticancer properties potentially valuable for application in drug products. PMID: 17391824 [PubMed - in process] 5: Modulation of inflammation-related genes of polysaccharides fractionated from mycelia of medicinal basidiomycete Antrodia camphorata. Acta Pharmacol Sin. 2007 Feb;28(2):258-67. Wu YY, Chen CC, Chyau CC, Chung SY, Liu YW. Graduate Institute of Biopharmaceutics, College of Life Sciences, National Chiayi University, Chiayi 600, Taiwan. ywlss@mail.ncyu.edu.tw AIM: To investigate the effect of water soluble-ethanol precipitation fraction (AC-1) and alkaline extraction-isoelectric precipitation fraction (AC-2) from Antrodia camphorata (Polyporaceae, Aphyllophorales) on lipopolysaccharide (LPS)-induced gene activation in mouse macrophages. METHODS: The AC-1 and AC-2 fractions were prepared, and their effects on LPS-induced gene expression were monitored by Western blotting and RT-PCR. RESULTS: Our results indicated that AC-2, but not AC-1 dose-dependently (50-200 mg/L) inhibited LPS-induced nitric oxide production as well as the protein and the mRNA expression of the inducible nitric oxide synthase (iNOS) gene. Neither AC-1 nor AC-2 inhibited LPS-induced cyclooxygenase-2 gene expression. Using the cytokine array assay, it showed that AC-2 also had the ability to inhibit LPS-induced the protein expression of interleukin (IL)-6, IL-10, the monocyte chemoattractant protein (MCP)-5, and regulated upon activation, normal T-cell expressed, and presumably secreted (RANTES). Like iNOS, AC-2 inhibiting LPS-induced IL-6 and IL-10 secretion resulted from inhibiting their mRNA expression. CONCLUSION: It was suggested that alkaline extraction-isoelectric precipitated the polysaccharide fraction of A camphorata and had the ability to inhibit LPS-induced iNOS, IL-6, IL-10, MCP-5, and RANTES expression in mouse macrophages. PMID: 17241529 [PubMed - in process]

6: Antihypertensive activities of a solid-state culture of Taiwanofungus camphoratus (Chang-chih) in spontaneously hypertensive rats. Biosci Biotechnol Biochem. 2007 Jan;71(1):23-30. Epub 2007 Jan 7. Liu DZ, Liang YC, Lin SY, Lin YS, Wu WC, Hou WC, Su CH. Graduate Institute of Biomedical Materials and Engineering, Taipei Medical University, Taiwan. Wild and solid-state cultures (SSC) of Taiwanofungus camphoratus (aka Antrodia camphorata and Chang-chih [CC]) were sequentially extracted with cold water, methanol, and hot water to get cold-water-soluble (CWS), methanol-soluble (MS), and hot-water-soluble (HWS) extracts, respectively. Only the MS extract exhibited angiotensin-converting enzyme (ACE) inhibitory activities. The antihypertensive effects of the MS extract (10 mg/kg BW) were measured in spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats. MS extract of the SSC type was able to effectively lower the systolic blood pressure (SBP) and diastolic blood pressure (DBP) of SHR, but not of WKY rats, the results being significantly different from those for distilled water only (the blank). However, wild CC and its MS extract were not as effective as the SSC type in reducing SHR blood pressure and had no effect on WKY rats. SSC-type CC might be developed into a health food with the ability to regulate blood pressure. PMID: 17213674 [PubMed - indexed for MEDLINE] 7: Biochemical characterization of 1-Cys peroxiredoxin from Antrodia camphorata. Appl Microbiol Biotechnol. 2007 Jan;73(6):1314-22. Epub 2006 Nov 14. Wen L, Huang HM, Juang RH, Lin CT. Department of Chemistry, Western Illinois University, 1 University Circle, Macomb, IL, 61455-1390, USA. Antrodia camphorata is a unique medicinal mushroom found only in Taiwan. It has been used as a remedy for various diseases in folk medicine. Antrodia camphorata has been shown to exhibit antioxidative effects. Peroxiredoxins play important roles in antioxidation and cell signaling. A gene encoding an antioxidant enzyme, 1-cysteine peroxiredoxin (1-Cys Prx), was identified in an expressed sequence tag database of the A. camphorata and cloned by polymerase chain reaction. The 1-Cys Prx cDNA (837 bp, accession no. AY870325) contains an open reading frame encoding a protein of 223 amino acid residues with calculated molecular mass of 25,081 Da. The deduced protein shared 44-58% identity with 1-Cys Prx from Homo sapiens, Bos taurus, and Saccharomyces cerevisia. The sequence surrounding the conserved cysteine DFTPVCTTE is conserved. The coding sequence was subcloned into a vector, pET-20b (+), and transformed into Escherichia coli. The recombinant 1-Cys Prx was purified by Ni(2+)-nitrilotriacetic acid (Sepharose). The purified enzyme was characterized under various conditions. The enzyme is thermostable because its half-life of inactivation was 15.5 min at 60 degrees C. It was stable under alkaline pH range from 7.8 to 10.2. The enzyme showed decreased activity with increasing concentration of imidazole. The enzyme is sensitive to trypsin and chymotrypsin treatment. PMID: 17103164 [PubMed - indexed for MEDLINE] 8: Protective effects of mycelia of Antrodia camphorata and Armillariella tabescens in submerged culture against ethanol-induced hepatic toxicity in rats. J Ethnopharmacol. 2007 Mar 1;110(1):160-4. Epub 2006 Oct 4. Lu ZM, Tao WY, Zou XL, Fu HZ, Ao ZH. The Key Laboratory of Industrial Biotechnology, Ministry of Education, Southern Yangtze University, Wuxi 214036, PR China. The hepatoprotective effects of the mycelia of Antrodia camphorata and Armillariella tabescens were evaluated in vivo using acute ethanol-intoxicated rats as an experimental model. Animals were orally treated with Antrodia camphorata (0.5 or 1.0 g/kg b.w.) or Armillariella tabescens (0.5 or 1.0 g/kg b.w.) for 10 days whereas controls received vehicle only. At the end of the experimental 10-day period, the animals were administered by gavage with an acute ethanol dose of 5.0 g/kg b.w. diluted in deionized water (6:4, v/v) and sacrificed at 18 h after ethanol administration. The degree of protection was measured by using biochemical parameters like serum transaminases (AST and ALT), alkaline phosphatase (ALP), bilirubin. Meanwhile, the histopathological studies were carried out to support the above parameters. Administration of Antrodia camphorata or Armillariella tabescens markedly prevented ethanol-induced elevation of levels of serum AST, ALT, ALP, and bilirubin comparable with standard drug silymarin. PMID: 17092673 [PubMed - in process]

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9: Phylogenetic analysis of Antrodia species and Antrodia camphorata inferred from internal transcribed spacer region. Antonie Van Leeuwenhoek. 2007 Apr;91(3):267-76. Epub 2006 Oct 28. Chiu HH. Department of Biotechnology, Fooyin University, 151 Chin-Hsueh Rd., Ta-Liao Hsiang, 831 Kaohsiung Hsien, Taiwan ROC. sc055@mail.fy.edu.tw The species of Antrodia are one of the difficult-to-classify and obscure groups of poroid Aphyllophorales based on morphological appearance. However, it is becoming increasingly important to reliably identify the entire suite of Antrodia camphorata strains and Antrodia species due to the potential pharmaceutical value of their biologically active ingredients. In this study, the internal transcribed spacer (ITS) region of the ribosomal RNA gene (rDNA) was sequenced and phylogenetically analyzed in a number of Antrodia fungal species and strains. ITS amplicons from the Antrodia species tested ranged in size from 543 to 610 bp; the size of the ITS of A. camphorata strains ranged from 592 to 596 bp. The overall sizes of ITS2 and 5.8S ribosomal RNA gene of all A. camphorata strains tested in this study were shown to be 217 and 158 bp, respectively. A phylogenetic analysis of ITS data generated, which included sequences of 11 A. camphorata strains and nine other Antrodia species, showed three clearly distinct groups. Group 1 includes A. camphorata, Antrodia salmonea, and Antrodia carbinca strains. Within Group 2, Antrodia sinuosa and Antrodia xantha were clustered together. Group 3 contained Antrodia albida, A. heteromorpha, A. serialis, and A. malicola. The observed sequence diversity among ITS alleles provided an effective tool for differentiating strains of A. camphorata, A. salmonea, A. xantha, A. sinuosa, or A. serialis. Polymorphisms arising within the ITS1-5.8S-ITS2 region can provide practical markers for establishing a foundation for the further expansion of an ITS sequence database of medically important fungi. PMID: 17072535 [PubMed - in process] 10: Biochemical characterization of a novel 2-Cys peroxiredoxin from Antrodia camphorata. Appl Microbiol Biotechnol. 2007 Feb;74(1):84-92. Epub 2006 Oct 10. Huang JK, Ken CF, Huang HM, Lin CT. Department of Chemistry, Western Illinois University, 1 University Circle, Macomb, IL 61455-1390, USA. Peroxiredoxins (Prxs) play important roles in antioxidation and cell signaling. A gene encoding a novel 2-Cys Prx was identified based on sequence homology in an expressed sequence tag database of the Antrodia camphorata, a medicinal mushroom found only in Taiwan. The 2-Cys Prx cDNA (940 bp) encodes a protein of 188 amino acid residues with calculated molecular mass of 20,965 Da and a pI of 5.89. The coding region was subcloned into pAVD10, transformed into Escherichia coli, and expressed as a His-tagged fusion protein. The purified enzyme was characterized under various conditions. The Prx retained 68% activity after being heated at 60 degrees C for 2 min. It was stable under a broad pH range from 5 to 11. The enzyme activity was slightly decreased in the presence of 1% sodium dodecyl sulfate. The enzyme was somewhat susceptible to chymotrypsin treatment but resistant to digestion by trypsin. PMID: 17031636 [PubMed - indexed for MEDLINE] 11: Antrodia camphorata extract induces replicative senescence in superficial TCC, and inhibits the absolute migration capability in invasive bladder carcinoma cells. J Ethnopharmacol. 2007 Jan 3;109(1):93-103. Epub 2006 Jul 11. Peng CC, Chen KC, Peng RY, Chyau CC, Su CH, Hsieh-Li HM. Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, 250 Wu-Xing Street, Taipei 110, Taiwan. The Antrodia camphorata crude extract (ACCE), an extract obtained from a precious traditional Chinese folkloric herbal medicine Zhan-Ku (a camphor tree mushroom) since the 18th century, has showed rather significant inhibitory effects on the growth and proliferation of the transitional cell carcinomas (TCC) cell lines RT4, TSGH-8301, and T24. On treatment with ACCE at 100 microg/mL, the p53-independent overexpression of p21 with simultaneous down alteration of pRb was observed in RT4, which was thus speculative of proceeding through a mechanism of replicative senescence. On the contrary treatment with ACCE, at 50 microg/mL, resulting in simultaneous down-regulations of Cdc2 and Cyclin B1, with suppression of the absolute migrating capability of the two cell lines TSGH-8301 and T24, and eventually the cell deaths. We conclude that ACCE can be rather effective and beneficial in suppression of both the superficial cancer cell line RT4 and the metastatic cell lines (TSGH-8301 and T24) through different mechanisms. 11

PMID: 16930895 [PubMed - indexed for MEDLINE] 12: Filtrate of fermented mycelia from Antrodia camphorata reduces liver fibrosis induced by carbon tetrachloride in rats. World J Gastroenterol. 2006 Apr 21;12(15):2369-74. Lin WC, Kuo SC, Lin WL, Fang HL, Wang BC. Department of Pharmacology, China Medical University, Taichung 404, Taiwan, China. wclin@mail.cmu.edu.tw AIM: To investigate the effects of filtrate of fermented mycelia from Antrodia camphorata (FMAC) on liver fibrosis induced by carbon tetrachloride (CCl(4)) in rats. METHODS: Forty Wistar rats were divided randomly into control group and model group. All model rats were given 200 mL/L CCl(4) (2 mL/Kg, po) twice a week for 8 wk. Four weeks after CCl(4) treatment, thirty model rats were further divided randomly into 3 subgroups: CCl(4) and two FMAC subgroups. Rats in CCl(4) and 2 FMAC subgroups were treated with FMAC 0, 0.5 and 1.0 g/kg, daily via gastrogavage beginning at the fifth week and the end of the eighth week. Spleen weight, blood synthetic markers (albumin and prothrombin time) and hepatic malondialdehyde (MDA) and hydroxyproline (HP) concentrations were determined. Expression of collagen I, tissue inhibitor of metalloproteinases (TIMP)-1 and transforming growth factor beta1 (TGF-beta1) mRNA were detected by RT-PCR. Histochemical staining of Masson's trichrome was performed. RESULTS: CCl(4) caused liver fibrosis, featuring increased prothrombin time, hepatic MDA and HP contents, and spleen weight and decreased plasma albumin level. Compared with CCl(4) subgroup, FMAC subgroup (1 g/kg) significantly decreased the prothrombin time (36.7+/-7.2 and 25.1+/-10.2 in CCl(4) and FMAC groups, respectively, P<0.05) and increased plasma albumin concentration (22.7+/-1.0 and 30.7+/-2.5 in CCl(4) and FMAC groups, respectively, P<0.05). Spleen weight was significantly lower in rats treated with CCl(4) and FMAC (1 g/kg) compared to CCl(4) treated rats only (2.7+/-0.1 and 2.4+/-0.2 in CCl(4) and FMAC groups, respectively, P<0.05). The amounts of hepatic MDA and HP in CCl(4)+FAMC (1 g/kg) subgroup were also lower than those in CCl(4) subgroup (MDA: 3.9+/-0.1 and 2.4+/-0.6 in CCl(4) and CCl(4)+FMAC groups, respectively, P<0.01; HP: 1730.7+/-258.0 and 1311.5+/-238.8 in CCl(4) and CCl(4)+FMAC groups, respectively, P<0.01). Histologic examinations showed that CCl(4)+FMAC subgroups had thinner or less fibrotic septa than CCl(4) group. RT-PCR analysis indicated that FMAC (1 g/kg) reduced mRNA levels of collagen I, TIMP-1 and TGF-beta1 (collagen I: 5.63+/-2.08 and 1.78+/-0.48 in CCl(4) and CCl(4)+FMAC groups, respectively, P<0.01; TIMP-1: 1.70+/-0.82 and 0.34+/-0.02 in CCl(4) and CCl(4)+FMAC groups, respectively, P<0.01; TGF-beta1:38.03+/-11.9 and 4.26+/-2.17 in CCl(4) and CCl(4)+FMAC groups, respectively, P<0.01) in the CCl(4)-treated liver. CONCLUSION: It demonstrates that FMAC can retard the progression of liver fibrosis induced by CCl(4) in rats. PMID: 16688827 [PubMed - indexed for MEDLINE] 13: Neuroprotective diterpenes from the fruiting body of Antrodia camphorata. J Nat Prod. 2006 Apr;69(4):689-91. Chen CC, Shiao YJ, Lin RD, Shao YY, Lai MN, Lin CC, Ng LT, Kuo YH. Department of Chemistry, National Taiwan University, Taipei, Taiwan 106. Three new compounds, 19-hydroxylabda-8(17)-en-16,15-olide (1), 3beta,19-dihydroxylabda-8(17),11E-dien-16,15-olide (2), and 13-epi-3beta,19-dihydroxylabda-8(17),11E-dien-16,15-olide (3), together with four known compounds, 19-hydroxylabda-8(17),13-dien-16,15-olide (4), 14-deoxy-11,12-didehydroandrographolide (5), 14-deoxyandrographolide, and pinusolidic acid, were isolated from the fruiting bodies of Antrodia camphorata. The structures of compounds 1-3 were elucidated by the analysis of their spectroscopic data. The in vitro neuroprotective activity of all compounds was evaluated, and compounds 1-5 protected neurons from Abeta damage by 39.2, 35.0, 36.7, 30.6, and 27.0%, respectively, at concentrations between 5 and 20 microM. PMID: 16643055 [PubMed - indexed for MEDLINE] 14: Antrodia camphorata in submerged culture protects low density lipoproteins against oxidative modification. Am J Chin Med. 2006;34(2):217-31. Yang HL, Hseu YC, Chen JY, Yech YJ, Lu FJ, Wang HH, Lin PS, Wang BC. Institute of Nutrition, China Medical University, Taichung, Taiwan. Antrodia camphorata is well known in Taiwan as a traditional Chinese medicine. In this study, we have investigated the 12

antioxidant properties of a fermented culture broth of Antrodia camphorata (FCBA) and the aqueous extracts of mycelia from Antrodia camphorata (AEMA) on the oxidative modification of human low-density lipoproteins (LDL), as induced by either copper sulfate (CuSO(4)) or 2,2'-azo-bis(2-amidinopropane) hydrochloride (AAPH). Under such oxidant stress, FCBA and AEMA appear to possess antioxidant properties with respect to oxidation of LDL in a time-and concentration-dependent manner, as assessed by inhibition of thiobarbituric acid-reactive substances (TBARS) formation, conjugated diene production, and cholesterol degradation of oxidized LDL. In addition, both FCBA and AEMA exhibited a remarkable ability to rescue the relative electrophoretic mobility and fragmentation of the Apo B moiety of the oxidized LDL. Furthermore, FCBA and AEMA effectively protected the endothelial cells from the damaging effects of the CuSO(4)-oxidized LDL. Our findings suggest that the antioxidant properties of Antrodia camphorata may also provide effective protection from atherosclerosis. PMID: 16552834 [PubMed - indexed for MEDLINE] 15: Human urinary bladder cancer T24 cells are susceptible to the Antrodia camphorata extracts. Cancer Lett. 2006 Nov 8;243(1):109-19. Epub 2006 Feb 7. Peng CC, Chen KC, Peng RY, Su CH, Hsieh-Li HM. Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan, ROC. Bladder cancer has been cited to result from the neoplastic lesion with environmental and/or occupational factors identified as causatives. Transitional cell carcinoma (TCC) is the most common type of bladder cancer. Most of the bladder cancer patients die from the invasive, metastatic TCC that has turned out to be resistant to chemotherapy. T24 cells, a cell line established from a human urinary bladder cancer patient, are high-grade and invasive TCC. T24 cells were found very susceptible to ACCE at concentration of 50 microg/mL. MTT assay showed that the cell growth and proliferation were inhibited to 50% of the control when treated with ACCE for 72 h, at which the cell proliferation suppressing rate revealed -4.4 x 10(3)cells/microg per day. Comparing the expressions of the cell cycle biomarkers Cdc2 and Cyclin B1 by the western blot analysis, a phase G(2)M arrest was confirmed. Both the wound scratch assay and the transwell motility assay indicated that ACCE was very effective anti-metastatic against T24 cells. Furthermore, the active form of matrix metalloproteinase-9 (MMP-9) was also found totally suppressed as revealed by zymography at 72 h post-incubation with ACCE, while the light and electron microscopic images have apparently revealed cell membrane damages on T24 cells when treated with ACCE (50 microg/mL). Moreover, both the wound scratch and the transwell assays have demonstrated the migration capability of T24 cells has been significantly retarded to 1.5-fold at same dosage of ACCE used. In conclusion, ACCE is a good anti-cancer agent, being effective in inducing phase G(2)M arrest, acting as an anti-proliferative, and an anti-metastatic agent against bladder cancer cell T24 cells. PMID: 16455193 [PubMed - indexed for MEDLINE] 16: Adenosine as an active component of Antrodia cinnamomea that prevents rat PC12 cells from serum deprivation-induced apoptosis through the activation of adenosine A(2A) receptors. Life Sci. 2006 Jun 13;79(3):252-8. Epub 2006 Jan 27. Lu MK, Cheng JJ, Lai WL, Lin YR, Huang NK. National Research Institute of Chinese Medicine, Taipei, Taiwan, No 155-1, Section 2, Li-Nung Street., Shipai, Pei-tou District (112), Taipei, Taiwan, ROC. Antrodia cinnamomea (formerly named Antrodia camphorata) is a rare medicinal fungus. We previously reported that it exhibits antioxidative, vasorelaxative, anti-inflammatory, and anti-angiogenic effects. When serum deprivation-induced apoptosis in neuronal-like PC12 cells was used as a stress model, the extract of A. cinnamomea displayed effectiveness in preventing serum-deprived apoptosis. Since our previous data show that the extract of A. cinnamomea contains adenosine (ADO), we attempt to investigate if the active component is ADO and to identify its targeting site in this study. After pre-incubation with ADO deaminase, neither ADO nor the extract of A. cinnamomea exerted any protection, demonstrating that the active component of A. cinnamomea is ADO. Furthermore, an ADO A(2A) receptor (A(2A)-R) antagonist was used and was able to block the protective effects of ADO and the extract of A. cinnamomea, demonstrating that the ADO targeting site in this model is A(2A)-R. Taken together, the protective effect of A. cinnamomea is owed to its active component, ADO, which acts through activation of A(2A)-R to prevent serum deprivation-induced PC12 cell apoptosis. PMID: 16443241 [PubMed - indexed for MEDLINE]

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17: Proteomic analysis of the effect of Antrodia camphorata extract on human lung cancer A549 cell. Proteomics. 2006 Feb;6(3):826-35. Wu H, Pan CL, Yao YC, Chang SS, Li SL, Wu TF. Department of Biotechnology, Southern Taiwan University of Technology, Tainan, Taiwan. Antrodia camphorata (niu-chang-chih) is a fungus native to Taiwan that is believed to be effective in preventing diseases. This study demonstrates that 0.2-2% v/v ethanol extracts of A. camphorata cultivated by solid-state fermentation (SACE) can effectively impede the proliferation of human non-small cell lung carcinoma A549 cells but not primary human fetal lung fibroblast MRC-5. The results of apoptotic analyses implicate that SACE might trigger the apoptosis in the A549 cells by inducing endoplasmic reticulum stress. Two-dimensional gel maps of non-treated and treated A549 cells were compared using PDQUEST analytical software to discover five statistically significant twofold or above-twofold differentially-expressed protein spots. The five protein spots that were significantly de-regulated were chosen for subsequent identification by high performance liquid chromatography electro-spray tandem mass spectrometry. The five proteins were later identified as human galectin-1, human eukaryotic translation initiation factor 5A, human Rho GDP dissociation inhibitor alpha, human calcium-dependent protease small subunit and human annexin V. All five proteins were confirmed to be down-regulated by Western blotting. The analytical results of this study help to provide insight into the effect of SACE on the gene expression of the tumor cells. PMID: 16411266 [PubMed - indexed for MEDLINE] 18: Growth inhibition and induction of apoptosis in MCF-7 breast cancer cells by Antrodia camphorata. Cancer Lett. 2006 Jan 18;231(2):215-27. Yang HL, Chen CS, Chang WH, Lu FJ, Lai YC, Chen CC, Hseu TH, Kuo CT, Hseu YC. Institute of Nutrition, China Medical University, Taichung, Taiwan, ROC. Antrodia camphorata (A. camphorata) is well known in Taiwan as a traditional Chinese medicine, and it has been shown to exhibit antioxidant and anticancer effects. In this study, therefore, its ability to induce apoptosis in cultured MCF-7 breast cancer cells was studied. Treatment of the MCF-7 cells with a variety of concentrations of the fermented culture broth of A. camphorata (25-150 microg/ml) resulted in dose- and time-dependent sequences of events marked by apoptosis, as shown by loss of cell viability, chromatin condensation, internucleosomal DNA fragmentation, and sub-G1 phase accumulation. Furthermore, apoptosis in the MCF-7 cells was accompanied by the release of cytochrome c, activation of caspase 3, and specific proteolytic cleavage of poly (ADP-ribose) polymerase (PARP). Although, the A. camphorata-induced apoptosis was associated with Bax protein levels, negligible Bcl-2 reduction was observed. Interestingly, A. camphorata induced dose-dependent reactive oxygen species (ROS) generation in MCF-7 cells. Analysis of the data suggests that A. camphorata exerts antiproliferative action and growth inhibition on MCF-7 cells through apoptosis induction, and that it may have anticancer properties valuable for application in drug products. PMID: 16399223 [PubMed - indexed for MEDLINE] 19: Anti-inflammatory potential of Antrodia Camphorata through inhibition of iNOS, COX-2 and cytokines via the NF-kappaB pathway. Int Immunopharmacol. 2005 Dec;5(13-14):1914-25. Epub 2005 Jul 18. Hseu YC, Wu FY, Wu JJ, Chen JY, Chang WH, Lu FJ, Lai YC, Yang HL. Department of Food Science, Changtai Institute of Health Sciences and Technology, Taiwan. Antrodia camphorata (A. camphorata), well known in Taiwan as a traditional Chinese medicine, has been shown to exhibit antioxidant and anticancer effects. In the present study, therefore, we have examined the effects of the fermented culture broth of A. camphorata (25-100 microg/ml) in terms of lipopolysaccharide (LPS)-induced nitric oxide (NO) and prostaglandin E2 (PGE2) production, and inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression in RAW 264.7 macrophages. Our results indicate concentration-dependent A. camphorata inhibition of LPS-induced NO and PGE2 production, without appreciable cytotoxicity on the RAW 264.7 cells. A. camphorata also attenuates the production of LPS-induced tumor necrosis factor (TNF-alpha) and interleukin (IL)-1beta. Furthermore, A. camphorata blocks the IkappaB-alpha degradation induced by LPS. These results indicate that A. camphorata inhibits LPS induction of cytokine, iNOS and COX-2 expression by blocking NF-kappaB activation. Therefore, we report the first confirmation of the anti-inflammatory potential of this traditionally employed herbal medicine in vitro. PMID: 16275626 [PubMed - indexed for MEDLINE] 14

20: Promotion of hyphal growth and underlying chemical changes in Antrodia camphorata by host factors from Cinnamomum camphora. Int J Food Microbiol. 2006 Jan 15;106(1):32-8. Epub 2005 Oct 10. Hsu FL, Chou CJ, Chang YC, Chang TT, Lu MK. Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan, ROC. The aim of this research was to investigate the hyphal growth-promoting factors (HGFs) of Antrodia camphorata from the host-related species, Cinnamomum camphora (CC) and the underlying chemical produced. The HGF was identified in the polysaccharide fraction of CC at levels ranging from 80 to 320 mg L(-1), and it maximally stimulated growth to 5.50 g L(-1) during a 14-day culture period compared to that of the control of 2.88 g L(-1). We also investigated the nature and chemical composition of the CC polysaccharide. Herein, size-exclusion column chromatography followed by high-performance anion-exchange chromatography after complete hydrolysis of the CC polysaccharide was performed to derive its molecular weight and sugar composition. The Mw values of the CC polysaccharide were determined to be 728.2, 187.5, 28.7, 7.5, and 1.9 kDa. Compositional analysis of the CC polysaccharide showed that galactosamine, mannose, and glucose were the major monosaccharides. Time-course studies of mycelial extracts of cultures revealed that prolonged incubation with the water-soluble extracts of CC resulted in an increase in the relative amounts of two lanostane-type compounds, i.e., dehydrosulphurenic acid and 15alpha-acetyl-dehydrosulphurenic acid, which are found in the fruiting bodies of A. camphorata. This finding offers the possibility of the reliable production of this medicinal fungus under laboratory conditions compared to its limited slow growth in nature. PMID: 16219379 [PubMed - indexed for MEDLINE] 21: Mycelia from Antrodia camphorata in Submerged culture induce apoptosis of human hepatoma HepG2 cells possibly through regulation of Fas pathway. J Agric Food Chem. 2005 Jul 13;53(14):5559-64. Song TY, Hsu SL, Yeh CT, Yen GC. Department of Food Science, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan. The objective of this study was to investigate the antiproliferative effect and the mechanism of the methanol extracts of mycelia (MEM) form Antrodia camphorata in submerged culture toward HepG2 cells. The results showed that MEM-induced cell apoptosis involved up-regulation of Fas and down-regulation of Bcl-2, DR3, DR4, TNFRI, and TNFRII in HepG2 cells, while no changes on the levels of Bax, Bid, Bad, and Bak protein were observed. On the basis of these results, the involvement of the Fas/Fas ligand (FasL) death-receptor pathway, in MEM-induced apoptosis in HepG2 cells, was investigated. The apoptosis inducing activity was significantly enhanced by a Fas activator and inhibited by a Fas antagonist. To know about the effect of MEM on the activation of the apoptotic pathway, the adenovirus transfected with Bcl-2 was infected on HepG2 cells. The data showed that the percentage of apoptotic cells induced by MEM in Bcl-2-infected HepG2 (Bcl-2 overexpression) was not significantly different from that of uninfected HepG2. These results demonstrate that MEM induces HepG2 apoptosis through inhibition of cell growth and up-regulation of Fas/FasL to activate the pathway of caspase-3 and -8 cascades. PMID: 15998114 [PubMed - indexed for MEDLINE] 22: Induction of apoptosis in human hepatoma cells by mycelia of Antrodia camphorata in submerged culture. J Ethnopharmacol. 2005 Aug 22;100(1-2):158-67. Song TY, Hsu SL, Yen GC. Department of Food Science, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan. The effect of methanolic extracts of mycelia (MEM) from Antrodia camphorata (Polyporaceac, Aphyllophorales) of submerged culture (ACSC) on the inhibition of cell viability and the mechanism of MEM-induced cytotoxic in hepatoma cells were investigated. The IC(50) of MEM on the cytotoxicity of HepG2 (wild type p53) and Hep3B (delete p53) were 49.5 and 62.7 microg/ml, respectively, on 48 h incubation. There is no observable cytotoxicity of MEM in Chang liver cells and rat primary hepatocytes at the concentration of 100 microg/ml. Cell cycle analysis revealed that MEM induced apoptosis on HepG2 via G0/G1 cell cycle arrest. MEM (100 microg/ml) treated HepG2 and Hep3B for 72 h, the apoptotic cells were 98.3 and 39.5%, respectively. The activities of caspase-3, -8 and -9 in HepG2 induced by MEM (50 microg/ml) were increased 5.3, 6.7 and 2.2-fold, respectively. MEM-induced apoptotic cell death was accompanied by up-regulation of 15

caspase-3 and -8 in HepG2 cells. Combined treatment with MEM and caspase-3, -8 and -9 inhibitors, the caspase-3 and -8 inhibitors were accounting for 63 and 47% inhibition in MEM-induced apoptosis, respectively; however, caspase-9 inhibitor exhibited no obvious inhibition effect on the apoptosis percentage (p>0.05). The results indicated that MEM induced HepG2 apoptosis through activation of caspase-3 and -8 cascades and regulation of the cell cycle progression to inhibit hepatoma cells proliferation. PMID: 15949907 [PubMed - indexed for MEDLINE] 23: Apoptotic effects of extract from Antrodia camphorata fruiting bodies in human hepatocellular carcinoma cell lines. Cancer Lett. 2005 Apr 18;221(1):77-89. Hsu YL, Kuo YC, Kuo PL, Ng LT, Kuo YH, Lin CC. Graduate Institute of Natural Products, College of Pharmacy, Kaohsiung Medical University, No. 100 Shin-Chuan 1st Road, Kaohsiung 807, Taiwan, ROC. The fruiting body of Antrodia camphorata is well known in Taiwan as a traditional medicine for treating cancer and inflammation. The purpose of this study was to evaluate the apoptotic effects of ethylacetate extract from A. camphorata (EAC) fruiting bodies in two human liver cancer cell lines, Hep G2 and PLC/PRF/5. Treatment with EAC decreased the cell growth of Hep G2 and PLC/PRF/5 cells in a dose dependent manner. In Fas/APO-1 positive-Hep G2 cells, EAC increased the expression level of Fas/APO-1 and its two forms of ligands, membrane-bound Fas ligand (mFasL) and soluble Fas ligand (sFasL), in a p53-indenpendent manner. In addition, EAC also initiated mitochondrial apoptotic pathway through regulation of Bcl-2 family proteins expression, release of cytochrome c, and activation of caspase-9 both in Hep G2 and PLC/PRF/5 cells. Furthermore, EAC also inhibited the cell survival signaling by enhancing the amount of IkappaBalpha in cytoplasm and reducing the level and activity of NF-kappaB in the nucleus, and subsequently attenuated the expression of Bcl-X(L) in Hep G2 and PLC/PRF/5 cells. EAC therefore decreased the cell growth and induced apoptosis both in Hep G2 and PLC/PRF/5 cells. PMID: 15797630 [PubMed - indexed for MEDLINE] 24: Antrodia camphorata prevents rat pheochromocytoma cells from serum deprivation-induced apoptosis. FEMS Microbiol Lett. 2005 Mar 1;244(1):213-9. Huang NK, Cheng JJ, Lai WL, Lu MK. National Research Institute of Chinese Medicine, No. 155-1, Li-Nung St., Sec. 2, Shih-Pai, Peitou, Taipei 112, Taiwan. Antrodia camphorata (A. camphorata) is a rare medicinal fungus with antioxidative, vasorelaxtative, anti-inflammatory and anti-hepatitive effects. However, the neuroprotective effect has not been studied. By using serum deprivation-induced apoptosis in neuronal-like PC12 cells as a cell stress model, we found that A. camphorata is effective in preventing serum-deprived apoptosis. Inhibitors of both a serine/threonine kinase and a specific protein kinase A (PKA) inhibited the protective effect of A. camphorata, indicating that A. camphorata prevents serum-deprived PC12 cell apoptosis through a PKA-dependent mechanism. A transcription inhibitor, actinomycin D, and a protein synthesis inhibitor, cyclohexamide, both attenuated the protective effect of A. camphorata, indicating a requirement for gene expression for protection by A. camphorata. On the other hand, A. camphorata also increased phosphorylated CREB, a transcription factor, which is H-89-inhibitable in this study, suggesting the possibility that A. camphorata prevents serum deprivation-induced PC12 cell apoptosis through a PKA/CREB-dependent pathway. PMID: 15727843 [PubMed - indexed for MEDLINE] 25: Characterization and functional study of Antrodia camphorata lipopolysaccharide. J Agric Food Chem. 2005 Jan 26;53(2):469-74. Cheng JJ, Yang CJ, Cheng CH, Wang YT, Huang NK, Lu MK. National Research Institute of Chinese Medicine, No. 155-1, Section 2, Li-Nung Street, Pei-tou District (112), Taipei, Taiwan. Lipopolysaccharide (LPS) is a highly proinflammatory molecule isolated from bacteria. This study demonstrated the existence of LPS in a medicinal fungus, Antrodia camphorata. Because no LPS had been identified in any fungus organism, the purification of LPS from A. camphorata was attempted. LPSs from six strains of A. camphorata (35396, 35398, 35716, B71, B85, and B86) were isolated. Chemical and functional properties were investigated on the fungus LPS. Compositional 16

analysis revealed that sorbitol, fucose, galactose, and glucose were the neutral sugars in LPS of A. camphorata. Galactosamine, glucosamine, galactose, and glucose were the predominant monosaccharide species in E. coli O129 LPS molecules, whereas galactosamine and glucosamine were absent in A. camphorata LPS. Because these properties are different from those of bacterial LPS, the functions between fungus and bacterial LPS are also discussed. The vascular endothelial lining of blood vessels, which controls leucocyte traffic and activation, may be one of the primary targets of LPS action during sepsis. Assays for biological activity were performed on endothelial cells with anti-inflammatory effects associated with sepsis. A. camphorata LPS apparently showed a lesser extent of cytotoxicity than bacterial LPS. In contrary to the proinflammatory property of bacterial LPS, LPS from A. camphorata differentially reversed bacterial LPS-induced intercellular adhersion molecule-1 and monocyte adhesion; both were indicators during inflammatory process. In conclusion, basic chemical properties categorized A. camphorata extracts into lipopolysaccharide. However, the detailed functional structures and bioactivities of A. camphorata LPS were totally different from those of bacterial LPS. The investigation of the existence and anti-inflammatory effect of fungus LPS is at present a truly novel and important finding. These results show that LPS isolated from A. camphorata offers a novel therapeutic target for anti-inflammation against E. coli infection. PMID: 15656690 [PubMed - indexed for MEDLINE] 26: Antitumor effects of the partially purified polysaccharides from Antrodia camphorata and the mechanism of its action. Toxicol Appl Pharmacol. 2004 Dec 1;201(2):186-93. Liu JJ, Huang TS, Hsu ML, Chen CC, Lin WS, Lu FJ, Chang WH. Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei, Taiwan. Antrodia camphorata is a popular folk medicine that has attracted great attention due to its fame for antitumor activity against cancer. However, there is little information available about its action. In the present study, we purified a unique polysaccharide component from A. camphorata mycelia (AC-PS) and found that it has pronounced anti-tumor effects on both in vitro and in vivo model. Our results showed that AC-PS alone did not show any direct cytotoxic effect to human leukemic U937 cells, even at high concentration (200 microg/ml). However, it could inhibit the proliferation of U937 cells via activation of mononuclear cells (MNCs). Treatment of U937 cells with AC-PS-stimulated-MNC-CM could significantly inhibit its proliferation with 55.3% growth inhibition rate. The in vitro antitumor activity was substantiated by the in vivo therapeutical study of AC-PS in sarcoma 180-bearing mice. Intraperitoneal and oral administration of AC-PS, 100 and 200 mg/kg significantly suppressed the tumor growth with the inhibition rate of 69.1% and 58.8%, respectively. In vivo studies also showed that several immunoparameters, such as the spontaneous proliferation of spleen cells, after AC-PS administration, were two-fold higher than in control mice. Furthermore, the cytolytic activity of spleen cells also increased from 9.8 +/- 1.1% in control mice to 34.2 +/- 5.5% and 48.2 +/- 2.5%, after oral and intraperitoneal treatment, respectively. Besides, the mice serum interleukin-12 levels increased significantly by AC-PS treatment. Considering all these results, it is suggested that AC-PS elicit its anti-tumor effect by promoting a Th1-dominant state and killer activities. PMID: 15541758 [PubMed - indexed for MEDLINE] 27: Induction of apoptosis by Antrodia camphorata in human premyelocytic leukemia HL-60 cells. Nutr Cancer. 2004;48(2):189-97. Hseu YC, Yang HL, Lai YC, Lin JG, Chen GW, Chang YH. Department of Food Science, Chungtai Institute of Health Sciences and Technology, Taichung, Taiwan. Antrodia camphorata (A. camphorata) is well known in Taiwan as a traditional Chinese medicine, and it has been shown to exhibit antioxidant effects. In this study, the ability of A. camphorata to induce apoptosis was studied in cultured human premyelocytic leukemia HL-60 cells. Treatment of the HL-60 cells with a variety of concentrations of the fermented culture broth of A. camphorata (25-150 microg/ml) resulted in dose- and time-dependent sequences of events marked by apoptosis, as shown by loss of cell viability, chromatin condensation, and internucleosomal DNA fragmentation. Furthermore, apoptosis in the HL-60 cells was accompanied by the release of cytochrome c, activation of caspase-3, and specific proteolytic cleavage of poly (ADP-ribose) polymerase (PARP). This increase in A. camphorata-induced apoptosis was also associated with a reduction in the levels of Bcl-2, a potent cell-death inhibitor, and an increase in those of the Bax protein, which heterodimerizes with and thereby inhibits Bcl-2. The data suggest that A. camphorata exerts antiproliferative action and growth inhibition on HL-60 cells through apoptosis induction and that it may have anticancer properties valuable for 17

application in drug products. PMID: 15231454 [PubMed - indexed for MEDLINE] 28: Evaluation of the anti-inflammatory activity of zhankuic acids isolated from the fruiting bodies of Antrodia camphorata. Planta Med. 2004 Apr;70(4):310-4. Shen YC, Wang YH, Chou YC, Chen CF, Lin LC, Chang TT, Tien JH, Chou CJ. National Research Institute of Chinese Medicine, Taipei, Taiwan. We have previously shown that a concentrated ethanol extract of the fruiting bodies of Antrodia camphorata exhibited immunomodulating effects in human leukocytes and fourteen compounds including zhankuic acids A, B, C, and antcin K were identified in the extract. In this study, an acute cellular model in isolated peripheral human neutrophils was established to elucidate the anti-inflammatory effects of these compounds. Reactive oxygen species (ROS) production and firm adhesion by neutrophils display two important responses during inflammation. To evaluate whether these compounds could prevent inflammatory responses by neutrophils, their effects on N-formyl-methionyl-leucyl-phenylalanine (fMLP) or phorbol 12-myristate-13-acetate (PMA)-activated peripheral human neutrophils were examined. Pretreatment with 1 - 25 microM of zhankuic acids A, B, C, or antcin K concentration-dependently diminished fMLP- or PMA-induced ROS production, as measured by a lucigenin-amplified chemiluminescence, with IC (50) (microM) around 5 - 20 microM. Zhankuic acids A, B, C, or antcin K also effectively inhibited the fMLP- or PMA-induced firm adhesion without interfering with the up-expression of surface Mac-1 (CD11b/CD18), a beta2 integrin mediating the firm adhesion of neutrophils to endothelium. The anti-inflammatory actions of these drugs were not due to cytotoxic effects because no significant difference in cell viability was observed compared to vehicle control. These data suggest that inhibition of both ROS production and firm adhesion by neutrophils has no significant cytotoxic effect that could give these drugs the potential to be anti-inflammatory agents for the clinical treatment. PMID: 15095145 [PubMed - indexed for MEDLINE] 29: Anti-inflammatory activity of the extracts from mycelia of Antrodia camphorata cultured with water-soluble fractions from five different Cinnamomum species. FEMS Microbiol Lett. 2004 Feb 9;231(1):137-43. Shen YC, Chou CJ, Wang YH, Chen CF, Chou YC, Lu MK. National Research Institute of Chinese Medicine, Room 739, 155-1, Section 2, Li-Nong Street, Pei-tou District (112), Taipei, Taiwan, ROC. We have previously reported that polysaccharides extracted from fruiting bodies or cultured mycelia of Antrodia camphorata exhibit an anti-hepatitis B virus effect. In this study, we intended to elucidate the anti-inflammatory potency of six mycelial extracts, namely PDB-ext, CK-ext, CM-ext, CO-ext, CC-ext, and CKO-ext, isolated from mycelia of A. camphorata cultured with six different media including potato dextrose broth (PDB) and five water-soluble fractions from the wood of different Cinnamomum species, i.e. C. kanehirae (CK), C. micranthum (CM), C. osmophloeum (CO), C. camphora (CC), and C. kotoense (CKO), against reactive oxygen species (ROS) production induced by N-formyl-methionyl-leucyl-phenylalanine (fMLP) or phorbol 12-myristate 13-acetate (PMA) in peripheral human neutrophils (PMN) or mononuclear cells (MNC). ROS produced by PMN or MNC act as inflammatory mediators and also signal immune responses. Pretreatment with these mycelial extracts (1-50 microg ml(-1)) concentration-dependently diminished fMLP- or PMA-induced ROS production in PMN or MNC, as measured by lucigenin-amplified chemiluminescence, with 50% inhibition concentrations (IC(50)) ranging from 2 to 20 microg ml(-1). Among these extracts evaluated, CM-ext, CO-ext, or CKO-ext exhibited higher potency than the others. Using high performance liquid chromatography, we identified two lanostane-type compounds, i.e. dehydrosulfurenic acid and 15alpha-acetyl-dehydrosulfurenic acid, which could be involved in the anti-inflammatory actions of these extracts. The anti-inflammatory actions of these extracts were not due to cytotoxic effects. In summary, these data suggest that extracts from cultured mycelia of A. camphorata display anti-inflammatory effects by inhibiting ROS production in human leukocytes at a pharmacologically applicable concentration. The biological activities of these extracts were further promoted when the culture medium was replaced with water-soluble fractions isolated from the wood of CM, CO or CKO. PMID: 14769478 [PubMed - indexed for MEDLINE]

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30: Five new maleic and succinic acid derivatives from the mycelium of Antrodia camphorata and their cytotoxic effects on LLC tumor cell line. J Nat Prod. 2004 Jan;67(1):46-8. Nakamura N, Hirakawa A, Gao JJ, Kakuda H, Shiro M, Komatsu Y, Sheu CC, Hattori M. Institute of Natural Medicine, Toyama Medical and Pharmaceutical University, 2630 Sugitani, Toyama 930-0194, Japan. Five new maleic and succinic acid derivatives were isolated from the mycelium of Antrodia camphorata. Their structures were determined by various spectroscopic means. Maleimide derivatives 2 and 3 showed appreciable cytotoxic activity against LLC cells. PMID: 14738384 [PubMed - indexed for MEDLINE] 31: The vasorelaxation of Antrodia camphorata mycelia: involvement of endothelial Ca(2+)-NO-cGMP pathway. Life Sci. 2003 Oct 10;73(21):2769-83. Wang GJ, Tseng HW, Chou CJ, Tsai TH, Chen CT, Lu MK. National Research Institute of Chinese Medicine, Room 739, 155-1, Section 2, Li-Nong Street, Pei-tou District (112), Taipei, Taiwan, ROC. Antrodia camphorata, a medicinal fungus, has been used to treat cardiovascular diseases such as hypertension for many years. The purpose of this study was to examine the effects of mycelia extracts, from five Antrodia camphorata strains, on vascular tension and underlying mechanisms were explored. In isolated rat aortic rings, accession B86 caused concentration-dependent vasorelaxation with maximal relaxation of 40.34 +/- 7.53% whereas accessions 35398, 35396 and B71 had mild vasorelaxing effects. Strain B85 evoked potent vasorelaxation, partly through an endothelium-dependent mechanism that was inhibited by Nomega-nitro-L-arginine and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ) but not by antagonist of K+ channels, tetraethylammonium. In cultured endothelial cells, B85 stimulated nitric oxide (NO) release and augmented the level of the intracellular Ca2+ concentration. HPLC and LC-MS-MS analysis revealed the presence of adenosine. Our results suggest that B85 produced strongest vasorelaxation in aortic preparations among five test strains. B85 acts in part on endothelial cells by activating the Ca(2+)-NO-cGMP pathway to reduce smooth muscle tone. However, K+ channels had no apparent roles. Adenosine could possibly be involved in the endothelium-dependent pathway of B85-induced vasorelaxation. PMID: 13679244 [PubMed - indexed for MEDLINE] 32: Antioxidative and hepatoprotective effects of Antrodia camphorata extract. J Agric Food Chem. 2003 May 21;51(11):3302-8. Hsiao G, Shen MY, Lin KH, Lan MH, Wu LY, Chou DS, Lin CH, Su CH, Sheu JR. Department of Pharmacology and Graduate Institute of Medical Sciences, Taipei Medical University, 250 Wu-Hsing Street, Taipei 110, Taiwan. Antrodia camphorata (A. camphorata) is well-known in Taiwan as a traditional Chinese medicine. The purpose of this study was to evaluate the ability of A. camphorata extracts to protect against oxidative stress in vitro and against carbon tetrachloride (CCl(4))-induced hepatic injury in vivo. An extract of A. camphorata inhibited nonenzymatic iron-induced lipid peroxidation in rat brain homogenates with an IC(50) value about 3.1 mg/mL. It also scavenged the stable free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH). The dose of the A. camphorata extract resulting in a decrease of 0.20 in the absorbance of DPPH was about 31 +/- 0.7 microg/mL. Furthermore, an A. camphorata extract dose-dependently (250-1250 mg/kg) ameliorated the increase in plasma aspartate aminotransferase (GOT) and alanine aminotransferase (GPT) levels caused by chronic repeated CCl(4) intoxication in mice. Moreover, A. camphorata extract significantly improved the CCl(4)-induced increase in hepatic glutathione peroxidase, reductase, and CCl(4)-induced decrease in superoxide dismutase activities. It also restored the decrement in the glutathione content and catalase activity of hepatic tissues in CCl(4)-intoxicated mice. Furthermore, it also dose-dependently inhibited the formation of lipid peroxidative products during CCl(4) treatment. Histopathological changes of hepatic lesions induced by CCl(4) were significantly ameliorated by treatment with an A. camphorata extract in a dose-dependent manner. These results suggest that A. camphorata extract exerts effective protection against chronic chemical-induced hepatic injury in vivo, by mediating antioxidative and free radical scavenging activities. PMID: 12744658 [PubMed - indexed for MEDLINE]

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33: Protective effects of fermented filtrate from Antrodia camphorata in submerged culture against CCl4-induced hepatic toxicity in rats. J Agric Food Chem. 2003 Mar 12;51(6):1571-7. Song TY, Yen GC. Department of Food Science, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan, Republic of China. The protective effects and the possible mechanisms of dry matter of fermented filtrate (DMF) from Antrodia camphorata in submerged culture (ACSC) on H(2)O(2)-induced cytotoxicity in HepG2 and carbon tetrachloride (CCl(4))-induced hepatotoxicity in Sprague-Dawley rats were investigated. The results showed that the inhibitory effect of DMF and its crude triterpenoids on lipid peroxidation occurred in a dose-response manner in an AAPH/linoleic acid system. When HepG2 cells were pretreated with DMF at the concentration of 0.10 mg/mL for 4 h and then induced by 1 h of treatment with H(2)O(2) (100 microM), lipid peroxidation was significantly (p < 0.05) decreased, as measured by the formation of malondialdehyde. The oral pretreatment with DMF [0.25 and 0.50 mg/kg of body weight (bw)] for 5 consecutive days prior to the administration of a single dose of 40% CCl(4) (0.10 mL/100 g of bw, ip) significantly prevented the increase in serum levels of hepatic enzyme markers (alanine and aspartate aminotransferase) and liver lipid peroxidation (p < 0.05). Histopathological evaluation of the rat liver revealed that DMF reduced the incidence of liver lesions, including neutrophil infiltration, hydropic swelling, and necrosis induced by CCl(4) in rats. Moreover, reduced glutathione (GSH)-dependent enzymes (glutathione peroxidase, glutathione reductase, and glutathione S-transferase) and the GSH/GSSG ratio were significantly improved in the oral pretreatment DMF of rats (p < 0.01). The results suggest that DMF may play a role in preventing oxidative damage in living systems by up-regulating hepatic GSH-dependent enzymes to preserve the normal GSH/GSSH ratio and scavenging free radicals formed during CCl(4) metabolism. PMID: 12617586 [PubMed - indexed for MEDLINE] 34: Protection of oxidative damage by aqueous extract from Antrodia camphorata mycelia in normal human erythrocytes. Life Sci. 2002 Jun 14;71(4):469-82. Hseu YC, Chang WC, Hseu YT, Lee CY, Yech YJ, Chen PC, Chen JY, Yang HL. Department of Medical Technology, Fooyin Institute of Technology, Kaohssiung, Taiwan. Antrodia camphorata (A. camphorata) is well known in Taiwan as a traditional Chinese medicine. The purpose of this study was to evaluate the ability of aqueous extract from A. camphorata mycelia to protect normal human erythrocytes against oxidative damage in vitro. Oxidative hemolysis and lipid/protein peroxidation of erythrocytes induced by the aqueous peroxyl radical [2,2'-Azobis(2-amidinopropane) dihydrochloride, AAPH] were suppressed by A. camphorata mycelia in a time-and concentration-dependent manner. A. camphorata mycelia also prevented the depletion of cytosolic antioxidant glutathione (GSH) and ATP in erythrocytes. Moreover, cultured human endothelial cell damage induced by AAPH was suppressed by A. camphorata mycelia. Interestingly, A. camphorata mycelia exhibited significant cytotoxicity against leukemia HL-60 cells but not against cultured human endothelial cells. These results imply that A. camphorata mycelia may have protective antioxidant and anticancer properties. PMID: 12044846 [PubMed - indexed for MEDLINE] 35: Antioxidant properties of Antrodia camphorata in submerged culture. J Agric Food Chem. 2002 May 22;50(11):3322-7. Song TY, Yen GC. Department of Food Science, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan, Republic of China. The biologically active compounds, antioxidant activities, and free radical scavenging effects of dry matter of cultural medium (DMCM), filtrate (DMF), and different solvent extracts of mycelia from Antrodia camphorata in submerged culture (ACSC) were investigated. DMF showed the strongest inhibition of lipid peroxidation as a function of its concentration, and was comparable to the antioxidant activity of BHA at the same concentration of 0.2 mg/mL. The hexane extract of mycelia had the weakest antioxidant ability, whereas other mycelial extracts exhibited a modest inhibition of lipid peroxidation. DMF and water extract of mycelia (WEM) showed marked activity in free radical scavenging. The antioxidant activities of filtrate and mycelial extracts were correlated with the presence of total polyphenols, the crude triterpenoids, and the 20

protein/polysaccharide ratio of the crude polysaccharides. It was found that DMCM had lower antioxidant ability than DMF in different model systems, indicating that the major antioxidant components in DMF must be derived from the secondary metabolites of mycelia. The results presented herein indicate that DMF could possibly act as a chemopreventing agent with respect to free radical-related diseases. PMID: 12010005 [PubMed - indexed for MEDLINE] 36: Antrodia camphorata polysaccharides exhibit anti-hepatitis B virus effects. FEMS Microbiol Lett. 2002 Mar 19;209(1):63-7. Lee IH, Huang RL, Chen CT, Chen HC, Hsu WC, Lu MK. China Medical College, 91 Hsueh-Shih Road, Taichung 40421, Taiwan. Polysaccharides were extracted from fruiting bodies and cultured mycelia from five Antrodia camphorata strains. Polysaccharide profiles of the five strains, as determined by high-performance anion-exchange chromatography, showed varying yields and composition of neutral sugars. A. camphorata fruiting bodies also had different polysaccharide patterns compared to the cultured mycelium. Analysis of 26-day-old mycelia showed that the neutral sugars galactose, glucose, mannose, and galactosamine were predominant. All mycelia polysaccharide preparations exhibited anti-hepatitis B virus activity. Polysaccharides from strain B86 at a concentration of 50 microg ml(-1) showed the highest level of anti-hepatitis B surface antigen effect, which was higher than alpha-interferon at a dosage of 1000 U ml(-1). Only strains B86 and 35398 had substantial anti-hepatitis B e antigen activities. None of the polysaccharides exhibited cytotoxic effects. Publication Types: Comparative Study PMID: 12007655 [PubMed - indexed for MEDLINE] 37: Anti-inflammatory Benzenoids from Antrodia camphorata. J Nat Prod. 2007 Jun 9; [Epub ahead of print] Chen JJ, Lin WJ, Liao CH, Shieh PC. Graduate Institute of Pharmaceutical Technology, Tajen University, Pingtung 907, Taiwan, Republic of China, and Graduate Institute of Natural Products, Chang Gung University, Taoyuan 333, Taiwan, Republic of China. Four new compounds, including three new benzenoids, antrocamphin A (1), antrocamphin B (2), and 2,3,4,5-tetramethoxybenzoyl chloride (3), and a new 1,3-dioxolan-2-one derivative, antrodioxolanone (4), together with 13 known compounds have been isolated from the fruiting body of Antrodia camphorata. The structures of these new compounds were determined through spectral analyses including extensive 2D-NMR data. Among the isolates, antrocamphin A (1), antcin A (10), and antcin B (11) exhibited potent inhibition against fMLP-induced superoxide production with IC50 values less than 10 muM. PMID: 17559265 [PubMed - as supplied by publisher] 38. Comparative anti-inflammatory characterization of wild fruiting body, liquid-state fermentation, and solid-state culture of Taiwanofungus camphoratus in microglia and the mechanism of its action Journal of Ethnopharmacology & J Ethnopharmacol. 2007 May 18; [Epub ahead of print] Der-Zen Liub, g, Hong-Jen Liangc, Chien-Ho Chena, Ching-Hua Sub, g, Tzong-Huei Leef, Chun-Ting Huanga, Wen-Chi Houf, g , Shyr-Yi Lind, g, Wen-Bin Zhonge, Pei-Jung Lina, Ling-Fang Hunga and Yu-Chih Lianga, g,
a

School of Medical Laboratory Science & Biotechnology, College of Medicine, Taipei Medical University, No. 250, Wu-Hsing Street, Taipei 11014, Taiwan b Graduate Institutes of Biomedical Materials, Taipei Medical University, Taipei, Taiwan c Department of Food Science, Yuanpei University, HsinChu, Taiwan d Department of Internal Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan e Department of Physiology and Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan f Graduate Institute of Pharmacognosy Science, College of Pharmacy, Taipei Medical University, Taipei, Taiwan g Traditional Herbal Medicine Research Center, Taipei Medical University Hospital, Taipei, Taiwan Received 14 November 2006; revised 7 February 2007; accepted 26 March 2007. Available online 18 May 2007. 21

Abstract: Taiwanofungus camphoratus (syn. Antrodia camphorata), a medicinal mushroom in Taiwan, is reputed to provide several therapeutic benefits, but the wild fruiting body is very rare. In this study, we used Taiwanofungus camphoratus extracts from wild fruiting bodies and two types of artificial cultivation (solid-state culture and liquid-state fermentation) to examine their anti-inflammatory effects in microglia cells and their possible roles in protection against neurodegenerative diseases. First, EOC13.31 microglia was treated with various kinds of Taiwanofungus camphoratus extracts and lipopolysaccharide (LPS) and interferon- (IFN-) to evaluate the iNOS expression. Western blot and RT-PCR analysis showed that among the various kinds of extracts from wild fruiting bodies, methanol extracts were the most potent inhibitors of iNOS expression. Secondly, the potency of methanol extracts could be ranked as follows: extracts of wild fruiting body > solid-state culture > liquid-state fermentation. To clarify the mechanisms involved, methanol extracts from fruiting body were found to inhibit the phosphorylation of extracellular signal-regulated protein kinases (ERK), c-Jun NH2-terminal protein kinases (JNK) and signal transducer and activator of transcription-1 (STAT-1) induced by LPS/IFN-. Methanol extracts from fruiting body also inhibited NF-B activation through the prevention of inhibitor B (IB) degradation. Moreover, methanol extracts from wild fruiting body inhibited both the iNOS and cyclooxygenase-2 (COX-2) expression induced by -amyloid in microglia in a dose-dependent manner. In an animal model, we confirmed that methanol extracts from fruiting bodies were able to suppress ear edema, indicating that they have anti-inflammatory activity in vivo. These results suggest that Taiwanofungus camphoratus exhibits an anti-inflammatory activity that might contribute to the prevention of neurodegenerative diseases. Keywords: Inflammation; iNOS; Microglia; Taiwanofungus camphoratus Abbreviations: iNOS, inducible nitric oxide synthase; NO, nitric oxide; LPS, lipopolysaccharide; NF-B, nuclear factor-B; IB, inhibitor B; IFN-, interferon-; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; PCR, polymerase chain reaction; TPA, 12-O-tetradecanoylphorbol-13-acetate; COX-2, cyclooxygenase 2; WC, cold water extracts from wild fruiting bodies of Taiwanofungus camphoratus; WH, hot water extracts from wild fruiting bodies of Taiwanofungus camphoratus; WM, methanol extracts from wild fruiting bodies of Taiwanofungus camphoratus; LM, methanol extracts from liquid-state fermentation of Taiwanofungus camphoratus; SM, methanol extracts from solid-state culture of Taiwanofungus camphoratus PMID: 17590297 [PubMed - as supplied by publisher] Corresponding author. Tel.: +886 2 27361661x3318; fax: +886 2 27393447. 39. A highly stable cambialistic-superoxide dismutase from Antrodia camphorata: Expression in yeast and enzyme properties J Biotechnol. 2007 Aug 1;131(1):84-91. Epub 2007 May 31. Yi-Jen Liaua, 1, Lisa Wenb, 1, Jei-Fu Shawc, 1 and Chi-Tsai Lina, a Institute of Bioscience and Biotechnology, National Taiwan Ocean University, 2 Pei-Ning Rd, Keelung 202, Taiwan b Department of Chemistry, Western Illinois University, 1 University Circle, Macomb, IL 61455-1390, USA c Department of Food Science and Biotechnology, National Chung Hsing University, Taichung 402, Taiwan Received 15 October 2006; revised 15 May 2007; accepted 21 May 2007. Available online 31 May 2007. Abstract A cDNA encoding a putative superoxide dismutase (SOD) was identified in expressed sequence tags of Antrodia camphorata, a medicinal mushroom found only in Taiwan. The deduced protein was aligned with Mn-SODs and Fe-SODs from other organisms, this SOD showed greater homology to Mn-SOD. Functional A. camphorata SOD protein was overexpressed in yeast and purified. The purified enzyme showed two active forms on a 12.5% native PAGE, a dimer and a monomer. The dimeric protein's half-life of deactivation at 80 C was 7 min, and its thermal inactivation rate constant Kd was 9.87 102 min1. The enzyme was stable in a broad pH range from 511; in the presence of 0.4 M imidazole and 2% SDS. The atomic absorption spectrometric assay showed that 1.0 atom of manganese/iron (9:1) was present in each SOD subunit. The high stability of the enzyme make it better suited than other cambialistic-SODs for use in cosmetics. The SOD also documents its future utility in developing anti-inflammatory agent and in the treatment of chronic diseases. Keywords: Mushroom; Antrodia camphorata; Expression; Cambialistic-superoxide dismutase (Mn/Fe-SOD); Yeast Corresponding author. Tel.: +886 2 24622192x5513; fax: +886 2 24622320. 1 These authors contributed equally to this paper. 40: A New Cytotoxic Agent from Solid-State Fermented Mycelium of Antrodia camphorata. 22

Planta Med. 2007 Oct 11; [Epub ahead of print] Lee TH, Lee CK, Tsou WL, Liu SY, Kuo MT, Wen WC. Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan, R.O.C. Antroquinonol ( 1), an ubiquinone derivative, was isolated from the solid-state fermented mycelium of ANTRODIA CAMPHORATA (Polyporaceae, Aphyllophorales), a parasitic fungus indigenous to Taiwan. The structure of compound 1 was elucidated by the analysis of their spectroscopic data. Its cytotoxic activities were evaluated against MCF-7, MDA-MB-231 (human breast carcinoma), Hep3B, HepG2 (human liver carcinoma) and DU-145, LNCaP (human prostate carcinoma) cell lines, and the IC (50) values ranged from 0.13 +/- 0.02 to 6.09 +/- 0.07 muM. PMID: 17932820 [PubMed - as supplied by publisher] 41: Evaluation of the anti-inflammatory and anti-proliferation tumoral cells activities of Antrodia camphorata, Cordyceps sinensis, and Cinnamomum osmophloeum bark extracts. J Ethnopharmacol. 2007 Oct 8;114(1):78-85. Epub 2007 Aug 2. Rao YK, Fang SH, Tzeng YM. Institute of Biotechnology, Chaoyang University of Technology, Wufeng 413, Taiwan, ROC. The extracts of chloroform (1) and methanol (2) from Antrodia camphorata (AC), and chloroform (3) and n-butanol (4) fractions of methanol extract from Cordyceps sinensis (CS), and hexane (5), ethyl acetate (6), and methanol (7) from Cinnamomum osmophloeum bark (CO) were evaluated for their anti-inflammatory as well as tumor-cell growth inhibitory activities in vitro. All the tested extracts dose dependently inhibited the enhanced production of inflammatory mediators such as nitric oxide (NO) through reducing inducible NO synthase expression, and cytokines (tumor necrosis factor (TNF)-alpha and interleukin (IL)-12 in LPS/IFN-gamma activated murine peritoneal macrophages. In addition, extracts 1 from AC, and 5 and 6 from CO significantly arrest the mitogen-stimulated spleen cells in G0/G1 stage. On the other hand, all these extracts were also evaluated for their tumor-cell proliferation activities in different type of cancer cell lines such as Jurkat, HepG2, PC 3, Colon 205, and MCF 7 as well as normal PBMCs. Compared to untreated controls, the extracts 1, 2, and 4-7 were most active and inhibited Jurkat cells with IC(50) value of 22, 40, 18, 4, 5, and 45mug/ml, respectively. In addition, the extracts 5, 6, and 7 from CO showed potent growth inhibition of HepG2 and PC 3 with IC(50) values of 35, 80, 55mug/ml; and 42, 125, and 50mug/ml, respectively. Similarly, the extracts 1 and 5 inhibited the growth of Colon 205 and MCF 7 cells with IC(50) values of 65, 33; and 95 and 30mug/ml, respectively. Interestingly, none of the tested extract has shown cytotoxicity towards normal PBMCs up to the concentration range studies (0-150mug/ml). Taken together, these data suggest that the anti-inflammatory and anti-cancer properties of AC, CS, and CO might result from the growth inhibition of NO, TNF-alpha and IL-12, and tumor cells proliferation, respectively. PMID: 17822865 [PubMed - in process] 42: In vivo immunomodulatory effects of Antrodia camphorata polysaccharides in a T1/T2 doubly transgenic mouse model for inhibiting infection of Schistosoma mansoni. Toxicol Appl Pharmacol. 2007 Nov 7 [Epub ahead of print] Cheng PC, Hsu CY, Chen CC, Lee KM. Institute of Tropical Medicine, National Yang-Ming University, Taipei, Taiwan. Antrodia camphorata (A. camphorata) is a fungus commonly used for treatment of viral hepatitis and cancer in Chinese folk medicine. Extract of A. camphorate is reported to possess anti-inflammatory, antihepatitis B virus and anticancer activities. In this study, we tested the in vivo effects of polysaccharides derived from A. camphorata (AC-PS) on immune function by detection of cytokine expression and evaluation of the immune phenotype in a T1/T2 doubly transgenic mouse model. The protective effect of AC-PS in mice was tested by infection with Schistosoma mansoni. The induction of large amounts of IFN-gamma, IL-2 and TNF- mRNA were detected after 2 and 4 weeks of oral AC-PS administration in BALB/c and C57BL/6 mice. In transgenic mice, 3 to 6 weeks of oral AC-PS administration increased the proportion of CD4(+) T cells and B cells within the spleen. More specifically, there was an increase of Th1 CD4(+) T cells and Be1 cells among spleen cells as observed by detection the of Type1/Type2 marker molecules. By using a disease model of parasitic infection, we found that AC-PS treatment inhibited infection with S. mansoni in BALB/C and C57BL/6 mice. AC-PS appears to influence the immune system of mice into developing Th1 responses and have potential for preventing infection with S. mansoni. PMID: 18078970 [PubMed - as supplied by publisher] 23

43. Fermented Antrodia cinnamomea Extract Protects Rat PC12 Cells from Serum Deprivation-Induced Apoptosis: The Role of the MAPK Family. J Agric Food Chem. 2008 Jan 11 [Epub ahead of print] Lu MK, Cheng JJ, Lai WL, Lin YJ, Huang NK. andrew@mail.nricm.edu.tw. Antrodia cinnamomea (formerly A. camphorata) has recently and commercially been used in the formulation of nutraceuticals and functional foods in Taiwan. Because of its diverse properties, the neuroprotective effect was investigated using a fermented A. cinnamomea extract in this study. Serum deprivation-induced apoptosis in neuronal-like pheochromocytoma (PC12) cells was used as a cell stress model, and it was found that A. cinnamomea was effective in preventing serum-deprived apoptosis according to results of an MTT assay and Hoechst staining. Serum deprivation resulted in decreased phosphorylation of extracellular signal-regulated kinase (ERK) and increased phosphorylations of c-Jun NH 2-terminal kinase (JNK) and p38, of the family of mitogen-activated protein kinases (MAPKs); however, A. cinnamomea reversed these phenomena, supporting the antagonistic effects between ERK and JNK-p38 in regulating cell survival. The previously identified active component of A. cinnamomea, adenosine (ADO), also exerted the same effects as A. cinnamomea in preventing apoptosis and regulating phosphorylations of MAPKs. Although an inhibitor of the ERK upstream activator blocked A. cinnamomea-induced ERK phosphorylations, it failed to block the protection of A. cinnamomea and ADO. A protein kinase A (PKA) inhibitor blocked the protection by both A. cinnamomea and ADO. Both JNK and p38 inhibitors were effective in preventing the phosphorylations of JNK and p38 and serum deprivation-induced apoptosis. Collectively, A. cinnamomea prevented serum deprivation-induced PC12 cell apoptosis through a PKA-dependent pathway and by suppression of JNK and p38 activities. PMID: 18186605 [PubMed - as supplied by publisher] 44. Isolation and analysis of genes specifically expressed during basidiomatal development in Antrodia cinnamomea by subtractive PCR and cDNA microarray. FEMS Microbiol Lett. 2008 Jan 22 [Epub ahead of print] Chu FH, Lee YR, Chou SJ, Chang TT, Shaw JF. School of Forestry and Resource Conservation, National Taiwan University, Taipei, Taiwan. cDNAs specifically expressed at the basidiome stage were isolated by using PCR-selected cDNA subtraction in order to study gene regulation during porous-hymenium basidiomatal formation in Antrodia cinnamomea. blastx results suggested that most of the expressed sequence tags (52.4-69.5%) had no significant protein homology to genes from other published living things. cDNAs particularly expressed at different growing conditions were identified using cDNA microarray analysis. Reverse transcriptase PCR analyses confirmed that the clone putative to P-type ATPase, various cytochrome P450s and some unknown genes were abundant at natural basidiomes while endoglucanase was abundant at the tissue from artificial medium. PMID: 18218021 [PubMed - as supplied by publisher] 45. DPPH radical scavenging and semicarbazide-sensitive amine oxidase inhibitory and cytotoxic activities of Taiwanofungus camphoratus (Chang-chih). Biosci Biotechnol Biochem. 2007 Aug;71(8):1873-8. Epub 2007 Aug 7. Wang GJ, Lin SY, Wu WC, Hou WC. National Research Institute of Chinese Medicine, Taipei, Taiwan. Wild, liquid state culture and solid state culture of Taiwanofungus camphoratus (Chang-chih) were sequentially extracted with cold water, methanol, and hot water to get cold water soluble, methanol soluble, and hot water soluble extracts respectively. The extracts from three Chang-chih were used to determine 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging, semicarbazide sensitive amine oxidase inhibitory, and cytotoxic activities against B16-F10 and HT-1080 cell lines. It was found that extracted fractions from three Chang-chih exhibited the different levels of biological activities. PMID: 17690475 [PubMed - indexed for MEDLINE] 46. Effects on Tyrosinase Activity by the Extracts of Ganoderma lucidum and Related Mushrooms. Mycopathologia. 2008 May 6 [Epub ahead of print] 24

Chien CC, Tsai ML, Chen CC, Chang SJ, Tseng CH. Graduate School of Biotechnology and Bioengineering, Yuan Ze University, 135 Yuan-Tung Rd, Chungli, 320, Taiwan, ccchien@saturn.yzu.edu.tw. The inhibitory effects on tyrosinase activity by extracts of several mushrooms belonging to Basidiomycetes were evaluated. Among the tested mushrooms (Ganoderma lucidum, Antrodia camphorata, Agaricus brasiliensis, and Cordyceps militaris), G. lucidum exhibited significant inhibition of tyrosinase activity (IC(50) value 0.32 mg/ml), compared to those prepared from other Basidiomycetes. Tyrosinase inhibitors are effective components of skin-lightening compounds and other cosmetics; currently many of the facial mask cosmetics in the market contain Ganoderma extracts in their ingredients. The finding that mushroom extracts contain tyrosinase activity inhibition will contribute to better understanding of how their 'healing' properties in various Chinese traditional herbal on skin care products. PMID: 18459064 [PubMed - as supplied by publisher] 47. Polysaccharides from Antrodia camphorata mycelia extracts possess immunomodulatory activity and inhibits infection of Schistosoma mansoni. Int Immunopharmacol. 2008 Mar;8(3):458-67. Epub 2007 Dec 28. Chen YJ, Cheng PC, Lin CN, Liao HF, Chen YY, Chen CC, Lee KM. Department of Radiation Oncology, Mackay Memorial Hospital, Taipei, Taiwan. Antrodia camphorata (AC) is a commonly used fungus in folk medicine for the treatment of viral hepatitis and cancer. AC polysaccharides (AC-PS) are reported to possess anti-inflammatory, anti-hepatitis B virus, and anticancer activities. In this study, we tested the in vivo effect of AC-PS on immune function by evaluating cytokine expression; on immunomodulation, by evaluating spleen cells; and on Schistosoma mansoni infection in mice. The induction of high levels of interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) mRNA was detected in BALB/c mice after 2, 4, and 6 weeks of oral AC-PS administration. After 6 weeks of oral AC-PS administration to the BALB/c mice, the number of splenic dendritic cells, macrophages, and the surface expression of CD8 alpha+ and major histocompatibility class II I-A/I-E on dendritic cells increased. The CD4+/CD8+ ratio and number of B cells among splenocytes were also augmented. By using a disease model of parasitic infection, we found that AC-PS treatment inhibited S. mansoni infection in BALB/c mice. AC-PS appears to modulate the immune system of mice and has potential for preventing S. mansoni infection. PMID: 18279800 [PubMed - indexed for MEDLINE] 48: The adjuvant effects of Antrodia Camphorata extracts combined with anti-tumor agents on multidrug resistant human hepatoma cells. J Ethnopharmacol. 2008 May 7. [Epub ahead of print] Chang CY, Huang ZN, Yu HH, Chang LH, Li SL, Chen YP, Lee KY, Chuu JJ. Institute of Biotechnology, College of Engineering, Southern Taiwan University, Tainan, Taiwan; Department of Medicine, Chi-Mei Medical Center, Yung-Kang City, Tainan Hsien, Taiwan. AIM OF THE STUDY: The objectives of this study were to investigate the adjuvant anti-tumor effects of Antrodia camphorate in human hepatoma cells (C3A and PLC/PRF/5) which are resistance to most anti-tumor agents, elucidate the possible regulation pathways, and measure the tumor growth and survival rate in xenograft-nude mice after combined with anti-tumor agents. MATERIALS AND METHODS: The AC extracts were measured by using a phenol/sulfuric acid method as previously described. The in vitro cell proliferation assay of ACs and anti-tumor agents was tested on C3A and PLC/PRF/5 cell lines. The percentage of human hepatoma cells undergoing apoptosis and distributing in different phases of cell cycle were determined by Flow cytometric analysis. Western blot analysis for MDR-1 and apoptosis- related proteins. The measurements of tumor growth and survival analysis of hepatoma implanted nude mice treated with Antrodia camphorata extracts and anti-tumor agents alone or in combinations. RESULTS: We have found that Antrodia camphorata extracts, when combined with anti-tumor agents, showed adjuvant antiproliferative effects on hepatoma cells (in vitro) and on xenografted cells in tumor-implanted nude mice (in vivo), which then extended their median survival days. Furthermore, solid-state extracts of Antrodia camphorata (AC-SS) showed its adjuvant effects through the inhibition of MDR gene expressions and the pathway of COX-2- dependent inhibition of p-AKT, which ultimately resulted in the induction of apoptosis in hepatoma cells. CONCLUSIONS: In this study, we have found that Antrodia camphorata extract, when combined with anti-tumor agents, showed adjuvant antiproliferative effects on hepatoma cells (in vitro) and on xenografted cells in tumor-implanted nude mice (in vivo). 25

PMID: 18571350 [PubMed - as supplied by publisher] 49: Anti-inflammatory Activities of New Succinic and Maleic Derivatives from the Fruiting Body of Antrodia camphorata. J Agric Food Chem. 2008 Jul 19. [Epub ahead of print] Chien SC, Chen ML, Kuo HT, Tsai YC, Lin BF, Kuo YH. yhkuo@ntu.edu.tw. Six new compounds, trans-3-isobutyl-4-[4-(3-methyl-2-butenyloxy)phenyl]pyrrolidine-2,5-dione ( 1), trans-1-hydroxy-3-(4-hydroxyphenyl)-4-isobutylpyrrolidine-2,5-dione ( 2), cis-3-(4-hydroxyphenyl)-4-isobutyldihydrofuran-2,5-dione ( 3), 3-(4-hydroxyphenyl)-4-isobutyl-1 H-pyrrole-2,5-dione ( 4), 3-(4-hydroxyphenyl)-4-isobutylfuran-2,5-dione ( 5), and dimethyl 2-(4-hydroxyphenyl)-3-isobutylmaleate ( 6), together with one known compound, 3-isobutyl-4-[4-(3-methyl-2-butenyloxy)phenyl]furan-2,5-dione ( 7), were isolated from the fruiting bodies of Antrodia camphorata. The structures of the compounds were elucidated by analysis of their spectroscopic data. To investigate the immunomodulatory potential of the compounds, RAW264.7 macrophage cells were treated with the compounds. Compound 1 significantly increased spontaneous TNF-alpha secretion from unstimulated RAW264.7 cells but suppressed IL-6 production [50% inhibition concentration value (IC 50) = 10 mug/mL] in LPS-stimulated cells. Compounds 3, 4, and 6 also suppressed IL-6 production with IC 50 values of 17, 18, and 25 mug/mL, respectively, suggesting that these four compounds may have an anti-inflammatory effect on macrophage-mediated responses. Of the six compounds, compound 1 was the most effective, exerting both immunostimulatory and anti-inflammatory effects. PMID: 18642845 [PubMed - as supplied by publisher] 50. Antioxidant activity of Antrodia camphorata on free radical-induced endothelial cell damage. J Ethnopharmacol. 2008 Jul 23;118(2):237-45. Epub 2008 Apr 12. Hseu YC, Chen SC, Yech YJ, Wang L, Yang HL. Department of Cosmeceutics, China Medical University, Taichung, Taiwan. AIM OF THE STUDY: Antrodia camphorata (A. camphorata) is well known in Taiwan as a traditional Chinese medicine. The purpose of this study is to evaluate the antioxidant activity of Antrodia camphorata on free radical-induced endothelial cell damage. MATERIALS AND METHODS: In this study, a human umbilical vein endothelial cell (EC) culture system was used to evaluate the effects of the fermented culture broth of A. camphorata (FCBA) and aqueous extracts of mycelia from A. camphorata (AEMA) against the oxidative cell damage induced by the free-radical generator AAPH. RESULTS: The present investigations show that FCBA (25-100mug/mL) and AEMA (50-200mug/mL) effectively protect the ECs from damage after exposure to 15mM AAPH for 16h. However, cell viability was not affected in ECs under controlled conditions after FCBA or AEMA treatment. An increase in EC prostacyclin (PGI(2)) production in response to AAPH exposure was positively and negatively correlated with cell damage and FCBA/AEMA concentration, respectively. Both FCBA and AEMA treatment significantly inhibited AAPH-apoptotic cell death in the ECs, as evidence by reduced DNA fragmentation, cytochrome c release, caspase-3 activation, and dysregulation of Bcl-2 and Bax. Moreover, the AAPH-induced reductions in EC SOD activity and protein levels are prevented by FCBA and AEMA. CONCLUSION: Our findings suggest that A. camphorata possesses antioxidant properties and improves endothelial function, further offering effective protection from atherosclerosis. PMID: 18486375 [PubMed - in process] 51. Effects on Tyrosinase Activity by the Extracts of Ganoderma lucidum and Related Mushrooms. Mycopathologia. 2008 Aug;166(2):117-20. Epub 2008 May 6. Chien CC, Tsai ML, Chen CC, Chang SJ, Tseng CH. Graduate School of Biotechnology and Bioengineering, Yuan Ze University, 135 Yuan-Tung Rd, Chungli, 320, Taiwan, ccchien@saturn.yzu.edu.tw. The inhibitory effects on tyrosinase activity by extracts of several mushrooms belonging to Basidiomycetes were evaluated. Among the tested mushrooms (Ganoderma lucidum, Antrodia camphorata, Agaricus brasiliensis, and Cordyceps militaris), G. lucidum exhibited significant inhibition of tyrosinase activity (IC(50) value 0.32 mg/ml), compared to those prepared from other Basidiomycetes. Tyrosinase inhibitors are effective components of skin-lightening compounds and other cosmetics; currently many of the facial mask cosmetics in the market contain Ganoderma extracts in their ingredients. The 26

finding that mushroom extracts contain tyrosinase activity inhibition will contribute to better understanding of how their 'healing' properties in various Chinese traditional herbal on skin care products. PMID: 18459064 [PubMed - in process] 52 Antrodia camphorata inhibits proliferation of human breast cancer cells in vitro and in vivo. Food Chem Toxicol. 2008 Aug;46(8):2680-8. Epub 2008 May 4. Hseu YC, Chen SC, Chen HC, Liao JW, Yang HL. Department of Cosmeceutics, China Medical University, Taichung, Taiwan. Antrodia camphorata (A. camphorata) has been shown to induce apoptosis in cultured human breast cancer cells (MDA-MB-231). In this study, we report the effectiveness of the fermented culture broth of A. camphorata in terms of tumor regression as determined using both in vitro cell culture and in vivo athymic nude mice models of breast cancer. We found that the A. camphorata treatment decreased the proliferation of MDA-MB-231 cells by arresting progression through the G1 phase of the cell cycle. This cell cycle blockade was associated with reductions in cyclin D1, cyclin E, CDK4, cyclin A, and proliferating cell nuclear antigen (PCNA), and increased CDK inhibitor p27/KIP and p21/WAF1 in a dose and time-dependent manner. Furthermore, the A. camphorata treatment was effective in delaying tumor incidence in the nude mice inoculated with MDA-MB-231 cells as well as reducing the tumor burden when compared to controls. A. camphorata treatment also inhibited proliferation (cyclin D1 and PCNA) and induced apoptosis (Bcl-2 and TUNEL) when the tumor tissue sections were examined histologically and immunohistochemically. These results suggest that the A. camphorata treatment induced cell cycle arrest and apoptosis of human breast cancer cells both in vitro and in vivo. PMID: 18550246 [PubMed - in process] 53. Active extracts of wild fruiting bodies of Antrodia camphorata (EEAC) induce leukemia HL 60 cells apoptosis partially through histone hypoacetylation and synergistically promote anticancer effect of trichostatin A. Arch Toxicol. 2008 Aug 16. [Epub ahead of print] Lu MC, Du YC, Chuu JJ, Hwang SL, Hsieh PC, Hung CS, Chang FR, Wu YC. Graduate Institute of Natural Products, School of Pharmacy, Kaohsiung Medical University, Kaohsiung, 807, Taiwan, Jinx6609@yahoo.com. The endemic species of Antrodia camphorate (AC) is a promising chemotherapeutic drug for cancer. We found that the ethanol extract from wild fruiting bodies of Antrodia camphorata (EEAC) could induce HL 60 cells apoptosis via histone hypoacetylation, up-regulation of histone deacetyltransferase 1 (HDAC 1), and down-regulation of histone acetyltransferase activities including GCN 5, CBP and PCAF in dose-dependent manner. In combination with histone deacetylase inhibitor, trichostatin A (TSA), did not block EEAC-induced apoptosis. Interestingly, combined treatment (100 nM of TSA and 100 mug/ml EEAC) caused synergistic inhibition of cell growth and increase of apoptotic induction. EEAC could effectively increase the cytotoxic sensitivity of TSA through the up-regulation of DR5 and NFkappaB activation. In this present study, bioassay-guided fractionation of EEAC led to a major active compound, zhankuic acid A, as the bioactive marker. Moreover, our findings may represent an experimental basis for developing EEAC as a potential chemotherapeutic adjuvant. PMID: 18709356 [PubMed - as supplied by publisher] 54: Effects of antrodia camphorata on viability, apoptosis, and [Ca2+]i in PC3 human prostate cancer cells. Chin J Physiol. 2008 Apr 30;51(2):78-84. Ho CM, Huang CC, Huang CJ, Cheng JS, Chen IS, Tsai JY, Jiann BP, Tseng PL, Kuo SJ, Jan CR. Department of Medical Education and Research, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan, Republic of China. Antrodia camphorata (AC) has been used as a health supplement in Asia to control different cancers; however, the cellular mechanisms of its effects are unclear. The effect of AC on cultured human prostate cancer cells (PC3) has not been explored. This study examined the effect of AC on viability, apoptosis, mitogen-activated protein kinases (MAPKs) phosphorylation and Ca2+ handling in PC3 cells. AC at concentrations of 5-50 microg/ml did not affect cell viability, but at 100-200 microg/ml decreased viability and induced apoptosis in a concentration-dependent manner. AC at concentrations of 25-200 microg/ml did not alter basal [Ca2+]i, but at a concentration of 25 microg/ml decreased the [Ca2+]i increases induced by ATP, bradykinin, histamine and thapsigargin. ATP, bradykinin and histamine increased cell viability whereas thapsigargin 27

decreased it. AC (25 microg/ml) pretreatment inhibited ATP-, bradykinin-, and histamine-induced enhancement on viability, but reversed thapsigargin-induced cytotoxicity. Immunoblotting showed that AC (200 microg/ml) did not induce the phosphorylation of ERK, JNK, and p38 MAPKs. Collectively, in PC3 cells, AC exerted multiple effects on viability and [Ca2+]i, caused apoptosis via pathways unrelated to [Ca2+]i signal and phosphorylation of ERK, JNK and p38 MAPKs. PMID: 18666710 [PubMed - in process] 55. Immunomodulatory effect of Antrodia camphorata mycelia and culture filtrate. J Ethnopharmacol. 2008 Nov 20;120(2):196-203. Epub 2008 Aug 19 Kuo MC, Chang CY, Cheng TL, Wu MJ. Department of Biotechnology, Chia-Nan University of Pharmacy and Science, Tainan, Taiwan. AIM OF THE STUDY: Antrodia camphorata, a precious folkloric medicinal mushroom, has been used to treat tumorigenic diseases in Taiwan. This study was to investigate the innate immunity augmentation effects of different fractions prepared from hot water extracts of submerged cultured Antrodia camphorata (AC). MATERIALS AND METHODS: The cytokine induction potency of AC fraction in diluted peripheral blood culture was measured by ELISA. The effects of AC fraction on phagocytic activity and CD11b expression were measured by the ingestion of FITC-labeled Escherichia coli and by labeling with PE-labeled CD11b monoclonal antibody, respectively, using flow cytometry. The molecular mass of hot water-soluble polysaccharides and content of adenosine in AC fraction were determined by gel permeation chromatography (GPC) and HPLC, respectively. RESULTS: The mycelia fraction, Fr. M II, and culture filtrate fractions, Fr. E II and Fr. E III, showed the strongest TNF-alpha and IL-6 induction effect as a function of their concentration. These fractions (20mug/ml) also showed marked activity in enhancing phagocytosis in human polymorphonuclear neutrophils (PMN) and monocytes. In parallel, the expression of CD11b, an early marker of PMN activation, was also up-regulated dose-dependently. Composition analysis suggested that immunomodulatory effect of mycelia is mainly attributed to the 10-20kDa polysaccharides and adenosine. CONCLUSIONS: These results provide evidences that Antrodia camphorata can modulate innate immunity and may serve as an adjuvant for tumor treatment. PMID: 18778764 [PubMed - in process 56. Probing Inhibitory Effects of Antrodia camphorata Isolates Using Insect Cell-Based Impedance Spectroscopy: Inhibition vs Chemical Structure. Chem Res Toxicol. 2008 Sep 25. [Epub ahead of print] Male KB, Rao YK, Tzeng YM, Montes J, Kamen A, Luong JH. Biotechnology Research Institute, National Research Council Canada, Montreal, Quebec, Canada H4P 2R2, Institute of Biotechnology, Chaoyang University of Technology, Wufeng, 41349 Taiwan, Republic of China, and Department of Chemistry, University College Cork, Cork, Ireland ymtzeng@cyut.edu.tw, john.luong@cnrc-nrc.gc.ca, j.luong@ucc.ie. A continuous online technique based on electric cell-substrate impedance sensing (ECIS) was used for probing inhibitory effects on Spodoptera frugiperda S f9 insect cells exposed to structurally similar compounds isolated and purified from the fruiting bodies of the fungus Antrodia camphorata. Such chemicals consisted of three ergostane-related steroids and five lanosta-related triterpenes, which are known for their diverse properties and use in the formulation of nutraceuticals and functional foods. The half-inhibition concentration (ECIS 50), the level at which 50% inhibition of the resistance response was obtained, was determined from the response function to establish inhibitory effects of the different isolates. A slight change in their chemical structures resulted in significant effects on inhibition as probed by impedance spectroscopy. The ergostane-related steroids were mostly inhibitory, but replacing their ketone groups with hydrogen or hydroxyl groups significantly reduced the inhibition. Similarly, the addition of methyl or carboxymethyl groups also lowered the inhibition. Removal of the double bond conjugation within the rings (sulfurenic acid) of the isolate drastically reduced the inhibition. PMID: 18816072 [PubMed - as supplied by publisher 57. An Extract of Antrodia camphorata Mycelia Attenuates the Progression of Nephritis in Systemic Lupus Erythematosus-Prone NZB/W F1 Mice Evid Based Complement Alternat Med. 2008 Sep 2. [Epub ahead of print] Chang JM, Lee YR, Hung LM, Liu SY, Kuo MT, Wen WC, Chen P. Division of Research and Development, Development Center for Biotechnology, 101, Lane 169, Kangning Street, Xizhi City, Taipei County, Taiwan 221, ROC. jiaming@ntu.edu.tw. 28

Antrodia camphorata is used in folk medicine for the treatment of inflammation syndromes and liver-related diseases in Taiwan. The goal of this study was to evaluate the efficacy of the mycelial extract of A. camphorata (ACE) for the treatment of systemic lupus erythematosus (SLE) in SLE-prone NZB/W F1 mice. After antibodies against double-stranded DNA appeared in NZB/W mice, the mice were orally administered varying dosages of ACE (100, 200 and 400 mg kg(-1)) for 5 consecutive days per week for 12 weeks via gavage. To assess the efficacy of ACE, we measured SLE-associated biochemical and histopathological biomarkers levels of blood urine nitrogen (BUN), blood creatinine, urine protein and urine creatinine and thickness of the kidney glomerular basement membrane by staining with periodic acid-Schiff. Antroquinonol, an active component of ACE, was investigated for anti-inflammation activity in lipopolysaccharide-induced RAW 267.4 cells.ACE at 400 mg kg(-1) significantly suppressed urine protein and serum BUN levels and decreased the thickness of the kidney glomerular basement membrane. Antroquinonol significantly inhibited the production of tumor necrosis factor-alpha and interleukin-1beta by 75 and 78%, respectively. In conclusion, ACE reduced urine protein and creatinine levels and suppressed the thickening of the kidney glomerular basement membrane, suggesting that ACE protects the kidney from immunological damage resulting from autoimmune disease. PMID: 18955361 [PubMed - as supplied by publisher] 58. Niuchangchih (Antrodia camphorata) and its potential in treating liver diseases. J Ethnopharmacol. 2009 Jan 21;121(2):194-212. Epub 2008 Nov 17 Ao ZH, Xu ZH, Lu ZM, Xu HY, Zhang XM, Dou WF. Laboratory of Pharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, No. 1800 Lihu Road, Wuxi 214122, PR China. Niuchangchih (Antrodia camphorata (M. Zang & C.H. Su) Sheng H. Wu, Ryvarden & T.T. Chang) is a basidiomycete endemic to Taiwan. It is well known as a Traditional Chinese Medicine (TCM), and Taiwanese aborigines used this species to treat liver diseases and food and drug intoxication. The compounds identified in Niuchangchih are predominantly polysaccharides, triterpenoids, steroids, benzenoids and maleic/succinic acid derivatives. Recent research has revealed that Niuchangchih possesses extensive biological activity, such as hepatoprotective, antihypertensive, anti-hyperlipidemic, immuno-modulatory, anticancer, anti-inflammatory and antioxidant activities. The fruiting bodies and fermented products of Niuchangchih have been reported to exhibit activity when treating liver diseases, such as preventing ethanol-, CCl(4)- and cytokine-induced liver injury, inhibiting the hepatitis B virus, ameliorating fatty liver and liver fibrosis, and inhibiting liver cancer cells. This review will address the protective effects of Niuchangchih on the pathological development of liver diseases, and the underlying mechanisms of action are also discussed. 59. New Constituents with iNOS Inhibitory Activity from Mycelium of Antrodia camphorata. Planta Med. 2009 Apr;75(5):512-6. Epub 2009 Feb 2. Yang SS, Wang GJ, Wang SY, Lin YY, Kuo YH, Lee TH. Department of Internal Medicine, Cathy General Hospital, Taipei, Taiwan. In continuing our investigation on the bioactive constituents of mycelium of ANTRODIA CAMPHORATA, antroquinonol B ( 1), 4-acetyl-antroquinonol B ( 2), 2,3-(methylenedioxy)-6-methylbenzene-1,4-diol ( 3) and 2,4-dimethoxy-6-methylbenzene-1,3-diol ( 4) along with antrodin D ( 5) were isolated by the guidance of an inducible nitric oxide synthase (iNOS) inhibitory assay and identified on the basis of their spectroscopic analysis. The effect of these compounds on the inhibition of NO production in lipopolysaccharide (LPS)-activated murine macrophages was further evaluated. Compounds 4 and 5 significantly inhibited NO production without any cytotoxicity, the IC (50) values being 32.2 +/- 0.1 and 26.3 +/- 1.6 microg/mL, respectively. Compounds 1 and 2 possessed greater effects on NO inhibition, with IC (50) values of 16.2 +/- 0.8 and 14.7 +/- 2.8 microg/mL, respectively, but displayed cytotoxicity at considerably higher concentrations. Compound 3 showed the lowest percent cell viability of 45.5 +/- 1.8 % as observed in treated cells at a concentration of 16.8 microg/mL. SUPPORTING INFORMATION available online at http://www.thieme-connect.de/ejournals/toc/plantamedica. PMID: 19189245 [PubMed - in process] 60. Purification, Cloning, and Functional Characterization of a Novel Immunomodulatory Protein from Antrodia camphorata (Bitter Mushroom) That Exhibits TLR2-Dependent NF-kappaB Activation and M1 Polarization within Murine Macrophages. 29

J Agric Food Chem. 2009 Apr 17. [Epub ahead of print] Sheu F, Chien PJ, Hsieh KY, Chin KL, Huang WT, Tsao CY, Chen YF, Cheng HC, Chang HH. Research Center of Food and Biomolecules. A new immunomodulatory protein, designated ACA, was purified from the mycelium extract of Antrodia camphorata , a well-known folk medicine bitter mushroom in Taiwan, and N-terminally sequenced. By taking advantage of its N-terminal amino acid sequence, the full-length ACA gene was cloned using rapid amplification of cDNA ends (RACE) approach. This gene encodes a 136 amino acid protein that is homologous to the phytotoxic proteins from fungi. On the basis of the data of N-terminal sequencing and N-glycosidase F treatment, the native ACA was confirmed to be a glycoprotein. The similarity in activation of TLR4-deficient macrophages by both the native ACA and recombinant ACA (rACA) suggested that the glycosyl group(s) of the native ACA was insignificant in macrophage activation. Moreover, the failure of rACA to induce TLR2-deficient macrophages and to activate the RAW 264.7 macrophages transfected with the dominate-negative MyD88 (dnMyD88) indicated that the ACA-mediated macrophage activation was TLR2/MyD88 dependent. Microarray assay of the ACA-activated NFkappaB-related gene expression showed that rACA demonstrated a LPS-mimetic proinflammatory response toward RAW 264.7 macrophages. Furthermore, rACA enhanced phagocytosis activity and CD86 (B7-2) expression as well as induced TNF-alpha and IL-1beta production within murine peritoneal macrophages. A time-dependent induction of mRNA expression of cytokines TNF-alpha, IL-1beta, IL-6, and IL-12 as well as chemokines CCL3, CCL4, CCL5, and CCL10, but not IL-10, CCL17, CCL22, and CCL24, was observed after the ACA treatment of the macrophages. These results proposed that ACA exhibited M1 polarization and differentiation in macrophages. Thus, ACA is an important immunomodulatory protein of A. camphorata. PMID: 19371137 [PubMed - as supplied by publishe] 61. Cytotoxic triterpenes from Antrodia camphorata and their mode of action in HT-29 human colon cancer cells. Cancer Lett. 2009 May 23. [Epub ahead of print] Yeh CT, Rao YK, Yao CJ, Yeh CF, Li CH, Chuang SE, Luong JH, Lai GM, Tzeng YM. National Institute of Cancer Research, National Health Research Institutes, Zhunan 35053, Taiwan, ROC. Five lanostane (2, 3, 4, 6 and 8) and three ergostane-type (1, 5 and 7) triterpenes isolated from the fruiting bodies of Antrodia camphorata were evaluated for their in vitro cytotoxic data against various cancer cell types. The three zhankuic acids, 1, 5 and 7 displayed the most potent cytotoxic effect with an IC(50) value of 22.3-75.0muM. The compound 3 was selectively cytotoxic in three colon cancer cell lines (HT-29, HCT-116 and SW-480) and a breast cancer model (MDA-MB-231), whereas 8 only showed its cytotoxicity against MDA-MB-231. None of these isolates was toxic to mammary epithelial (MCF10A) and primary foreskin fibroblast (HS68) cells, two human normal cell lines. The compounds 1, 5 and 7 were also demonstrated to induce apoptosis in HT-29 and SW-480 cells, as confirmed by sub-G1 cell cycle arrest. In HT-29 cells, the expression of apoptosis-associated proteins poly-(ADP-ribose) polymerase cleavage, Bcl-2 and procaspase-3 were suppressed by compounds 1, 5 and 7. A mixture containing 4muM each of compounds 1, 5 and 7 also showed a synergistic cytotoxic effect in HT-29 cells. PMID: 19477064 [PubMed - as supplied by publisher 62. Cyclodextrin-modified capillary electrophoresis for achiral and chiral separation of ergostane and lanostane compounds extracted from the fruiting body of Antrodia camphorata. Electrophoresis. 2009 Jun 10;30(11):1967-1975. [Epub ahead of print] Majid E, Male KB, Tzeng YM, Omamogho JO, Glennon JD, Luong JH. Biotechnology Research Institute, National Research Council of Canada, Montreal, Quebec, Canada. A CD-modified capillary electrophoretic method has been developed for achiral and chiral analysis of seven bioactive compounds isolated from the fruiting body of Antrodia camphorata. Such important target analytes exhibit similar chemical structures and are known for their diverse properties including antioxidant and anticancer effects. The analytes were separated in 25 min using a pH 9.3, 20 mM sodium borate buffer containing 20 mM methyl-beta-CD and 30 mM sulfobutylether-beta-CD. With the exception of the optical isomer pairs (antcin B or zhankuic acid A, zhankuic acid C, and antcin A), the remaining bioactive compounds including the chiral pair antcin C were baseline-separated. Analysis time was noticeably longer to baseline separate all of the above chiral pairs ( approximately 38 min) by adding 5% DMF to the running buffer. The migration order was reversed compared with the HPLC elution. More hydrophobic compounds complexed favorably with methyl-beta-CD and emerged earlier in the electropherogram than their more hydrophilic 30

counterparts which were strongly associated with sulfobutylether-beta-CD. The simple capillary electrophoretic method developed was applicable for rapid separation and characterization of several important bioactive compounds isolated from the fruiting body of A. camphorata. PMID: 19517437 [PubMed - as supplied by publisher] 63. Review of Pharmacological Effects of Antrodia camphorata and its Bioactive Compounds. Evid Based Complement Alternat Med. 2009 Aug 17. [Epub ahead of print] Geethangili M, Tzeng YM. Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Wufeng, Taiwan, ROC. ymtzeng@cyut.edu.tw. Antrodia camphorata is a unique mushroom of Taiwan, which has been used as a traditional medicine for protection of diverse health-related conditions. In an effort to translate this Eastern medicine into Western-accepted therapy, a great deal of work has been carried out on A. camphorata. This review discusses the biological activities of the crude extracts and the main bioactive compounds of A. camphorata. The list of bioactivities of crude extracts is huge, ranging from anti-cancer to vasorelaxation and others. Over 78 compounds consisting of terpenoids, benzenoids, lignans, benzoquinone derivatives, succinic and maleic derivatives, in addition to polysaccharides have been identified. Many of these compounds were evaluated for biological activity. Many activities of crude extracts and pure compounds of A. camphorata against some major diseases of our time, and thus, a current review is of great importance. It is concluded that A. camphorata can be considered as an efficient alternative phytotherapeutic agent or a synergizer in the treatment of cancer and other immune-related diseases. However, clinical trails of human on A. camphorata extracts are limited and those of pure compounds are absent. The next step is to produce some medicines from A. camphorata, however, the production may be hampered by problems related to mass production. PMID: 19687189 [PubMed - as supplied by publisher] 64. Preferential blockade of dioxin-induced activation of the Aryl hydrocarbon receptor by Antrodia camphorata. Biol Pharm Bull. 2009 Sep;32(9):1510-5. Mukai M, Hayakawa K, Okamura M, Tagawa Y, Nakajima S, Saito Y, Takahashi S, Yao J, Nishimura D, Sugi M, Matsunaga M, Kitamura M. Department of Molecular Signaling, University of Yamanashi, Chuo, Japan. Halogenated and polycyclic aromatic hydrocarbons are widely distributed pollutants in environments. These toxic substances activate the aryl hydrocarbon receptor (AhR) and thereby cause a broad spectrum of pathological changes. Development of AhR inhibitors will be useful for prevention of diseases caused by AhR activation. Using the dioxin responsive element (DRE)-based sensing via secreted alkaline phosphatase (DRESSA), we examined effects of Antrodia camphorata, a mycerial extract, on the activation of AhR by halogenated and polycyclic aromatic hydrocarbons. We found that Antrodia camphorata markedly suppressed activation of AhR triggered by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In contrast, activation of AhR by polycyclic aromatic hydrocarbons (benzo[a]pyrene and 3-methylcholanthrene) was inhibited only modestly by this mycelium. Similarly, Antrodia camphorata only mildly attenuated activation of AhR by cigarette smoke that contains polycyclic aromatic hydrocarbons. Consistent with these results, Northern blot analysis revealed that DRE-driven exogenous and endogenous gene expression triggered by TCDD was abolished by Antrodia camphorata, whereas it did not substantially affect DRE-induced transcription triggered by benzo[a]pyrene, 3-methylcholanthrene or cigarette smoke. We also found that the inhibitory effect of Antrodia camphorata on TCDD-induced AhR activation was ascribed to neither down-regulation of AhR, down-regulation of the AhR nuclear translocator, nor up-regulation of the AhR repressor. These results suggest that Antrodia camphorata preferentially inhibits AhR activation and DRE-dependent gene expression triggered by dioxin. PMID: 19721224 [PubMed - in process] 65. Rapid Evaluation of Antrodia camphorata Natural Products and Derivatives in Tumourigenic Liver Progenitor Cells with a Novel Cell Proliferation Assay. ChemMedChem. 2009 Sep 3. [Epub ahead of print] Stewart SG, Ho LA, Polomska ME, Percival AT, Yeoh GC.

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The School of Biomedical, Biomolecular & Chemical Sciences, The University of Western Australia, Crawley, Crawley, WA 6009 (Australia), Fax: (+61) 8 6488 1005. We report the syntheses of five natural product maleimide and maleic anhydrides from the mushroom Antrodia camphorata. The ability of these compounds to affect proliferation in non-tumourigenic and tumourigenic liver progenitor cell lines was monitored by the Cellscreen system, a novel and nondestructive rapid-screening instrument. Additionally, a range of new aryl-functionalised differentiated derivatives were prepared through a Suzuki cross-coupling reaction to influence cell-growth effects. Several derivatives radically slowed the proliferation of liver progenitor cells; however, of particular interest were two maleic anhydride derivatives containing aryl tethers. These analogues demonstrated selectivity for limiting the proliferation of tumourigenic progenitor cells in comparison with their non-tumourigenic counterparts. Also highlighted is the application of the Cellscreen system in medicinal chemistry to rapidly measure the effect of compound libraries on cell proliferation. PMID: 19731279 [PubMed - as supplied by publisher] 66. Effects of Antrodia camphorata extracts on the viability, apoptosis, [Ca2+]i, and MAPKs phosphorylation of OC2 human oral cancer cells. Chin J Physiol. 2009 Jun 30;52(3):128-35. Huang CC, Cheng HH, Wang JL, Cheng JS, Chai KL, Fang YC, Kuo CC, Chu ST, Ho CM, Lin KL, Tsai JY, Jan CR. Department of Nursing, Tzu Hui Institute of Technology, Pingtung, Taiwan, Republic of China. The effect of Antrodia camphorata (AC) on human oral cancer cells has not been explored. This study examined the effect of AC on the viability, apoptosis, mitogen-activated protein kinases (MAPKs) phosphorylation and Ca2+ regulation of OC2 human oral cancer cells. AC at a concentration of 25 microM induced an increase in cell viability, but AC at concentrations > or = 50 microg/ml decreased viability in a concentration-dependent manner. AC at concentrations of 100-200 microg/ml induced apoptosis in a concentration-dependent manner as demonstrated by propidium iodide staining. AC (25 microg/ml) did not alter basal [Ca2+]i, but decreased the [Ca2+]i increases induced by ATP, bradykinin, histamine and thapsigargin. ATP, bradykinin, and histamine increased cell viability whereas thapsigargin decreased it. AC (25 microg/ml) pretreatment failed to alter ATP-induced increase in viability, potentiated bradykinin-induced increase in viability, decreased histamine-induced increase in viability and reversed thapsigargin-induced decrease in viability. Immunoblotting suggested that AC induced phosphorylation of ERK and JNK MAPKs, but not p38 MAPK. Collectively, for OC2 cells, AC exerted multiple effects on their viability and [Ca2+]i, induced their ERK and JNK MAPK phosphorylation, and probably evoked their apoptosis. PMID: 19777798 [PubMed - indexed for MEDLINE] 67. Protective effect of Antrodia Camphorata on bladder ischemia/reperfusion injury. Int Urol Nephrol. 2009 Sep 17. [Epub ahead of print] Juan YS, Mannikarottu A, Chuang SM, Li S, Lin AD, Chang-Chou L, Schuler C, Leggett RE, Levin RM. Department of Urology, Kaohsiung Municipal Hsiao-Kang Hospital, Kaohsiung, Taiwan. INTRODUCTION: Obstructive bladder dysfunction is directly related to ischemia/reperfusion injury characterized by damage to nerves, synapses and smooth muscle cells within the bladder wall. Antrodia Camphorata (AC) has significant antioxidant, antiinflammatory and cell-cycle inhibition properties. The specific aim of this study was to evaluate whether orally administered AC can protect rabbit bladders from the progressive dysfunctions induced by bilateral ischemia/reperfusion (I/R). METHODS: Twenty-four male NZW rabbits were separated into 4 groups of 6 animals each. Rabbits in groups 1 and 2 were fed Antrodia Camphorata (AC) suspensions; those in groups 3 and 4 received vehicle. Each rabbit in groups 2 and 4 were subjected to in vivo bilateral ischemia for 2 h and then allowed to recover for 1 week. The rabbits in groups 1 and 3 received sham operation and served as control groups. Cystometry, contractile responses to field stimulation, carbachol, ATP and KCl were determined. Biochemical and immuno-histochemical studies were also performed. RESULTS: I/R resulted in decreased compliance, decreased contractile responses, decreased nerve density, and increased apoptosis. AC pretreatment of rabbits subjected to I/R significantly protected the bladder from all contractile, biochemical, and structural dysfunctions resulting in significantly improved bladder. PMID: 19760512 [PubMed - as supplied by publisher]

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68. Taiwanofungus camphoratus activates peroxisome proliferator-activated receptors and induces hypotriglyceride in hypercholesterolemic rats. Biosci Biotechnol Biochem. 2008 Jul;72(7):1704-13. Epub 2008 Jul 7. Suk FM, Lin SY, Chen CH, Yen SJ, Su CH, Liu DZ, Hou WC, Hung LF, Lin PJ, Liang YC. Department of Internal Medicine, Taipei Medical University Hospital & Wan-Fang Hospital, Taipei, 11014, Taiwan. Taiwanofungus camphoratus (T. camphoratus), a fungus and a Taiwan-specific, well-known traditional Chinese medicine, has long been used to treat diarrhea, hypertension, itchy skin, and liver cancer. To gain a large amount of T. camphoratus, several culture techniques have been developed, including solid-state culture and liquid-state fermentation. Peroxisome proliferator-activated receptor gamma (PPARgamma) has been described as a hypoglycemic agent that increases insulin sensitivity in peripheral tissues and results in reduced blood glucose, insulin, and triglyceride levels in insulin-resistant animals and in type-2 (non-insulin-dependent) diabetic patients. In this study, we investigate the possibility that T. camphoratus might activate PPARgamma in vitro and hypolipidemic activity in vivo. The results show that an aqueous extract of the wild fruiting bodies of T. camphoratus was able to increase the PPARgamma activity in cells transfected with the PPARgamma expression plasmid and the AOx-TK reporter plasmid. Based on the cell experiment, we examined the hypolipidemic effect of wild fruiting bodies (WFT) and a solid-state culture (SST) of T. camphoratus on SD rats fed on a high-cholesterol (HC) diet. The results show that WFT significantly decreased the serum triglyceride level, but could not affect the cholesterol level. SST only slightly decreased the serum triglyceride level. In addition, both WFT and SST significantly decreased the serum alanine transaminase (ALT) level and protected against the liver damage induced by the HC diet from the results of a histological examination. These results suggest that T. camphoratus might contain PPARgamma ligands and result in a hypotriglyceridemic effect, and that it also exhibits a liver protective activity. PMID: 18603804 [PubMed - indexed for MEDLINE] 69. Cloning, expression, and characterization of an enzyme possessing both glutaredoxin and dehydroascorbate reductase activity from Taiwanofungus camphorata. J Agric Food Chem. 2009 Nov 11;57(21):10357-62. Ken CF, Lin CY, Jiang YC, Wen L, Lin CT. Institute of Biotechnology, National Changhua University of Education, Changhua 500 Taiwan.Glutaredoxins (Grxs) play important roles in the reduction of disulfides via reduced glutathione as a reductant. A cDNA (503 bp, EU193660) encoding a putative Grx was cloned from Taiwanofugus camphorata (Tc). The deduced amino acid sequence is conserved among the reported dithiol Grxs. A 3D homology structure was created for this TcGrx. To characterize the TcGrx enzyme, the coding region was subcloned into an expression vector pET-20b(+) and transformed into Escherichia coli . Functional TcGrx was expressed and purified by Ni(2+)-nitrilotriacetic acid Sepharose. The purified enzyme showed bands of approximately 15 kDa on 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The TcGrx encodes a protein possessing both Grx and dehydroascorbate reductase (DHAR) activity. The Michaelis constant (K(m)) values for beta-hydroxyethyl disulfide (HED) and dehydroascorbate (DHA) were 0.57 and 1.85 mM, respectively. The half-life of deactivation of the protein at 100 degrees C was 8.5 min, and its thermal inactivation rate constant K(d) was 6.52 x 10(-2) min(-1). The enzyme was active under a broad pH range from 6.0 to 10.0 and in the presence of imidazole up to 0.4 M. The enzyme was susceptible to SDS denaturation and protease degradation/inactivation. PMID: 19886686 [PubMed - in process] 70. Inhibition of Anchorage-Independent Proliferation and G0/G1 Cell-Cycle Regulation in Human Colorectal Carcinoma Cells by 4,7-Dimethoxy-5-methyl-l,3-benzodioxole Isolated from the Fruiting Body of Antrodia camphorate. Evid Based Complement Alternat Med. 2009 Mar 17. [Epub ahead of print] Lien HM, Lin HW, Wang YJ, Chen LC, Yang DY, Lai YY, Ho YS. Taipei Medical University, No. 250, Wu-Hsing Street, Taipei 110, Taiwan. Tel: +886-2-27361661. Ext. 3327; hoyuansn@tmu.edu.tw or Ya-Yun Lai, Department of Applied Chemistry, Chung Shan Medical University, No. 110, Sec. 1, Jianguo N. Rd., Taichung 402, Taiwan. yayun819@csmu.edu.tw. In this study, 4,7-dimethoxy-5-methyl-l,3-benzodioxole (SY-1) was isolated from three different sources of dried fruiting bodies of Antrodia camphorate (AC). AC is a medicinal mushroom that grows on the inner heartwood wall of Cinnamomum kanehirai Hay (Lauraceae), an endemic species that is used in Chinese medicine for its anti-tumor and 33

immunomodulatory properties. In this study, we demonstrated that SY-1 profoundly decreased the proliferation of human colon cancer cells (COLO 205) through G0/G1 cell-cycle arrest (50-150 muM) and induction of apoptosis (>150 muM). Cell-cycle arrest induced by SY-1 was associated with a significant increase in levels of p53, p21/Cip1 and p27/Kip1, and a decrease in cyclins D1, D3 and A. In contrast, SY-1 treatment did not induce significant changes in G0/G1 phase cell-cycle regulatory proteins in normal human colonic epithelial cells (FHC). The cells were cultured in soft agar to evaluate anchorage-independent colony formation, and we found that the number of transformed colonies was significantly reduced in the SY-1-treated COLO 205 cells. These findings demonstrate for the first time that SY-1 inhibits human colon cancer cell proliferation through inhibition of cell growth and anchorage-independent colony formation in soft agar. However, the detailed mechanisms of these processes remain unclear and will require further investigation. PMID: 19293251 [PubMed - as supplied by publisher] 71. The augmented anti-tumor effects of Antrodia camphorata co-fermented with Chinese medicinal herb in human hepatoma cells. Am J Chin Med. 2009;37(4):771-83. Li SL, Huang ZN, Hsieh HH, Yu WC, Tzeng WY, Lee GY, Chen YP, Chang CY, Chuu JJ. Institute of Biotechnology, College of Engineering, Southern Taiwan University, Yung-Kang City, Tainan, Taiwan. Antrodia camphorata, unique fungal specie, has been used as a folk medicine in Taiwan for many years. The purpose of this study was to compare the extracts from the solid-state culture of A. camphorata co-fermented with Chinese medicinal herb (AC-CF) with two other extracts from fruiting bodies (AC-FB) or solid-state culture (AC-SS), for their anti-tumor effects in human hepatoma HepG2 cells. We measured in vitro cell proliferation, percentage of apoptosis, population distribution of cell cycles, Western blot analysis of multiple drugs resistance-1 (MDR-1), and apoptosis-related proteins in HepG2 cells treated with three different preparations of A. camphorate extracts. Our results showed that AC-CF had better anti-proliferation effect on human hepatoma HepG2 cells than AC-FB or AC-SS dose-dependently. In addition, AC-CF in combination with anti-tumor agents (mitomycin C or methotrexate) showed better adjuvant anti-tumor effects than AC-FB or AC-SS. We further demonstrated the augmented adjuvant anti-tumor effects of AC-CF not only through down regulation of MDR-1 expression but also through a COX-2 dependent apoptosis pathway, involving down-regulation of COX-2 and p-AKT and up-regulation of PARP-1. In conclusion, in this study, we have demonstrated a novel strategy of fermenting A. camphorata with Chinese medicinal herb (AC-CF), which augmented their anti-tumor effects in human hepatoma HepG2 cells as compared to the traditional ones (AC-FB or AC-SS). PMID: 19655414 [PubMed - indexed for MEDLINE] 72. Antroquinonol displays anticancer potential against human hepatocellular carcinoma cells: a crucial role of AMPK and mTOR pathways. Biochem Pharmacol. 2010 Jan 15;79(2):162-71. Epub 2009 Aug 31. Chiang PC, Lin SC, Pan SL, Kuo CH, Tsai IL, Kuo MT, Wen WC, Chen P, Guh JH. School of Pharmacy, National Taiwan University, No. 1, Sect. 1, Jen-Ai Rd, Taipei 100, Taiwan. 5'AMP-activated protein kinase (AMPK) and the mammalian target of rapamycin (mTOR) are two serine/threonine protein kinases responsible for cellular energy homeostasis and translational control, respectively. Evidence suggests that these two kniases are potential targets for cancer chemotherapy against hepatocellular carcinoma (HCC). Antroquinonol that is isolated from Antrodia camphorate, a well-known Traditional Chinese Medicine for treatment of liver diseases, displayed effective anticancer activity against both HBV DNA-positive and -negative HCC cell lines. The rank order of potency against HCCs is HepG2>HepG2.2.15>Mahlavu>PLC/PRF/5>SK-Hep1>Hep3B. Antroquinonol completely abolished cell-cycle progression released from double-thymidine-block synchronization and caused a subsequent apoptosis. The data were supported by down-regulation and reduced nuclear translocation of G1-regulator proteins, including cyclin D1, cyclin E, Cdk4 and Cdk2. Further analysis showed that the mRNA expressions of the G1-regulator proteins were not modified by antroquinonol, indicating an inhibition of translational but not transcriptional levels. Antroquinonol induced the assembly of tuberous sclerosis complex (TSC)-1/TSC2, leading to the blockade of cellular protein synthesis through inhibition of protein phosphorylation including mTOR (Ser(2448)), p70(S6K) (Thr(421)/Ser(424) and Thr(389)) and 4E-BP1 (Thr(37)/Thr(46) and Thr(70)). Furthermore, the AMPK activity was elevated by antroquinonol. Compound C, a selective AMPK inhibitor, significantly reversed antroquinonol-mediated effects suggesting the crucial role of AMPK. Besides, the loss of mitochondrial membrane potential and depletion of mitochondrial content indicated the mitochondrial stress caused by 34

antroquinonol. In summary, the data suggest that antroquinonol displays anticancer activity against HCCs through AMPK activation and inhibition of mTOR translational pathway, leading to G1 arrest of the cell-cycle and subsequent cell apoptosis. PMID: 19723512 [PubMed - in process] 73. Dehydroeburicoic acid induces calcium- and calpain-dependent necrosis in human U87MG glioblastomas. Chem Res Toxicol. 2009 Nov;22(11):1817-26. Deng JY, Chen SJ, Jow GM, Hsueh CW, Jeng CJ. Institute of Anatomy and Cell Biology, School of Medicine, National Yang-Ming University, No. 155, Section 2, Li-Non Street, Taipei 12212, Taiwan. Dehydroeburicoic acid (DeEA) is a triterpene purified from medicinal fungi such as Antrodia camphorate, the crude extract of which is known to exert cytotoxic effects against several types of cancer cells. We aim to test the hypothesis that DeEA possesses significant cytotoxic effects against glioblastomas, one of the most frequent and malignant brain tumors in adults. 3-(4,5-Dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase release assays indicated that DeEA inhibited the proliferation of the human glioblastoma cell U87MG. In addition, Annexin V and propidium iodide staining showed that DeEA treatment led to a rapid increase of glioblastomas in the necrotic/late apoptotic fraction, whereas cell cycle analysis revealed that DeEA failed to significantly enhance the population of U87MG cells in the hypodiploid (sub-G1) fraction. Using electron microscopy, we found that DeEA induced significant cell enlargements, massive cytoplasmic vacuolization, and loss of mitochondrial membrane integrity. DeEA treatment triggered an intracellular Ca(2+) increase, and DeEA-induced cell death was significantly attenuated by BAPTA-AM but not ethylenediaminetetraacetic acid or ethylene glycol tetraacetic acid. DeEA instigated a reduction of both mitochondrial transmembrane potential and intracellular ATP level. Moreover, DeEA induced proteolysis of alpha-spectrin by calpain, and DeEA cytotoxicity in U87MG cells was caspase-independent but was effectively blocked by calpain inhibitor. Interestingly, DeEA also caused autophagic response that was prevented by calpain inhibitor. Taken together, these results suggest that in human glioblastomas, DeEA induces necrotic cell death that involves Ca(2+) overload, mitochondrial dysfunction, and calpain activation. PMID: 19848398 [PubMed - in process] 74. Administration of polysaccharides from Antrodia camphorata modulates dendritic cell function and alleviates allergen-induced T helper type 2 responses in a mouse model of asthma. Immunology. 2009 Aug 17. [Epub ahead of print] Liu KJ, Leu SJ, Su CH, Chiang BL, Chen YL, Lee YL. School of Medical Laboratory Science and Biotechnology, Taipei Medical University, Taipei, Taiwan. Summary Asthma is a chronic disease characterized by airway inflammation caused by the dysregulated production of cytokines secreted by allergen-specific type 2 T helper (Th2) cells. Antrodia camphorata is a commonly used fungus in Asian folk medicine, and A. camphorata polysaccharides are reported to possess anti-cancer activities. In this study, the immunomodulatory effects of purified fractionated polysaccharides (GF2) from A. camphorata on dendritic cells (DCs) and their potential preventive effects against ovalbumin (OVA) -induced asthma were investigated. In the presence of GF2, lipopolysaccharide (LPS) -activated DCs exhibited up-regulated expression of major histocompatibility complex (MHC) class II and co-stimulatory molecules, as well as enhanced interleukin-10 (IL-10) and IL-12 production. GF2 treatment on LPS-activated DCs suppressed na CD4(+) T-cell proliferation and Th2 cell polarization with IL-10 production in an ve allogeneic mixed lymphocyte reaction. In animal experiments, a high dose of GF2 efficiently reduced expression levels of OVA-specific immunoglobulin G1 (IgG1) and IgE. However, lower doses of GF2 significantly enhanced OVA-specific IgG2a production. Our data also showed that administration of GF2 dose-dependently inhibited the development of airway hyperresponsiveness, airway eosinophilia and Th2 responses. OVA-specific CD4(+) T cells from higher doses of GF2-treated mice had significantly lower proliferative capacities compared with control mice. Moreover, treatment with GF2 significantly increased the high levels of IL-10 and low levels of interferon-gamma produced by T cells. Taken together, these data indicate that administration of A. camphorata polysaccharides (GF2) may have therapeutic potential when used as an adjuvant for the immunomodulatory treatment of allergic asthma. PMID: 19909376 [PubMed - as supplied by publisher]

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75. Apoptotic effects of a high performance liquid chromatography (HPLC) fraction of Antrodia camphorata mycelia are mediated by down-regulation of the expressions of four tumor-related genes in human non-small cell lung carcinoma A549 cell. J Ethnopharmacol. 2009 Dec 6. [Epub ahead of print] Chan YY, Chang CS, Chien LH, Wu TF. Department of Biotechnology, Southern Taiwan University, 1 Nan-Tai Street, YungKang City, Tainan County 701, Taiwan. AIM OF THE STUDY: Antrodia camphorata (niu-chang-chih) is a fungus native to Taiwan which is believed to be effective in preventing diseases. Recent reports demonstrate that Antrodia camphorata products induce the apoptosis of various kinds of tumor cells. In this study we determined the inhibitory effects of alcohol extract and individual fractions of alcohol extract on the proliferation of human non-small cell lung carcinoma A549 cell and clarified the mechanism underlying the anti-cancer activities. MATERIALS AND METHODS: Alcohol extracts of Antrodia camphorata mycelia were prepared by the serial extraction with the solvents with increasing polarity and fractionated using HPLC. Cell viability was determined by MTT assay. Apoptosis detection was carried out by subG(1) analysis and annexin V/propidium iodide staining using flow cytometry. The impacts of HPLC fractions on the expression levels of apoptosis- and cancer-related proteins were evaluated by western blotting. RESULTS: Three HPLC fractions, fractions 5-7, had robust inhibition of human A549 cells and among them fraction 6 (Fr-6) possessed the most potent effectiveness. Apoptotic assay showed that Fr-6-induced human A549 cell apoptosis by triggering the mitochondrial pathway and endothelium reticulum (ER) stress. Immunoblotting results demonstrated that Fr-6 possibly activated ER stress by lowering the expression level of calpain 1/2 small subunit and Fr-6-mediated decrease in cell proliferation might attribute to the suppressive effect on the Erk 1/2 pathway, which arose from Fr-6-derived low galectin-1 expression. Furthermore Fr-6 could diminish Rho GDP dissociation inhibitor alpha (RhoGDI-alpha) expression and subsequently activated c-Jun NH(2)-terminal kinase (JNK) pathway, which is linked to cell apoptosis. Fr-6 also could decrease the production level of eukaryotic translation initiation factor 5A, which is a potential cancer intervention target. CONCLUSION: These results suggested that the anti-cancer activity of Antrodia camphorata might be due to multiple active metabolites, which work together to induce cell apoptosis via various pathways. Copyright 2009 Elsevier Ireland Ltd. All rights reserved. PMID: 19995598 [PubMed - as supplied by publisher] 76. Antrocamphin A, an Anti-inflammatory Principal from the Fruiting Body of Taiwanofungus camphoratus , and Its Mechanisms. J Agric Food Chem. 2010 Feb 3. [Epub ahead of print] Hsieh YH, Chu FH, Wang YS, Chien SC, Chang ST, Shaw JF, Chen CY, Hsiao WW, Kuo YH, Wang SY. Graduate Institute of Biotechnology, National Chung-Hsing University, Taichung, Taiwan. The fungus Taiwanofungus camphoratus is commonly used for medicinal purposes in Taiwan. It is used as a detoxicant for food poisoning and considered to be a precious folk medicine for hepatoprotection and anti-inflammation. In this study, a lipopolysaccaride (LPS)-challenged ICR mouse acute inflammation model and a LPS-induced macrophage model were used to evaluate the anti-inflammatory activity of T. camphoratus. Ethanol extract of T. camphoratus significantly inhibited expression of iNOS and COX-2 in the liver of LPS-challenged acute inflammatory mice. The ethyl acetate fraction and its isolated compound, antrocamphin A, significantly suppressed nitrite/nitrate concentration in LPS-challenged RAW 264.7 cells. Antrocamphin A showed potent anti-inflammatory activity by suppressing pro-inflammatory molecule release via the down-regulation of iNOS and COX-2 expression through the NF-kappaB pathway. This study, therefore, first demonstrates the bioactive compound of T. camphoratus and illustrates the mechanism by which it confers its anti-inflammatory activity. PMID: 20192205 [PubMed - as supplied by publisher] 77. Fruiting Body of Niuchangchih (Antrodia camphorata) Protects Livers against Chronic Alcohol Consumption Damage. J Agric Food Chem. 2010 Mar 1. [Epub ahead of print] Huang CH, Chang YY, Liu CW, Kang WY, Lin YL, Chang HC, Chen YC. Department of Biomedical Engineering, National Cheng Kung University, Tainan City 701, Taiwan. Abstract: An alcoholic fatty liver disease was induced by drinking water containing 20% (w/w) alcohol. Therapeutic groups were orally administrated dosages of 0.25 g silymarin/kg body weight (BW) and a low dosage of Niuchangchih (Antrodia camphorata) (0.025 g/kg BW) and a high dosage of Niuchangchih (0.1 g/kg BW) per day. Niuchangchih, 36

especially at the high dosage, not only showed a hypercholesterolemic effect (p < 0.05) but also reduced (p < 0.05) hepatic lipids in alcohol-fed rats. Those beneficial effects could be partially attributed to higher (p < 0.05) fecal cholesterol and bile acid outputs, as well as downregulations (p < 0.05) of 3-hydroxy-3-methylglutaryl-CoA reductase, sterol regulatory element-binding protein-1c, acetyl-CoA carboxylase, fatty acid synthase, and malic enzyme gene expressions; meanwhile, there was an upregulation of low-density lipoprotein receptor and peroxisome proliferator-activated alpha gene expression. Besides, Niuchangchih also enhanced (p < 0.05) the liver glutathione, Trolox equivalent antioxidant capacity, and activities of superoxide dismutase, catalase, and glutathione peroxidase and decreased the liver malondialdehyde content, which also partially contributed to the lowered (p < 0.05) serum aspartate aminotransferase levels and no observed lesion in the histological examination of alcohol-fed rats. PMID: 20128588 [PubMed - as supplied by publisher] 78. Quality evaluation of mycelial Antrodia camphorata using high-performance liquid chromatography (HPLC) coupled with diode array detector and mass spectrometry (DAD-MS). Chin Med. 2010 Jan 29;5:4. Zhao SS, Leung KS. Department of Chemistry, Hong Kong Baptist University, Kowloon, Hong Kong SAR, China. s9362284@hkbu.edu.hk. ABSTRACT: BACKGROUND: Antrodia camphorata (AC) is an important fungus native to Taiwanese forested regions. Scientific studies have demonstrated that extracts of AC possess a variety of pharmacological functions. This study aims to identify the full profile fingerprint of nucleosides and nucleobases in mycelial AC and to assess the quality of two commercial mycelial AC products. METHODS: High-performance liquid chromatography coupled with diode array detector and mass spectrometry was employed to identify the major components in mycelial AC. The chemical separation was carried out using a gradient program on a reverse phase Alltima C18 AQ analytical column (250 x 4.6 mm, 5 mum) with the mobile phase consisting of deionized water and methanol. RESULTS: Ten nucleosides and nucleobases, two maleimide derivatives, and a sterol were identified as the major constituents in mycelial AC. These groups of chemical compounds constitute the first chromatographic fingerprint as an index for quality assessment of this medicinal fungus. CONCLUSIONS: This study provides the first chromatographic fingerprint to assess the quality of mycelial AC. PMID: 20205903 [PubMed - in process] 79. Cloning, expression, and characterization of a thioredoxin reductase cDNA from Taiwanofungus camphorata J Agric Food Chem. 2010 Apr 28;58(8):4825-30. Huang CY, Ken CF, Chi HH, Wen L, Lin CT. Institute of Bioscience and Biotechnology and Marine Center for Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 202, Taiwan. Abstract: A cDNA encoding putative thioredoxin reductase (TR) was identified from a medicinal mushroom, Taiwanofungus camphorata (T. camphorata). Alignment of the deduced amino acid sequence with TRs from other organisms showed high levels of identity (59-74%). A three-dimensional (3-D) homology structure was created for this TR. Functional T. camphorata TR (TcTR) was overexpressed in yeast and purified. The purified enzyme showed a monomic form on a 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme's half-life of deactivation at 60 degrees C was 12.9 min, and its thermal inactivation rate constant K(d) was 5.37 x 10(-2) min(-1). The optimal pH for the enzyme was pH 8 and retained about 76% activity in the presence of 0.1 M imidazole. The enzyme showed 50% activity after 10 min of incubation at 37 degrees C with chymotrypsin. The Michaelis constant (K(m)) value for dithionitrobenzoate (DTNB) was 1.59 mM. PMID: 20307095 [PubMed - in process] 80. Promoting effect of Antrodia camphorata as an immunomodulating adjuvant on the antitumor efficacy of HER-2/neu DNA vaccine. Cancer Immunol Immunother. 2010 Apr 14. [Epub ahead of print] Huang CH, Chang CC, Lin CM, Wang ST, Wu MT, Li EI, Chang HC, Lin CC. Institute of Biomedical Engineering, National Cheng Kung University, Tainan, Taiwan, ROC. Abstract: It is well known that DNA vaccines induce protective humoral and cell-mediated immune responses in several animal models. Antrodia camphorata (AC) is a unique basidiomycete fungus of the Polyporaceae family that only grows on 37

the aromatic tree Cinnamomum kanehirai Hayata (Lauraceae) endemic to Taiwan. Importantly, AC has been shown to be highly beneficial in the treatment and prevention of cancer. The goal of this study was to investigate whether AC is able to augment the antitumor immune properties of a HER-2/neu DNA vaccine in a mouse model in which p185neu is overexpressed in MBT-2 tumor cells. Compared with the mice that received the HER-2/neu DNA vaccine alone, co-treatment with AC suppressed tumor growth and extended the survival rate. This increase in the antitumor efficacy was attributed to the enhancement of the Th1-like cellular immune response by the HER-2/neu DNA vaccine-AC combination. Evidence for this came from the marked increase in the IFN-gamma mRNA expression in CD4(+) T cells in the draining inguinal lymph nodes, an increase in the number of functional HER-2/neu-specific CTLs, and the increased tumor infiltration of both CD4(+) and CD8(+) T cells, depletion of which abolishes the antitumor effect of the HER-2/neu DNA vaccine-AC therapy. Our results further indicate that the treatment of mice with AC enhanced DC activation and production of Th1-activating cytokines (e.g. IL-12, and IFN-alpha) in the draining lymph nodes, which were sufficient to directly stimulate T cell proliferation and higher IFN-gamma production in response to ErbB2. Overall, these results clearly demonstrate that AC represents a promising immunomodulatory adjuvant that could enhance the therapeutic potency of HER-2/neu DNA vaccines in cancer therapy. PMID: 20390417 [PubMed - as supplied by publisher] 81. Analgesic Effects and the Mechanisms of Anti-inflammation of Ergostatrien-3beta-ol from Antrodia camphorata Submerged Whole Broth in Mice. J Agric Food Chem. 2010 May 27. [Epub ahead of print] Huang GJ, Huang SS, Lin SS, Shao YY, Chen CC, Hou WC, Kuo YH. Institute of Chinese Pharmaceutical Sciences, China Medical University, Taichung 404, Taiwan, Republic of China. Abstract: Ergostatrien-3beta-ol (ST1), an active and major ingredient from Antrodia camphorata (AC) submerged whole broth was evaluated for the analgesic and anti-inflammatory effects. Treatment of male imprinting control region (ICR) mice with ST1 (1, 5, and 10 mg/kg) significantly inhibited the numbers of acetic-acid-induced writhing response in 10 min. Also, our result showed that ST1 (10 mg/kg) significantly inhibited the formalin-induced pain in the late phase (p < 0.001). In the anti-inflammatory test, ST1 (10 mg/kg) decreased the paw edema at 4 and 5 h after lambda-carrageenin (Carr) administration and increased the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the liver tissue. We also demonstrated that ST1 significantly attenuated the malondialdehyde (MDA) level in the edema paw at 5 h after Carr injection. ST1 (1, 5, and 10 mg/kg) decreased the nitric oxide (NO) levels on both the edema paw and serum level at 5 h after Carr injection. Also, ST1 (5 and 10 mg/kg) diminished the serum tumor necrosis factor (TNF-alpha) at 5 h after Carr injection. Western blotting revealed that ST1 (10 mg/kg) decreased Carr-induced inducible nitric oxide synthase (iNOS), and cycloxyclase (COX-2) expressions at 5 h in the edema paw. An intraperitoneal (ip) injection treatment with ST1 also diminished neutrophil infiltration into sites of inflammation, as did indomethacin (Indo). The anti-inflammatory mechanisms of ST1 might be related to the decrease in the level of MDA, iNOS, and COX-2 in the edema paw via increasing the activities of CAT, SOD, and GPx in the liver through the suppression of TNF-alpha and NO. PMID: 20507140 [PubMed - as supplied by publisher] 82. Development of a LC-MS/MS method for the determination of antrodin B and antrodin C from Antrodia camphorata extract in rat plasma for pharmacokinetic study. J Pharm Biomed Anal. 2010 Jun 11. [Epub ahead of print] Liu Y, Di X, Liu X, Shen W, Leung KS. School of Pharmacy, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang 110016, PR China; Department of Chemistry, Hong Kong Baptist University, 224 Waterloo Road, Kowloon Tong, Hong Kong, PR China. Abstract: A selective and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the determination of antrodin B and antrodin C in rat plasma. Both target compounds, together with the internal standard (diazepam), were extracted from rat plasma samples by liquid-liquid extraction with ethyl acetate. Chromatographic separation was carried out on an Agilent XDB-C(8) column with an isocratic mobile phase consisting of acetonitrile and water (70:30, V/V) at a flow rate of 0.5mL/min. The mass spectrometric detection was performed by selected reaction monitoring (SRM) mode via atmospheric pressure chemical ionization (APCI) source operating in positive ionization mode. The assay exhibited a linear dynamic range of 47.6-4760ng/mL for antrodin B and 56.6-5660ng/mL for antrodin C. The intra- and inter-day precision was less than 5.3% and the accuracy was less than 2.7% for both analytes. The 38

validated method has been applied to the pharmacokinetic study of antrodin B and antrodin C in rats following oral administration of Antrodia camphorata extract. Copyright 2010. Published by Elsevier B.V. PMID: 20542396 [PubMed - as supplied by publisher] 83. Methyl Antcinate A from Antrodia camphorata Induces Apoptosis in Human Liver Cancer Cells through Oxidant-Mediated Cofilin- and Bax-Triggered Mitochondrial Pathway. Chem Res Toxicol. 2010 Jun 17. [Epub ahead of print] Hsieh YC, Rao YK, Wu CC, Huang CY, Geethangili M, Hsu SL, Tzeng YM. Department of Education and Research, Taichung Veterans General Hospital, Taichung, Taiwan, Republic of China, Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Wufeng, Taiwan, Republic of China, Institute of Medical and Molecular Toxicology, Chung Shan Medical University, Taichung, Taiwan, Republic of China, and Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan, Republic of China. Abstract: We investigated the effects of antcin A, antcin C, and methyl antcinate A (MAA) isolated from Antrodia camphorata on the proliferation of human liver cancer cell lines Huh7, HepG2, and Hep3B and the normal cell rat hepatocytes. The three compounds selectively inhibit the proliferation of tumor cells rather than normal cells, with IC(50) values ranging from 30.2 to 286.4 muM. The compound MAA was a more potent cytotoxic agent than antcins A and C with IC(50) values of 52.2, 78.0, and 30.2 muM against HepG2, Hep3B, and Huh7 cells, respectively. To elucidate the molecular mechanism, treatment of Huh7 cells with 100 muM MAA induced an apoptotic cell death, which was characterized by the appearance of sub-G1 population, DNA fragmentation, TUNEL positive cells, and caspase activation. MAA triggered the mitochondrial apoptotic pathway, as indicated by an increase in the protein expression of Bax, Bak, and PUMA, as well as a decrease in Bcl-(XL) and Bcl-2 and disruption of mitochondrial membrane potential and promotion of mitochondrial cytochrome c release, as well as activation of caspases-2, -3, and -9. We also found that pretreatment with inhibitors of caspases-2, -3, and -9 noticeably blocked MAA-triggered apoptosis. Furthermore, intracellular reactive oxygen species (ROS) generation and NADPH oxidase activation were observed in MAA-stimulated Huh7 cells. Mechanistic studies showed that MAA induces mitochondrial translocation of cofilin. When Huh7 cells were treated with cyclosporine A and bongkrekic acid, an inhibitor of the mitochondria permeability transition pore, the levels of cell death induced by MAA were significantly attenuated. Additionally, pretreatment of Huh7 cells with antioxidants ascorbic acid and N-acetyl cysteine markedly attenuated the MAA-induced apoptosis by upregulation of Bax, Bak, and PUMA, mitochondrial translocation of cofilin, activation of caspase-3, and cell death. Taken together, our results provide the first evidence of the activation of the ROS-dependent cofilin- and Bax-triggered mitochondrial pathway as a critical mechanism of MAA-induced cell death in liver cancer cells. PMID: 20557081 [PubMed - as supplied by publisher] 84. Dietary effect of Antrodia Camphorate extracts on immune responses in WEHI-3 leukemia BALB/c mice. Lin SY, Sheen LY, Chiang BH, Yang JS, Pan JH, Chang YH, Hsu YM, Chiang JH, Lu CC, Wu CL, Chung JG. Hung-Kuang University, Sha Lu, Taichung, Taiwan. Nutr Cancer. 2010 Jul;62(5):593-600. Abstract: Antrodia camphorata has been recognized to be a traditional Chinese medicine for abdominal pain, diarrhea, and to protect against hepatitis virus infection. Several ingredients derived from A. camphorata possess various pharmacological and biological activities such as antioxidant and anticancer. In this study, its ability to promote immune responses and to exhibited antileukemia activity in WEHI-3 leukemia BALB/c mice were investigated. The results indicated A. camphorata significantly prolonged the survival rate and prevented the body weight loss in leukemia mice. Four mg/kg of A. camphorata treatment significantly decreased the weight of the spleen. Both doses (2 and 4 mg/kg) of A. camphorata did not affect Mac-3 marker in leukocytes. However, the 4 mg/kg of A. camphorata decreased the levels of CD11b and both doses of treatment increased CD3 and CD19. With lipopolysaccharide stimulation, the 4 mg/kg of A. camphorata promoted the significant proliferation of leukocytes; but with concanavalin A stimulation, both doses promoted the significant proliferation of leukocytes. YAC-1 target cells were killed by NK cells from the mice after treatment with A. camphorata at 4 mg/kg in target cells at a ratio of 50:1. The percentage of macrophages with phagocyted at A. camphorata treatment increased, and these effects were in dose-dependent manners. PMID: 20574920 [PubMed - in process]

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85. Further studies on the hepatoprotective effect of Antrodia camphorata in submerged culture on ethanol-induced acute liver injury in rats. Lu ZM, Tao WY, Xu HY, Ao ZH, Zhang XM, Xu ZH. Nat Prod Res. 2010 Jul 9:1-12. [Epub ahead of print] Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, PR China. Abstract: To further understand the hepatoprotective activity of Antrodia camphorata in living systems and the possible mechanisms of this protection, the effects of fractions from A. camphorata in submerged culture on the liver and its antioxidative system in acute ethanol intoxicated rats were investigated. The results showed that the ethanolic extract (Fr-I) of A. camphorata was the most effective in the prevention of ethanol-induced acute liver injury and free radical generation in rats. The ethanolic extract administrated prior to ethanol significantly prevented the increase in serum levels of hepatic enzyme markers such as aspartate aminotransferase and alanine aminotransferase. It also normalised the increase of hepatic malondialdehyde concentration and the decrease of glutathione levels in the liver. Moreover, Fr-I improved the ethanol-induced decrease of hepatic glutathione peroxidase and reductase activities. On the basis of these results, the ethanolic extract of A. camphorata may exert its hepatoprotective activity by up-regulating GSH-dependent enzymes and inhibiting free radical formation in the liver. PMID: 20623423 [PubMed - as supplied by publisher] 86. Antrodia camphorata suppresses lipopolysaccharide-induced nuclear factor-kappaB activation in transgenic mice evaluated by bioluminescence imaging. Hseu YC, Huang HC, Hsiang CY. Food Chem Toxicol. 2010 August - September;48(8-9):2319-2325. Epub 2010 Jun 1. Department of Cosmeceutics, China Medical University, Taichung, Taiwan. Abstract: In an earlier study, we found that Antrodia camphorata inhibited the production of lipopolysaccharide (LPS)-induced cytokines, inducible nitric oxide synthase, and cyclooxygenase-2 by blocking nuclear factor-kappaB (NF-kappaB) activation in cultured RAW 264.7 macrophages. This study was aimed at evaluating the inhibitory effects of the fermented culture broth of A. camphorata in terms of LPS-induced NF-kappaB activation in transgenic mice by using a non-invasive, real-time NF-kappaB bioluminescence imaging technique. Transgenic mice carrying the luciferase gene under the control of NF-kappaB were given A. camphorata (570mg/kg, p.o.) for three consecutive days and then injected with LPS (4mg/kg, i.p.). In vivo imaging showed that treatment with LPS increased the luminescent signal, whereas A. camphorata suppressed the LPS-induced inflammatory response significantly. Ex vivo imaging showed that A. camphorata suppressed LPS-induced NF-kappaB activity in the small intestine, mesenteric lymph nodes, liver, spleen, and kidney. Immunohistochemical staining revealed that A. camphorata suppressed production of the LPS-induced tumour necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and NF-kappaB p65 subunit in these organs. Furthermore, A. camphorata attenuated the productions of LPS-induced TNF-alpha and IL-1beta in serum from transgenic mice. We report the first confirmation of the anti-inflammatory action in vivo of this potentially beneficial mushroom. Copyright 2010 Elsevier Ltd. All rights reserved. PMID: 20621584 [PubMed - as supplied by publisher] 87. Structure and functions of gamma-dodecalactone isolated from Antrodia camphorata for NK cell activation. Chen CJ, Vijaya Krishna R, Tsai CC, Wu WH, Chao LK, Hwang KH, Chien CM, Chang HY, Chen ST. Bioorg Med Chem. 2010 Jul 16. [Epub ahead of print] Institute of Biochemical Sciences, National Taiwan University, Taipei, Taiwan; Institute of Biological Chemistry and Genomic Research Center, Academia Sinica, 128, Sec. 2, Academia Rd., Taipei 115, Taiwan. Abstract: The preserved fungal species Antrodia camphorata has diverse health-promoting effects and has been popularly used in East Asia as a traditional herb. We isolated a volatile compound from the culture medium of A. camphorata and identified it as gamma-dodecalactone (gamma-DDL). Cytomic screening for immune-modulating activity revealed that gamma-DDL can activate human NK cells to express the early activation marker CD69. Further experiments showed that gamma-DDL not only can induce NK cells to express CD69 but also stimulate NK cells to secrete cytotoxic molecules (FasL and granzyme B) and Th1 cytokines (TNF-alpha and INF-gamma). Measuring the distribution of gamma-DDL in the subcellular compartments of NK cells revealed that gamma-DDL has been converted to 4-hydroxydodecanoic acid (an acyclic isomer of gamma-DDL) in a time-dependent manner in the cytoplasm. Synthetic (R,S)-4-hydroxydodecanoic acid 40

activated NK cells to express CD69 mRNA within 10min, in contrast to gamma-DDL, which activated NK cells to express CD69 within 50min. This faster activation suggests that gamma-DDL has converted to 4-hydroxydodecanoic acid and to stimulate the NK cells to express CD69. Optically pure (R)-(+)-4-hydroxydodecanoic acid and (S)-(-)-4-hydroxydodecanoic acid were obtained via: (1) synthesis of its diastereomeric esters of (R,S)-4-hydroxydodecanoic (R)-(-)-2-phenylpropionate; (2) separation of diastereomers via preparative HPLC, and (3) subsequent hydrolysis of the obtained optical pure ester of (R)-(+)-4-hydroxydodecanoic acid (R)-(-)-2-phenylpropionate and (R)-(-)-4-hydroxydodecanoic acid (R)-(-)-2-phenylpropionate, respectively. Further assays of NK cells activation using each enantiomer showed that only the (R)-(+)-4-hydroxydodecanoic acid can activate NK cells. PMID: 20708940 [PubMed - as supplied by publisher] 88. Methylantcinate A induces tumor specific growth inhibition in oral cancer cells via Bax-mediated mitochondrial apoptotic pathway. Bioorg Med Chem Lett. 2010 Oct 15;20(20):6145-8. Epub 2010 Aug 6. Tsai WC, Rao YK, Lin SS, Chou MY, Shen YT, Wu CH, Geethangili M, Yang CC, Tzeng YM. School of Medical Laboratory and Biotechnology, Chung Shan Medical University, Taichung, Taiwan, ROC; Department of Clinical Laboratory, Chung Shan Medical University Hospital, Taichung, Taiwan, ROC. Abstract: An ergostane type triterpenoid methylantcinate A (MAA) isolated from the fruiting bodies of Antrodia camphorata inhibited the growth of oral cancer cell lines OEC-M1 and OC-2 in a dose-dependent manner, without cytotoxic to normal oral gingival fibroblast cells. The major mechanism of growth inhibition was apoptosis induction, as shown by flow cytometric analysis of annexin V-FITC and propidium iodide staining, caspase-3 activation and DNA fragmentation. The increased expression of pro-apoptotic Bax, poly-(ADP-ribose) polymerase cleavage, and activated caspase-3 and decreased expression of anti-apoptotic Bcl-2 and Bcl-xL were also observed. These results provide the first evidence that the anti-oral cancer effects of MAA may involve a mechanism through the mitochondrial dependent pathway. Thus, results reported here may offer further impulse to the development of MAA analogues as potential chemotherapeutic targets for oral cancer complications. PMID: 20817519 [PubMed - in process] 89. Anti-metastatic activities of Antrodia camphorata against human breast cancer cells mediated through suppression of the MAPK signaling pathway. Food Chem Toxicol. 2010 Nov 4. [Epub ahead of print] Yang HL, Kuo YH, Tsai CT, Huang YT, Chen SC, Chang HW, Lin E, Lin WH, Hseu YC. Institute of Nutrition, China Medical University, Taichung, Taiwan. Abstract: The fermented culture broth of Antrodia camphorata (A. camphorata) has been shown to promote cell cycle arrest and apoptosis of human estrogen-nonresponsive MDA-MB-231 cells. Herein, we demonstrate that non-cytotoxic concentrations (20-80g/mL) of A. camphorata markedly inhibited the invasion/migration of highly metastatic MDA-MB-231 cells as shown by an in vitro transwell and a wound-healing repair assay. The results of a gelatin zymography assay showed that A. camphorata suppressed the activity of matrix metalloproteinase (MMP)-9 and urokinase plasminogen activator (uPA). Western blot results demonstrated that treatment with A. camphorata decreased the expression of MMP-9, MMP-2, uPA, uPA receptor (uPAR) and vascular endothelial growth factor (VEGF); while the expression of the endogenous inhibitors of these proteins, i.e., tissue inhibitors of MMP (TIMP-1 and TIMP-2), and plasminogen activator inhibitor (PAI)-1, increased. Further investigation revealed that A. camphorata suppressed the phosphorylation of ERK1/2, p38, and JNK1/2. A. camphorata treatment also led to a dose-dependent inhibition on NF-B binding and activation. This is the first report confirming the anti-metastatic activity of this potentially beneficial mushroom against human breast cancer. Copyright 2010 Elsevier Ltd. All rights reserved. PMID: 21056076 [PubMed - as supplied by publisher] 90. First total synthesis of antrocamphin A and its analogs as anti-inflammatory and anti-platelet aggregation agents. Org Biomol Chem. 2010 Nov 18. [Epub ahead of print] Lee CL, Huang CH, Wang HC, Chuang DW, Wu MJ, Wang SY, Hwang TL, Wu CC, Chen YL, Chang FR, Wu YC. Graduate Institute of Natural Products, Kaohsiung Medical University, Kaohsiung 807, Taiwan. aaronfrc@kmu.edu.tw. Abstract: Naturally occurring antrocamphin A (1) is a potent anti-inflammatory compound from the edible fungus Antrodia camphorata (Taiwanofungus camphoratus), whose wild fruiting body is used as a valuable folk medicine in Taiwan. This 41

study is the first total synthesis of antrocamphin A (1) and its analogs. Their inhibition ability on NO release, superoxide anion generation, elastase release and platelet aggregation are reported herein. PMID: 21088769 [PubMed - as supplied by publisher] 91. Biologically active constituents from the fruiting body of Taiwanofungus camphoratus. Bioorg Med Chem. 2010 Oct 20. [Epub ahead of print] Shi LS, Chao CH, Shen DY, Chan HH, Chen CH, Liao YR, Wu SJ, Leu YL, Shen YC, Kuo YH, Lee EJ, Qian K, Wu TS, Lee KH. Department of Biotechnology, National Formosa University, Yunlin 632, Taiwan. Abstract: Five new benzenoids, benzocamphorins A-E (1-5), and 10 recently isolated triterpenoids, camphoratins A-J (16-25), together with 23 known compounds including seven benzenoids (6-12), three lignans (13-15), and 13 triterpenoids (26-38) were isolated from the fruiting body of Taiwanofungus camphoratus. Their structures were established by spectroscopic analysis. Selected compounds were examined for cytotoxic and anti-inflammatory activities. Compounds 9 and 21 showed moderate cytotoxicity against MCF-7 and Hep2 cell lines with ED(50) values of 3.4 and 3.0g/mL, respectively. Compounds 21, 25, 26, 29-31, 33, and 36 demonstrated potent anti-inflammatory activity by inhibiting lipopolysaccharide (LPS)-induced nitric oxide (NO) production with IC(50) values of 2.5, 1.6, 3.6, 0.6, 4.1, 4.2, 2.5, and 1.5M, respectively, which were better than those of the nonspecific nitric oxide synthase (NOS) inhibitor N-nitro-l-arginine methyl ester (l-NAME) (IC(50): 25.8M). These results may substantiate the use of T. camphoratus in traditional Chinese medicine (TCM) for the treatment of inflammation and cancer-related diseases. The newly discovered compounds deserve further development as anti-inflammatory candidates. Copyright 2010 Elsevier Ltd. All rights reserved. PMID: 21115251 [PubMed - as supplied by publisher] 92. Identification of Antrocin from Antrodia camphorata as a Selective and Novel Class of Small Molecule Inhibitor of Akt/mTOR Signaling in Metastatic Breast Cancer MDA-MB-231 Cells. Chem Res Toxicol. 2010 Dec 15. [Epub ahead of print] Rao YK, Wu AT, Geethangili M, Huang MT, Chao WJ, Wu CH, Deng WP, Yeh CT, Tzeng YM. Institute of Biochemical Sciences and Technology, Chaoyang University of Technology , Wufeng, Taiwan, ROC. Abstract: The PI3K/Akt/mTOR pathway is considered to be an attractive target for the development of novel anticancer molecules. This paper reports for the first time that a small molecule, antrocin (MW = 234), from Antrodia camphorata was a potent antagonist in various cancer types, being highest in metastatic breast cancer MDA-MB-231 cells (MMCs) with an IC(50) value of 0.6 M. Antrocin was a superior antiproliferator in MMCs as compared with doxorubicin and cisplatin, prevents colony formation, and was nontoxic to nontumorgenic MCF10A and HS-68 cells. Antrocin induced dose-dependent apoptosis in MMCs and caused cleavage of caspase-3 and poly(ADP-ribose) polymerase. Antrocin also caused a time-dependent decrease in protein expression of anti-apoptotic Bcl-2, Bcl-xL, survivin, and their mRNA, with concomitant increase in pro-apoptotic Bax and cytosolic cytochrome c. In a mechanistic study, antrocin suppressed the phosphorylation of Akt and its downstream effectors mTOR, GSK-3, and NF-B. Furthermore, down-regulation of Akt by small interfering RNA prior to antrocin treatment resulted in enhanced cell growth inhibition and apoptosis. Thus, antrocin as an Akt/mTOR dual inhibitor has broad applicability in the development of a clinical trial candidate for the treatment of metastatic breast cancer. PMID: 21158420 [PubMed - as supplied by publisher] 93. Antroquinonol inhibits NSCLC proliferation by altering PI3K/mTOR proteins and miRNA expression profiles. Mutat Res. 2010 Dec 23. [Epub ahead of print] Kumar VB, Yuan TC, Liou JW, Yang CJ, Sung PJ, Weng CF. Department of Life Science and Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan. Abstract: Antroquinonol a derivative of Antrodia camphorata has been reported to have antitumor effects against various cancer cells. However, the effect of antroquinonol on cell signalling and survival pathways in non-small cell lung cancer (NSCLC) cells has not been fully demarcated. Here we report that antroquinonol treatment significantly reduced the proliferation of three NSCLC cells. Treatment of A549 cells with antroquinonol increased cell shrinkage, apoptotic vacuoles, pore formation, TUNEL positive cells and increased Sub-G1 cell population with respect to time and dose dependent manner. 42

Antroquinonol treatment not only increased the Sub-G1 accumulation but also reduced the protein levels of cdc2 without altering the expression of cyclin B1, cdc25C, pcdc2, and pcdc25C. Antroquinonol induced apoptosis was associated with disrupted mitochondrial membrane potential and activation of Caspase 3 and PARP cleavage in A549 cells. Moreover, antroquinonol treatment down regulated the expression of Bcl2 proteins, which was correlated with the decreased PI3K and mTOR protein levels without altering pro apoptotic and anti apoptotic. Results from the microarray analysis demonstrated that antroquinonol altered the expression level of miRNAs compared with untreated control in A549 cells. The data collectively suggested the antiproliferative effect of antroquinonol on NSCLC A549 cells, which provides useful information for understanding the anticancer mechanism influenced by antroquinonol and is the first report to suggest that antroquinonol may be a promising chemotherapeutic agent for lung cancer. Copyright 2010. Published by Elsevier B.V. PMID: 21185843 [PubMed - as supplied by publisher] 94. Antroquinonol reduces oxidative stress through enhancing Nrf2 signaling pathway and inhibits inflammation and sclerosis in focal segmental glomerulosclerosis mice. Free Radic Biol Med. 2011 Mar 1. [Epub ahead of print] Tsai PY, Ka SM, Chao TK, Chang JM, Lin SH, Li CY, Kuo MT, Chen P, Chen A. Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei, Taiwan, R.O.C. Abstract: Oxidative stress, inflammation, and fibrosis are involved in development and progression of focal segmental glomerulosclerosis (FSGS), a common form of idiopathic nephrotic syndrome and represents a therapeutic challenge due to it have a poor response to steroids. Antroquinonol (Antroq), a purified compound, is a major active component of a mushroom namely Antrodia camphorata growing in the camphor tree in Taiwan, and has inhibitory effects on nitric oxide production and inflammatory reactions. We hypothesized that Antroq might ameliorate FSGS renal lesions by modulating pathogenic pathways of oxidative stress, inflammation, and glomerular sclerosis in the kidney. We demonstrated that Antroq significantly (1) attenuated proteinuria, renal dysfunction, and glomerulopathy, including epithelial hyperplasia lesions and podocyte injury; (2) reduced oxidative stress, leukocyte infiltration, and expression of fibrosis-related proteins in the kidney; (3) increased renal nuclear factor E2-related factor 2 (Nrf2) and glutathione peroxidase activity; and (4) inhibited renal nuclear factor-kappa B (NF-B) activation and decreased levels of transforming growth factor (TGF)-1 in the serum and kidney tissue in a mouse FSGS model. Our data suggest that Antroq might be a potential therapeutic agent for FSGS acting by boosting Nrf2 activation and suppressing NF-B-dependent inflammatory and TGF-1-mediated fibrosis pathways in the kidney. Copyright 2010. Published by Elsevier Inc. PMID: 21376112 [PubMed - as supplied by publisher] 95. Monothiol Glutaredoxin cDNA from Taiwanofungus camphorata : A Novel CGFS-type Glutaredoxin Possessing Glutathione Reductase Activity. J Agric Food Chem. 2011 Mar 11. [Epub ahead of print] Ken CF, Chen IJ, Lin CT, Liu SM, Wen L, Lin CT. Institute of Bioscience and Biotechnology and Center for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University , Keelung, Taiwan. Abstract: Glutaredoxins (Grxs) play important roles in the redox system via reduced glutathione as a reductant. A TcmonoGrx cDNA (1039 bp, EU158772) encoding a putative monothiol Grx was cloned from Taiwanofungus camphorata (formerly named Antrodia camphorata ). The deduced amino acid sequence is conserved among the reported monothiol Grxs. Two 3-D homology structures of the TcmonoGrx based on known structures of human Grx3 (pdb: 2DIY_A) and Mus musculus Grx3 (pdb: 1WIK_A) have been created. To characterize the TcmonoGrx protein, the coding region was subcloned into an expression vector pET-20b(+) and transformed into E. coli C41(DE3). The recombinant His6-tagged TcmonoGrx was overexpressed and purified by Ni(2+)-nitrilotriacetic acid Sepharose. The purified enzyme showed a predominant band on 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme exhibited glutathione reductase (GR) activity via dithionitrobenzoate (DTNB) assay. The Michaelis constant (K(M)) values for GSSG and NADPH were 0.064 and 0.041 mM, respectively. The enzyme's half-life of deactivation at 60 C was 10.5 min, and its thermal inactivation rate constant (k(d)) was 5.37 10(-2) min(-1). The enzyme was active under a broad pH range from 6

43

to 8. The enzyme retained 50% activity after trypsin digestion at 37 C for 40 min. Both mutants C(40)S(40) and C(165)S(165) lost 40-50% GR activity, whereas the mutant S(168)C(168) showed a 20% increase in its GR activity. PMID: 21395221 [PubMed - as supplied by publisher] 96. Effects of Antrodia camphorata on Alcohol Clearance and Antifibrosis in Livers of Rats Continuously Fed Alcohol. J Agric Food Chem. 2011 Mar 14. [Epub ahead of print] Wu MT, Tzang BS, Chang YY, Chiu CH, Kang WY, Huang CH, Chen YC. Biotechnology Division, Taiwan Agricultural Research Institute , Council of Agriculture, Taichung County 413, Taiwan. Abstract: Alcoholic fatty liver disease (AFLD) is the result of an excessive or chronic consumption of alcohol. Nine male Wistar rats per group were randomly assigned to one of the following drinking treatments: a 20% (w/w) alcohol solution (ALC); a 20% (w/w) alcohol solution cotreated with 0.25 g silymarin/kg BW/day; or a 20% (w/w) alcohol solution cotreated with 0.025 g Niuchangchih (Antrodia camphorata )/kg BW/day for 4 weeks. Rats with cotreatments of silymarin or Niuchangchih had smaller (p < 0.05) relative liver size, less (p < 0.05) liver lipid accumulation, and lower (p < 0.05) liver damage indices [aspartate aminotransferase (AST) and alkaline phosphatase (ALP) values]. In the regulation of alcohol metabolism, the lower serum alcohol level was observed only in alcohol-fed rats supplemented with Niuchangchih. Meanwhile, cotreatment of silymarin or Niuchangchih increased (p < 0.05) CAT and ALDH activities but did not (p > 0.05) affect ADH and CYP2E1 expressions, which accelerate alcohol metabolism in the body. Additionally, neither silymarin nor Niuchangchih (p > 0.05) influenced serum/hepatic MMP-2 activities and NF-B, AP1, and -SMA gene expressions, but serum/hepatic MMP-9 activities and TNF-, KLF-6, and TGF-1 gene expressions of alcohol-fed rats were down-regulated (p < 0.05) by silymarin or Niuchangchih, which also could explain the lower liver damage observed in rats chronically fed alcohol. PMID: 21401100 [PubMed - as supplied by publisher] 97. Is 2,3,4,5-tetramethoxybenzoyl chloride a natural product? J Nat Prod. 2011 May 27;74(5):1348-50. Epub 2011 Apr 14. Punch KA, Ghisalberti EL, Piggott MJ. Source: School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia , Crawley, Western Australia, 6009. Abstract: The title compound, which was reported to be a constituent of the fruiting body of the fungus Antrodia camphorata, has been synthesized. The reactivity and spectroscopic properties of the synthetic material do not match those of the natural product. There is currently insufficient information for a definitive structural reassignment. PMID: 21491924, [PubMed - in process] 98. Apoptotic Cell Death and Inhibition of Wnt/-Catenin Signaling Pathway in Human Colon Cancer Cells by an Active Fraction (HS7) from Taiwanofungus camphoratus. Evid Based Complement Alternat Med. 2011;2011:750230. Epub 2011 Mar 9. Yeh CT, Yao CJ, Yan JL, Chuang SE, Lee LM, Chen CM, Yeh CF, Li CH, Lai GM. Cancer Center, Shuang Ho Hospital, Taipei Medical University, Taipei 235, Taiwan. Abstract: Aberrant activation of Wnt/-catenin signaling plays an important role in the development of colon cancer. HS7 is an active fraction extracted from Taiwanofungus camphoratus, which had been widely used as complementary medicine for Taiwan cancer patients in the past decades. In this study, we demonstrated the effects of HS7 on the growth inhibition, apoptosis induction, and Wnt/-catenin signaling suppression in human colon cancer cells. HS7 significantly inhibited proliferation of HT29, HCT116, and SW480 colon cancer cells in a dose- and time-dependent manner. The apoptosis induction was evidenced by DNA fragmentation and subG1 accumulation, which was associated with increased Bax/Bcl-2 ratio, activation of caspase-3 and cleavage of PARP. By using Tcf-dependent luciferase activity assay, HS7 was found to inhibit the -catenin/Tcf transcriptional activities. In addition, HS7 strongly suppressed the binding of Tcf complexes to its DNA-binding site shown in electrophoretic mobility shift assay. This inhibition was further confirmed by the decreased protein levels of Tcf-4 and -catenin. The -catenin/Tcf downstream target genes, such as survivin, c-myc, cyclin D1, MMP7, and MT1-MMP involved in apoptosis, invasion, and angiogenesis were also diminished as well. These results indicate that Taiwanofungus camphoratus may provide a benefit as integrative medicine for the treatment of colon cancer. PMID: 21423639 [PubMed - in process]PMCID: PMC3057579 44

99. Macrophage Mediated Anti-Proliferation Effects of Anthodia camphorata Non-Polysaccharide Based Extracts on Human Hepatoma Cells. Biosci Biotechnol Biochem. 2011 May 10;75(4):624-32. Epub 2011 Apr 22. Chang CY, Cheng TJ, Chang FR, Wang HY, Kan WC, Li SL, Huang LH, Chen YC, Tsai WC, Huang CH, Cheng CH, Lee GY, Shyue SW, Chen YP, Lin KC, Chuu JJ. Source: Institute of Biotechnology, College of Engineering, Southern Taiwan University. Abstract: It has been reported that medicinal mushrooms might induce different types of immune responses. Anthodia camphorata (A. camphorata) has attracted much attention for its therapeutic effects in treating hepatoma. We tested this anti-tumor effects using immunomodulation of macrophages and extracts of A. camphorata. We evaluated the anti-proliferation effects of various extracts of A. camphorata from fruiting bodies (AC-FB), mycelium of solid-state cultures (AC-SS), liquid-state cultures (AC-LS) and polyaccharide extracts from liquid-state cultures (AC-PS), and extracts of A. camphorata stimulated RAW 264.7 macrophage cell-conditioned mediums (MC-CMs). We measured cell proliferation and, did migration assays by cell cycle analysis and by observing apoptosis-related proteins (AKT, PARP-1, and NF-B) and the mRNA expression of cytokines (TNF- and IL-1) of macrophages in human hepatoma cell lines. Our results revealed that two of the extracts (AC-FB and AC-SS) had better anti-proliferation effects, implying an immunomodulatory role the macrophages might play. This outcome is consistent with findings that AC-FB and AC-SS increase mRNA expression of TNF- and the corresponding expression of apoptosis-related proteins on activation of MC-CMs, while A. camphorata polysaccharides induce macrophage-derived anti-tumor activities in human hepatoma cells via IL-1 and Akt activation. These results indicate that anti-tumor effects exerted by modulation of macrophage activation of A. camphorate may be influenced by the other constituents which (contained little or no polysaccharide) of A. camphorata. Acta Pharmacol Sin. 2011 May 23. [Epub ahead of print] 100. Antcin A, a steroid-like compound from Antrodia camphorata, exerts anti-inflammatory effect via mimicking glucocorticoids. Chen YC, Liu YL, Li FY, Chang CI, Wang SY, Lee KY, Li SL, Chen YP, Jinn TR, Tzen JT. Source: Graduate Institute of Biotechnology, National Chung Hsing University, Taichung 40227, Taiwan, China. Abstract: Aim: To determine the active ingredient of Niuchangchih (Antrodia camphorata) responsible for its anti-inflammatory effects and the relevant molecular mechanisms. Methods: Five major antcins (A, B, C, H, and K) were isolated from fruiting bodies of Niuchangchih. Structural similarity between the antcins and 2 glucocorticoids (cortisone and dexamethasone) was compared. After incubation with each compound, the cytosolic glucocorticoid receptor (GR) was examined for its migration into the nucleus. Mo lecular docking was performed to model the tertiary structure of GR associated with antcins.Results:Incubation with cortisone, dexamethasone or antcin A (but not antcins B, C, H, and K) led to the migration of glucocorticoid receptor into the nucleus. The minimal concentration of antcin A, cortisone and dexamethasone to induce nuclear migration of glucocorticoid receptor was 10, 1, and 0.1 mol/L, respectively. The results are in agreement with the simulated binding affinity scores of these three ligands docking to the glucocorticoid receptor. Molecular modeling indicates that C-7 of antcin A or glucocorticoids is exposed to a hydrophobic region in the binding cavity of the glucocorticoid receptor, and the attachment of a hydrophilic group to C-7 of the other four antcins presumably results in their being expelled when docking to the cavity. Conclusion: The anti-inflammatory effect of Niuchangchih is, at least, partly attributed to antcin A that mimics glucocorticoids and triggers translocation of glucocorticoid receptor into nucleus to initiate the suppressing inflammation. PMID: 21602840, [PubMed - as supplied by publisher] 101. Study of the Anti-Proliferative Activity of 5-Substituted 4,7-Dimethoxy-1,3-Benzodioxole Derivatives of SY-1 from Antrodia camphorata on Human COLO 205 Colon Cancer Cells. Evid Based Complement Alternat Med. 2011;2011:450529. Epub 2011 May 3. Lien HM, Kuo PT, Huang CL, Kao JY, Lin H, Yang DY, Lai YY. Department of Chemistry, Tunghai University, Taichung, Taiwan. Abstract: A set of 10 4,7-dimethoxy-1,3-benzodioxole derivatives based on a lead compound previously discovered by our group, SY-1, which was isolated from Antrodia camphorata, were evaluated for their in vitro inhibitory activity on human 45

colorectal carcinoma cells (COLO 205). Structure-activity relationship studies of the 10 compounds indicated the importance of the chain length of the alkyl group at the 5-position, and the 2-propenyl substituent named "apiole" exhibited the most potent inhibitory activity. In the present study, we demonstrate that the SY-1 analogue "apiole" decreased the proliferation of COLO 205 cells, but not that of normal human colonic epithelial cells (FHC). The G0/G1 cell cycle arrest induced by apiole (75-225M) was associated with significantly increased levels of p53, p21 and p27 and decreased levels of cyclin D1. Concerning COLO 205 cell apoptosis, apiole (>150M) treatment significantly increased the levels of cleaved caspases 3, 8, 9 and bax/bcl-2 ratio and induced ladder formation in DNA fragmentation assay and sub-G1 peak in flow cytometry analysis. These findings suggest that apiole can suppress COLO 205 cell growth; however, the detailed mechanisms of these processes require further investigation. PMID: 21785624, [PubMed - in process] PMCID: PMC3138483 102. A Preclinical Evaluation of Antrodia camphorata Alcohol Extracts in the Treatment of Non-Small Cell Lung Cancer Using Non-Invasive Molecular Imaging. Evid Based Complement Alternat Med. 2011;2011:914561. Epub 2011 Mar 13. Chiou JF, Wu AT, Wang WT, Kuo TH, Gelovani JG, Lin IH, Wu CH, Chiu WT, Deng WP. Cancer Center and Department of Radiatioin Oncology, Taipei Medical University Hospital, Taipei, Taiwan. Abstract: This study was carried out to provide a platform for the pre-clinical evaluation of anti-cancer properties of a unique CAM (complementary and alternative medicine) agent, Antrodia camphorata alcohol extract (ACAE), in a mouse model with the advantageous non-invasive in vivo bioluminescence molecular imaging technology. In vitro analyses on the proliferation, migration/invasion, cell cycle and apoptosis were performed on ACAE-treated non-small cell lung cancer cells, H441GL and control CGL1 cells. In vivo, immune-deficient mice were inoculated subcutaneously with H441GL followed by oral gavages of ACAE. The effect of ACAE on tumor progression was monitored by non-invasive bioluminescence imaging. The proliferation and migration/invasion of H441GL cells were inhibited by ACAE in a dose-dependent manner. In addition, ACAE induced cell cycle arrest at G0/G1 phase and apoptosis in H441GL cells as shown by flow cytometric analysis, Annexin-V immunoflourescence and DNA fragmentation. In vivo bioluminescence imaging revealed that tumorigenesis was significantly retarded by oral treatment of ACAE in a dose-dependent fashion. Based on our experimental data, ACAE contains anti-cancer properties and could be considered as a potential CAM agent in future clinical evaluation. PMID: 21785640, [PubMed - in process] PMCID: PMC3137791 103. Antcin A contributs to anti-inflammatory effect of Niuchangchih (Antrodia camphorata). Acta Pharmacol Sin. 2011 Aug;32(8):981-2. doi: 10.1038/aps.2011.104. Lu ZM, Xu ZH. Laboratory of Pharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, Wuxi 214122, China. PMID: 21818107, [PubMed - in process] 104. Changes in volatile compound composition of Antrodia camphorata during solid state fermentation. J Sci Food Agric. 2011 Aug 5. doi: 10.1002/jsfa.4488. [Epub ahead of print] Xia Y, Zhang B, Li W, Xu G. Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, People's Republic of China. Abstract: BACKGROUND: Although the volatiles present in mushrooms and fungi have been investigated by many researchers, including Antrodia camphorata in submerged fermentation, there are few data available regarding changes in volatile compounds during fermentation. Our research has revealed that solid state fermentation of A. camphorata is highly odiferous compared with submerged cultures and the odor changed with increasing culture time. Therefore the aim of this study was to investigate the changes in volatile compound composition of A. camphorata during solid state fermentation. RESULTS: Altogether, 124 major volatile compounds were identified. The volatile compounds produced by A. camphorata during growth in solid state fermentation were quite different. Oct-1-en-3-ol, octan-3-one and methyl 2-phenylacetate were predominant in exponential growth phase production, while the dominant volatiles produced in stationary phase were octan-3-one and methyl 2-phenylacetate. In stationary phase, lactone compounds in A. camphorata, 46

such as 5-butyloxolan-2-one, 5-heptyloxolan-2-one, 6-heptyloxan-2-one, contributed greatly to peach and fruit-like flavor. Terpene and terpene alcohol compounds, such as 1-terpineol, L-linalool, T-cadinol, (E, E)-farnesol, -elemene, cis--bisabolene and -muurolene, made different contributions to herbal fresh aroma in A. camphorata. Nineteen volatile sesquiterpenes were detected from solid state fermentation of A. camphorata. The compounds 5-n-butyl-5H-furan-2-one, -ionone, (-)-caryophyllene oxide, aromadendrene oxide, diepi--cedrene epoxide, -elemene, -selinene, -muurolene, azulene, germacrene D, -cadinene and 2-methylpyrazine have not hitherto been reported in A. camphorata. CONCLUSION: The preliminary results suggest that the aroma-active compounds produced by A camphorata in solid state fermentation might serve as an important source of natural aroma compounds for the food and cosmetic industries or antibiotic activity compounds. The sesquiterpenes could be identified as possible taxonomic markers for A. camphorata. Copyright 2011 Society of Chemical Industry. PMID: 21823126, [PubMed - as supplied by publisher] 105. Antroquinonol, a natural ubiquinone derivative, induces a cross talk between apoptosis, autophagy and senescence in human pancreatic carcinoma cells. J Nutr Biochem. 2011 Aug 11. [Epub ahead of print] Yu CC, Chiang PC, Lu PH, Kuo MT, Wen WC, Chen P, Guh JH. School of Pharmacy, National Taiwan University, Taipei 100, Taiwan. Abstract: Pancreatic cancer is a malignant neoplasm of the pancreas. A mutation and constitutive activation of K-ras occurs in more than 90% of pancreatic adenocarcinomas. A successful approach for the treatment of pancreatic cancers is urgent. Antroquinonol, a ubiquinone derivative isolated from a camphor tree mushroom, Antrodia camphorata, induced a concentration-dependent inhibition of cell proliferation in pancreatic cancer PANC-1 and AsPC-1 cells. Flow cytometric analysis of DNA content by propidium iodide staining showed that antroquinonol induced G1 arrest of the cell cycle and a subsequent apoptosis. Antroquinonol inhibited Akt phosphorylation at Ser(473), the phosphorylation site critical for Akt kinase activity, and blocked the mammalian target of rapamycin (mTOR) phosphorylation at Ser(2448), a site dependent on mTOR activity. Several signals responsible for mTOR/p70S6K/4E-BP1 signaling cascades have also been examined to validate the pathway. Moreover, antroquinonol induced the down-regulation of several cell cycle regulators and mitochondrial antiapoptotic proteins. In contrast, the expressions of K-ras and its phosphorylation were significantly increased. The coimmunoprecipitation assay showed that the association of K-ras and Bcl-xL was dramatically augmented, which was indicative of apoptotic cell death. Antroquinonol also induced the cross talk between apoptosis, autophagic cell death and accelerated senescence, which was, at least partly, explained by the up-regulation of p21(Waf1/Cip1) and K-ras. In summary, the data suggest that antroquinonol induces anticancer activity in human pancreatic cancers through an inhibitory effect on PI3-kinase/Akt/mTOR pathways that in turn down-regulates cell cycle regulators. The translational inhibition causes G1 arrest of the cell cycle and an ultimate mitochondria-dependent apoptosis. Moreover, autophagic cell death and accelerated senescence also explain antroquinonol-mediated anticancer effect. Copyright 2011 Elsevier Inc. All rights reserved. PMID: 21840189, [PubMed - as supplied by publisher] 106. Optimization of fermentation medium for triterpenoid production from Antrodiacamphorata ATCC 200183 using artificial intelligence-based techniques. Appl Microbiol Biotechnol. 2011 Aug 26. [Epub ahead of print] Lu ZM, Lei JY, Xu HY, Shi JS, Xu ZH. Laboratory of Pharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, People's Republic of China. Abstract: In this study, alteration in morphology of submergedly cultured Antrodia camphorata ATCC 200183 including arthroconidia, mycelia, external and internal structures of pellets was investigated. Two optimization models namely response surface methodology (RSM) and artificial neural network (ANN) were built to optimize the inoculum size and medium components for intracellular triterpenoid production from A. camphorata. Root mean squares error, R (2), and standard error of prediction given by ANN model were 0.31%, 0.99%, and 0.63%, respectively, while RSM model gave 1.02%, 0.98%, and 2.08%, which indicated that fitness and prediction accuracy of ANN model was higher when compared to RSM model. Furthermore, using genetic algorithm (GA), the input space of ANN model was optimized, and maximum triterpenoid production of 62.84 mg l(-1) was obtained at the GA-optimized concentrations of arthroconidia (1.7810(5) ml(-1)) and 47

medium components (glucose, 25.25 gl(-1); peptone, 4.48 gl(-1); and soybean flour, 2.74 gl(-1)). The triterpenoid production experimentally obtained using the ANN-GA designed medium was 64.792.32 mg l(-1) which was in agreement with the predicted value. The same optimization process may be used to optimize many environmental and genetic factors such as temperature and agitation that can also affect the triterpenoid production from A. camphorata and to improve the production of bioactive metabolites from potent medicinal fungi by changing the fermentation parameters. PMID: 21870045, [PubMed - as supplied by publisher] 107. Antrodia camphorata Induces Apoptosis and Enhances the Cytotoxic Effect of Paclitaxel in Human Ovarian Cancer Cells. Int J Gynecol Cancer. 2011 Sep 2. [Epub ahead of print] Liu FS, Yang PY, Hu DN, Huang YW, Chen MJ. *Cancer Center and Department of Medical Research, Show Chwan Memorial Hospital, Changhua, Taiwan, ROC; Tissue Culture Center, New York Eye & Ear Infirmary, New York, NY. Abstract INTRODUCTION: Antrodia camphorata is a Chinese herb. Recently, several reports demonstrated that it had growth-inhibiting effects on some cancer cells. In this study, we investigated whether the crude extract of A. camphorata could inhibit the growth of ovarian cancer cells and examined the possible mechanisms involved. We also examined whether the cytotoxic effect of paclitaxel on ovarian cancer cells would be affected by A. camphorata. MATERIALS AND METHODS: Two human ovarian cancer cell lines, SKOV-3 and TOV-21G, were treated with A.camphorata (3-300 g/mL). An MTT assay was used to test its cytotoxic effect. The apoptosis-related factors including the activity of caspase-3, -8, and -9 and the cytochrome c level released from mitochondria were analyzed. The expression of Bcl-2 family proteins (Bcl-2, Bcl-xL, Bax, Bim, Bad, and Bak) was examined by Western blot analysis. Cell lines were further treated with paclitaxel or paclitaxel plus A. camphorata to examine the cytotoxic efficiency. RESULTS: The MTT assay revealed that A. camphorata was cytotoxic to both the ovarian cancer cells in a dose- and time-dependent manner. Activities of caspase-3, -8, and -9 and release of mitochondrial cytochrome c increased in both ovarian cancer cell lines with increased dose of A. camphorata. Western blot analysis of Bcl-2 family proteins revealed an increased expression of Bad in SKOV-3 cells, whereas increased expression of Bim and Bak and decreased expression of Bcl-xL were noted in TOV-21G cells. In addition, the cytotoxic effect of paclitaxel on SKOV-3 and TOV-21G cells was increased significantly with the addition of A. camphorata (P < 0.01) by MTT assay. CONCLUSIONS: These in vitro results suggest that A. camphorata causes a cytotoxic effect on ovarian cancer cells through the induction of apoptosis. It may also enhance the antitumor effect of paclitaxel. Further studies with the ultimate goal of conducting clinical trials are warranted. PMID: 21897275, [PubMed - as supplied by publisher] 108. Pretreatment with an ethanolic extract of Taiwanofungus camphoratus (Antrodiacamphorata) enhances the cytotoxic effects of Amphotericin B. J Agric Food Chem. 2011 Sep 7. [Epub ahead of print] Chen LY, Sheu MT, Liu DZ, Liao CK, Ho HO, Kao WY, Ho YS, Lee WS, Su CH. Abstract: Taiwanofungus camphoratus, a well-known Chinese medicine used in Taiwan, possesses several pharmacological functions, including anticancer effects. In the present study, we aimed to investigate a novel anticancer effect by pretreating cancer cells with an ethanolic extract of Taiwanofungus camphoratus (TCEE) followed by the administration of an antifungal agent Amphotericin B (AmB). Both TCEE and AmB showed significant dose-dependent cytotoxicity in HT29 cells. Pretreatment with a non-toxic dose of TCEE enhanced the cytotoxicity of AmB. Furthermore, significant apoptotic cell death was found in cells treated with TCEE and AmB. Combination treatment with AmB plus TCEE resulted in a significant repression of tumor growth in HT29 xenografts. Collectively, our results indicated that combined treatment with AmB and TCEE effectively induced apoptosis and inhibited tumor growth. In the future, TCEE may serve as a potential complementary and alternative medicine to treat patients suffering from colorectal cancer. 48

PMID: 21899275, [PubMed - as supplied by publisher] 109. Antroquinonol differentially modulates T cells activity, reduces IL-18 production, but enhances Nrf2 activation in accelerated severe lupus nephritis. Arthritis Rheum. 2011 Sep 8. doi: 10.1002/art.33328. [Epub ahead of print] Tsai PY, Ka SM, Chang JM, Lai JH, Dai MS, Jheng HL, Kuo MT, Chen P, Chen A. Graduate Institute of Medical Sciences, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan, R.O.C. Abstract OBJECTIVE: Accelerated severe lupus nephritis (ASLN), with an acute onset of severe clinical manifestations and histopathological renal lesions, may represent transformation of mild lupus nephritis to a severe form of glomerulonephritis. Abnormal activation of T and B cells and/or oxidative stress may play a major role in the pathogenesis of ASLN. We tested the hypothesis that antroquinonol, a purified compound and a major effective component of Antrodia camphorata with anti-inflammatory and antioxidant activities, might prevent the transformation of mild lupus nephritis into higher grade (severe form) nephritis in ASLN model. METHODS: Experimental ASLN was induced in (NZBxNZW)F1 mice by twice weekly intraperitoneal injections of Salmonella type lipopolysaccharide and daily administration of antroquinonol by gavage for different durations starting two days after the first dose of lipopolysaccharide. RESULTS: Antroquinonol significantly ameliorated the proteinuria, hematuria, impairment of renal function, and development of severe renal lesions, especially cellular crescent formation, neutrophil infiltration, fibrinoid necrosis, and proliferation in the glomerulus and peri-glomerular interstitial inflammation. Mechanistic analyses revealed that antroquinonol administration: [1] inhibited T cell activation/proliferation, but enhanced regulatory T cell suppression and reduced renal IL-18 production; [2] inhibited the production of reactive oxygen species and nitric oxide, but increased activation of nuclear factor erythroid-2-related factor-2 (Nrf2) in the kidney; and [3] suppressed renal inflammation via blocking nuclear factor-kappaB activation. CONCLUSION: We demonstrated that antroquinonol may have therapeutic potential for the early treatment of ASLN via differentially regulating T cell function and lowering IL-18 production, but promoting Nrf2 activation. Copyright 2011 by the American College of Rheumatology. PMID: 21905011, [PubMed - as supplied by publisher] 110. Antcin B and its Ester Derivative from Antrodia camphorata Induce Apoptosis in Hepatocellular Carcinoma Cells Involves Enhancing Oxidative Stress Coincident with Activation of Intrinsic and Extrinsic Apoptotic Pathway. J Agric Food Chem. 2011 Sep 14. [Epub ahead of print] Hsieh YC, Rao YK, Whang-Peng J, Huang CY, Shyue SK, Hsu SL, Tzeng YM. ABSTRACT: The triterpenoids methylantcinate B (MAB) and antcin B (AB) isolated from the medicinal mushroom Antrodia camphorata has been identified as strong cytotoxic agents against various type of cancer cells, however, the mechanisms of MAB and AB-induced cytotoxicity have not been adequately explored. This study investigated the roles of caspase cascades, ROS, DNA damage, mitochondrial disruption, and Bax and Bcl-2 proteins in MAB and AB-induced apoptosis of hepatocellular carcinoma (HCC) HepG2 cells. Here, we showed that MAB and AB induced apoptosis in HepG2 cells, as characterized by increased DNA fragmentation, cleavage of PARP, sub-G1 population, chromatin condensation, loss of mitochondrial membrane potential and release of cytochrome c. Increasing the levels of caspase-2, -3, -8, and -9 activities was involved in MAB and AB-induced apoptosis, and they could be attenuated by inhibitors of specific caspases, indicating that MAB and AB triggered caspase-dependent apoptotic pathway. Additionally, the enhanced apoptotic effect correlates with high expression of Fas, Fas ligand as well as Bax and decreased protein levels of Bcl-XL and Bcl-2, suggesting that both the extrinsic and intrinsic apoptosis pathways were involved in the apoptotic processes. Incubation of HepG2 cells with antioxidant enzymes superoxide dismutase, catalase and, antioxidants N-acetylcysteine and ascorbic acid attenuated the ROS generation and apoptosis induced by MAB and AB, which indicate that ROS plays a pivotal role in cell death. NADPH oxidase activation was observed in MAB- and AB-stimulated HepG2 cells; however, inhibition of such activation by diphenylamine significantly blocked MAB and AB-induced ROS production and increased cell viability. Taken together, our results provide the first evidence that triterpenoids MAB and AB induced a NADPH oxidase-provoked oxidative stress and extrinsic and intrinsic apoptosis as a critical mechanism of cause cell death in HCC cells. 49

PMID: 21916504, [PubMed - as supplied by publisher] 111. Chemical profiling of the cytotoxic triterpenoid-concentrating fraction and characterization of ergostane stereo-isomer ingredients from Antrodiacamphorata. J Pharm Biomed Anal. 2011 Sep 16. [Epub ahead of print] Du YC, Wu TY, Chang FR, Lin WY, Hsu YM, Cheng FT, Lu CY, Yen MH, Tsui YT, Chen HL, Hou MF, Lu MC, Wu YC. Graduate Institute of Natural Products, College of Pharmacy, Kaohsiung Medical University, Kaohsiung 807, Taiwan. Abstract: Antrodia camphorata (AC), also known as Antrodia cinnamomea, an endemic species in Taiwan, is one of the treasured medicinal mushrooms. AC is traditionally used for its chemopreventive biofunctions. In this investigation, we report a convenient method for concentrating the antiproliferative active triterpenoid-rich fraction (FEA), from ethanolic extract of AC (EEAC). A series of stereo-isomers of zhankuic acids (1-8) from the FEA was purified by HPLC using an efficient acidic solvent system. The structures of compounds 1-8 were elucidated based on spectroscopic data analysis, and the absolute configuration of -chiral carboxylic acid at C-25 in the structures was assigned based on reaction with (R)- and (S)-1-(9-anthryl)-2,2,2-trifluoroethanol. Major ingredients of FEA (eight ergostanes 1-8 and two lanostanes 9-10) were further characterised by high-performance liquid chromatography-photodiode array detection/mass spectrometry (HPLC-PDA/MS). Compounds 1-8 and their pair mixture forms (antcin K, antcin C, zhankuic acid C, and zhankuic acid A) were subjected to anti-proliferative assay against three human leukemia cell lines. Among them, the derivatives with carbonyl group at C-3 showed cytotoxicity with IC(50) values ranging from 16.44 to 77.04g/ml. Copyright 2011. Published by Elsevier B.V. PMID: 21975021, [PubMed - as supplied by publisher] 112. Production of bioactive exopolysaccharides from bitter medicinal mushroom, Antrodia camphorata (M. Zang et C.H. Su) Sh.H. Wu et al. (Aphyllophoromycetideae) in submerged cultivation. Int J Med Mushrooms. 2011;13(1):51-60. Lung MY, Lee CY, Chen WX, Huang E. Source: Department of Chemical and Materials Engineering, Minghsin University of Science and Technology, Xinfeng, Hsinchu, Taiwan, ROC. mylong@must.edu.tw Abstract: This study examines the effects of various fructose concentrations in media on the production and quality of bioactive exopolysaccharides (EPS) from Bitter medicinal mushroom, Antrodia camphorata in submerged cultures. The fructose in media of submerged cultures of A. camphorata significantly affected the production, average molecular weight (Mn), and antioxidant activity of exopolysaccharides. The specific growth rate decreased monotonically from 0.33 to 0.25 1/day as the fructose concentration increased from 10 to 60 g/L; however, maximum production and productivity for EPS increased from 75.23 to 164.87 mg/L and 6.27 to 9.70 mg/L/day, respectively. In addition, the fed-batch culture used in this study significantly improved the production of EPS (2.43-fold enhancement, from 75.23 to 182.99 mg/L), number average molecular weights of EPS (1.47-fold enhancement, from 5.44 x 10, to 7.98 x 10(5) Da), protein/exopolysaccharide ratios (1.63-fold enhancement, from 16% to 26%), and antioxidant activity of EPS (1.32-fold enhancement, from 60% to 79%), as compared with corresponding properties of batch fermentation at 10 g/L fructose in an air-lift bioreactor. The antioxidant activity of EPS was highly correlated with number average molecular weights (R2 = 0.90) and protein/exopolysaccharide ratios (R2 = 0.96). The positive results of this research have successfully verified the promotion efficiency on the production and quality of EPS from the medicinal mushroom A. camphorata. PMID: 22135904, [PubMed - in process] 113. In vitro and in vivo comparisons of the effects of the fruiting body and mycelium of Antrodia camphorata against amyloid -protein-induced neurotoxicity and memory impairment. Appl Microbiol Biotechnol. 2012 Feb 21. [Epub ahead of print] Wang LC, Wang SE, Wang JJ, Tsai TY, Lin CH, Pan TM, Lee CL. Source: Continuing Education School, National Taitung Junior College, Taitung, Taiwan, Republic of China. Abstract: Antrodia camphorata is a particular and precious medicinal mushroom, and its fruiting body was found to provide more efficient protection from oxidative stress and inflammation than its mycelium because of its higher content of triterpenoids, total phenols, and so on. In the previous in vitro studies, the mycelium of A. camphorata is proven to provide strong neuroprotection in neuron cells and suggested to have the potential of protection against neurotoxicity of amyloid -protein (A) known as the risk factor toward Alzheimer's disease (AD) development. However, the in vivo study and the 50

comparison study with the fruiting body have not yet been investigated. This study compared the effect of the fruiting body and mycelium of A. camphorata on alleviating the A40-induced neurocytotoxicity in the in vitro A-damaged neuron cell model (PC-12 cell treated with A40) and memory impairment in the in vivo AD animal model induced with a continuous brain infusion of A40. In the results of in vitro and in vivo studies, the fruiting body possessed stronger anti-oxidative and anti-inflammatory abilities for inhibiting neurocytotoxicity in A40-treated PC-12 cells and A40 accumulation in A40-infused brain than mycelium. Moreover, hyperphosphorylated tau (p-tau) protein expression, known as an important AD risk factor, was suppressed by the treatment of fruiting body rather than that of mycelium in the in vitro and in vivo studies. These comparisons supported the reasons why the fruiting body resulted in a more significant improvement effect on working memory ability than mycelium in the AD rats. PMID: 22350319 [PubMed - as supplied by publisher] 114. In Vivo Antitumor Effects of 4,7-Dimethoxy-5-methyl-1,3-benzodioxole Isolated from the Fruiting Body of Antrodia camphorata through Activation of the p53-Mediated p27/Kip1 Signaling Pathway. J Agric Food Chem. 2012 Mar 29. [Epub ahead of print] Tu SH, Wu CH, Chen LC, Huang CS, Chang HW, Chang CH, Lien HM, Ho YS. Source: Graduate Institute of Medical Sciences, Taipei Medical University , Taipei, Taiwan. Abstract: In this study, 4,7-dimethoxy-5-methyl-1,3-benzodioxole (SY-1) was isolated from three different sources of dried Antrodiacamphorata (AC) fruiting bodies. AC is a medicinal mushroom that grows on the inner heartwood wall of Cinnamomum kanehirai Hay (Lauraceae), which is an endemic species that is used in Chinese medicine for its antitumor properties. We demonstrated that SY-1 [given as a 1-30 mg/kg body weight intraperitoneal (ip) injection three times per week] profoundly decreased the growth of COLO-205 human colon cancer cell tumor xenografts in an athymic nude mouse model. We further demonstrated that significant AC extract-mediated antitumor effects were observed at the highest concentration (5 g/kg body weight/day). No gross toxicity signs were observed (i.e., body weight changes, general appearance, or individual organ effects). Frozen COLO-205 xenograft tumors were pulverized in liquid N(2), and the expression of cell cycle regulatory proteins was detected by immunoblotting. We found that the p53-mediated p27/Kip1 protein was significantly induced in the low-dose (1 mg/kg body weight) SY-1-treated tumors, whereas the p21/Cip1 protein levels did not change. The G0/G1 phase cell cycle regulators induced by SY-1 were also associated with a significant decrease in cyclins D1, D3, and A. These results provide further evidence that SY-1 may have significance for cancer chemotherapy. PMID: 22429157, [PubMed - as supplied by publisher] 115. [Analyze on volatile compounds of Antrodia camphorata using HS-SPME-GC-MS]. Zhong Yao Cai. 2011 Nov;34(11):1722-5. [Article in Chinese] He Z, Lu ZM, Xu HY, Shi JS, Xu ZH. Source: Laboratory of Pharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, Wuxi 214122, China. Abstract: OBJECTIVE: To analyze the volatile compounds of Antrodia camphorata in solid-state and submerged cultures. METHODS: A headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry(GC-MS) were used to evaluate the profile of the volatile compounds. RESULTS: 49 volatile compounds were identified in A. camphorata mycelia in submerged culture, while 43 volatile compounds were identified in mycelia in solid-state culture. 1-octen-3-ol, 3-octanone, 1-octen-3-ylacetate, acetic acid octyl ester and ethanol were the main volatile compounds in A. camphorata mycelia in submerged culture, while 1-octen-3-ol, 3-octanone, 3-methyl-butyraldenhyde, gamma-podecalactone and methyl 2-furozte were the most potent key volatile compounds in mycelia in solid-state culture. CONCLUSION: The volatile compounds in the mycelia of A. camphorata in solid-state and submerged cultures are similar but their relative contents are different. PMID: 22506397, [PubMed - in process] 116. [Medium optimization for mycelia production of Antrodia camphorata based on artificial neural network-genetic algorithm]. Sheng Wu Gong Cheng Xue Bao. 2011 Dec;27(12):1773-9. [Article in Chinese] Lu Z, He Z, Xu H, Shi J, Xu Z. 51

Source: Laboratory ofPharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, Wuxi 214122, China. Abstract: To illustrate the complex fermentation process of submerged culture of Antrodia camphorata ATCC 200183, we observed the morphology change of this filamentous fungus. Then we used two optimization models namely response surface methodology (RSM) and artificial neural network (ANN) to model the fermentation process of Antrodia camphorata. By genetic algorithm (GA), we optimized the inoculum size and medium components for Antrodia camphorata production. The results show that fitness and prediction accuracy of ANN model was higher when compared to those of RSM model. Using GA, we optimized the input space of ANN model, and obtained maximum biomass of 6.2 g/L at the GA-optimized concentrations of spore (1.76x 10(5) /mL) and medium components (glucose, 29.1 g/L; peptone, 9.3 g/L; and soybean flour, 2.8 g/L). The biomass obtained using the ANN-GA designed medium was (6.1+/-0.2) g/L which was in good agreement with the predicted value. The same optimization process may be used to improve the production of mycelia and bioactive metabolites from potent medicinal fungi by changing the fermentation parameters. PMID: 22506418, [PubMed - in process] 117. Antileukemia component, dehydroeburicoic acid from Antrodia camphoratainduces DNA damage and apoptosis in vitro and in vivo models. Phytomedicine. 2012 Apr 17. [Epub ahead of print] Du YC, Chang FR, Wu TY, Hsu YM, El-Shazly M, Chen CF, Sung PJ, Lin YY, Lin YH, Wu YC, Lu MC. Source: Graduate Institute of Natural Products, College of Pharmacy, Kaohsiung Medical University, Kaohsiung 807, Taiwan. Abstract: Antrodia camphorata (AC) is a native Taiwanese mushroom which is used in Asian folk medicine as a chemopreventive agent. The triterpenoid-rich fraction (FEA) was obtained from the ethanolic extract of AC and characterized by high performance liquid chromatography (HPLC). FEA caused DNA damage in leukemia HL 60 cells which was characterized by phosphorylation of H2A.X and Chk2. It also exhibited apoptotic effect which was correlated to the enhancement of PARP cleavage and to the activation of caspase 3. Five major triterpenoids, antcin K (1), antcin C (2), zhankuic acid C (3), zhankuic acid A (4), and dehydroeburicoic acid (5) were isolated from FEA. The cytotoxicity of FEA major components (1-5) was investigated showing that dehydroeburicoic acid (DeEA) was the most potent cytotoxic component. DeEA activated DNA damage and apoptosis biomarkers similar to FEA and also inhibited topoisomerase II. In HL 60 cells xenograft animal model, DeEA treatment resulted in a marked decrease of tumor weight and size without any significant decrease in mice body weights. Taken together, our results provided the first evidence that pure AC component inhibited tumor growth in vivo model backing the traditional anticancer use of AC in Asian countries. Copyright 2012 Elsevier GmbH. All rights reserved. PMID: 22516893, [PubMed - as supplied by publisher] 118. Polysaccharides from extracts of Antrodia camphorata mycelia and fruiting bodies modulate inflammatory mediator expression in mice with polymicrobial sepsis. Nutrition. 2012 Apr 25. [Epub ahead of print] Meng LM, Pai MH, Liu JJ, Yeh SL. Source: School of Nutrition and Health Sciences, Taipei Medical University, Taipei, Taiwan. Abstract OBJECTIVES: Antrodia camphorata (AC) is a traditional Chinese medicine, and the polysaccharides contained within AC (AC-PSs) are reported to possess various biological functions. This study extracted AC-PSs from mycelia and fruiting bodies and evaluated their influences on inflammatory mediator expressions in septic mice. METHODS: There were one normal control (NC) and three experimental groups. The normal control group underwent a sham operation, whereas the experimental groups underwent cecal ligation and puncture (CLP) to induce sepsis. Mice in the experimental groups were further divided into saline, mycelia, and fruiting body treatment groups. Saline or AC-PSs were injected intraperitoneally twice at 0.5 and 1 h after CLP and the mice were sacrificed at 6 or 16 h after sepsis for further analysis. RESULTS: Compared with the normal control group, interleukin (IL)-6, tumor necrosis factor-, IL-10, and monocyte chemotactic protein-1 levels in plasma and/or peritoneal lavage fluid in the septic mice dramatically increased after CLP. The increased levels of these inflammatory mediators in the two AC-PS-treated groups had decreased by 16 h after CLP. 52

Messenger RNA expressions of tumor necrosis factor-, IL-6, and IL-10 in the splenocytes were lower in the 2 AC-PS-treated groups than in the saline group. Consistent with the results, lung nuclear factor-B expressions decreased and less severe interstitial inflammation was observed in the histologic finding after CLP in mice that had received AC-PSs. The fruiting body group had higher white blood cell counts and lower IL-6 levels in the peritoneal lavage fluid 6 h after CLP, whereas the interferon- level was higher 16 h after CLP than in the saline group. These alterations were not found in mice injected with the mycelia extract. CONCLUSION: The administration of AC-PSs from mycelia or fruiting bodies decreased the inflammatory mediator expressions at the location of injury and in the circulation, especially in the late stage of sepsis. AC-PSs from fruiting bodies seemed to be more effective in decreasing the inflammatory response than those from mycelia. These findings suggest that AC-PSs from mycelia and fruiting bodies have potential protective effects against polymicrobial sepsis. Copyright 2012 Elsevier Inc. All rights reserved. PMID: 22541057, [PubMed - as supplied by publisher] 119. Review of Biological and Pharmacological Activities of the Endemic Taiwanese Bitter Medicinal Mushroom, Antrodia camphorata (M. Zang et C. H. Su) Sh. H. Wu et al. (Higher Basidiomycetes). Int J Med Mushrooms. 2012;14(3):241-56. Yue PY, Wong YY, Chan TY, Law CK, Tsoi YK, Leung KS. Source: Department of Biology, Faculty of Science, Hong Kong Baptist University, Kowloon, Hong Kong SAR. Abstract: Antrodia camphorata is an extremely rare fungus native to the forested regions of Taiwan. It is also a traditional Chinese medicine, and Taiwanese aborigines applied it for treating liver diseases and protecting from food and drug intoxication. Scientific studies have demonstrated that A. camphorata crude extracts and pure compounds possess a variety of beneficial functions, such as anti-hypertensive, anti-hyperlipidemic, anti-inflammatory, anti-oxidant, anti-tumor, and immuno-modulatory activities. Recent studies have shown that many of these biological and pharmacological activities can be attributed to various active constituents, including polysaccharides, terpenoids, steroids, lignans, benzoquinone derivatives, benzenoids, and maleic and succinic acid derivatives. A. camphorata has been considered as a novel phytotherapeutic agent. However, detailed mechanistic studies or even clinical trials on A. camphorata are still rare. With the help of modern analytical techniques, it is not surprising that many novel constituents are being identified or fractionated from A. camphorata mycelium and fruiting bodies. This review summarizes the latest published results from A. camphorata research, focusing on the biological and pharmacological activities of the crude extract and known constituents of A. camphorata. PMID: 22577975, [PubMed - in process] 120. Taiwanofungus camphoratus (Syn Antrodia camphorata) Extract and Amphotericin B Exert Adjuvant Effects via Mitochondrial Apoptotic Pathway. Integr Cancer Ther. 2012 Jul 12. [Epub ahead of print] Chen LY, Sheu MT, Liao CK, Tsai FC, Kao WY, Su CH. Source: Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan, ROC. Abstract: The use of multiple drugs in cancer therapy increases the efficacy of the potential therapeutic effects. In this study, the authors investigated the adjuvant effects of an ethanol extract of solid-state cultivated Taiwanofungus camphoratus (TCEE) and amphotericin B (AmB) in the human cancer cell lines RPMI7951 and MG63. Taiwanofungus camphoratus is a well-known Chinese medicine in Taiwan, and AmB is a widely used antifungal agent. The authors demonstrated that TCEE pretreatment followed by AmB treatment effectively inhibited cell growth. The combination of sublethal doses of TCEE and AmB revealed a significant growth inhibitory effect in both cell lines. The combination of TCEE and AmB but not AmB alone induced phosphatidylserine externalization and loss of mitochondrial membrane potential. Cell cycle analyses revealed that combination of TCEE and AmB triggered G2/M arrest and significant apoptosis after 48 hours. These effects were greater than those achieved using TCEE or AmB alone. Furthermore, the authors demonstrated that the drugs increased the levels of p21(Cip1/Waf1) and pro-apoptotic protein Bax and reduced the level of anti-apoptotic protein Bcl-2. Taken together, the results showed that the combination treatment of TCEE and AmB displays strong adjuvant effects, which are indicated by the inhibition of cell proliferation in 2 human cancer cell lines, RPMI7951 and MG63. These findings suggest possible therapeutic applications and alternative medicines using this drug combination. PMID: 22791310, [PubMed - as supplied by publisher] 53

121. Anticancer Effects of Eleven Triterpenoids Derived from Antrodia camphorata. Anticancer Res. 2012 Jul;32(7):2727-34. Lee YP, Tsai WC, Ko CJ, Rao YK, Yang CR, Chen DR, Yang MH, Yang CC, Tzeng YM. Source: Chung Shan Medical University, 110, Section 1, Chien-Kuo North Road, Taichung, Taiwan 40201, R.O.C. Tel: +886 424730022/12415, cyang@csmu.edu.tw. Abstract: Eleven derivatives from Antrodia camphorata were isolated in order to evaluate their selective cytotoxicity toward 14 types of human cancer cell and two non-transformed cell types. Among these triterpenoids, methyl antcinate A (MAA) exhibited the most potent spectrum of anticancer effects in KB cells, four different oral cancer cell lines (TSCCa, GNM, OC-2, and OEC-M1), Panc-1, BT474, PC-3, OVCAR-3, HeLa, and U2OS cells with high selectivity indices (CC(50)/IC(50)). The expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and poly(ADP-ribose) polymerase (PARP) of PC-3 cells tested by western blotting suggested that MAA exerts cell death through the caspase-dependent cascade and the Bax-mediated mitochondrial apoptotic pathway, not only on liver and oral cancer cells but on other types as well, including prostate cancer, in a dose-dependent manner. In addition to MAA, methyl antcinate B, dehydroeburicoic acid, and 15-acetyl-dehydrosulfurenic acid also exhibited significant selective cytotoxic effects to respective cancer cells. Modifications of these triterpenoids may lead to the development of more potent anticancer drugs. PMID: 22753732, [PubMed - in process] 122. Inhibition of Cell Growth and Induction of Apoptosis by Antrodia camphorata in HER-2/neu-Overexpressing Breast Cancer Cells through the Induction of ROS, Depletion of HER-2/neu, and Disruption of the PI3K/Akt Signaling Pathway. Evid Based Complement Alternat Med. 2012;2012:702857. Epub 2012 Jun 3. Lee CC, Yang HL, Way TD, Kumar KJ, Juan YC, Cho HJ, Lin KY, Hsu LS, Chen SC, Hseu YC. Source: Department of Health and Nutrition Biotechnology, Asia University, Taichung 41354, Taiwan. Abstract: Previously, we demonstrated that a submerged fermentation culture of Antrodia camphorata (AC) promotes cell-cycle arrest and apoptosis in human estrogen receptor-positive/negative breast cancer cells. However, whether AC is effective against HER-2/neu-overexpressing breast cancers has not been thoroughly elucidated. In the present study, we showed that AC exhibited a significant cytotoxic effect against HER-2/neu-overexpressing MDA-MB-453 and BT-474 cells. Immunoblot analysis demonstrated that HER-2/neu and their tyrosine phosphorylation were inhibited by AC in a dose-dependent manner. An increase in intracellular reactive oxygen species (ROS) was observed in AC-treated cells, whereas antioxidant N-acetylcysteine (NAC) significantly prevented AC induced HER-2/neu depletion and cell death, which directly indicates that AC-induced HER-2/neu depletion and cell death was mediated by ROS generation. Also, AC significantly downregulated the expression of cyclin D1, cyclin E, and CDK4 followed by the suppression of PI3K/Akt, and their downstream effectors GSK-3 and -catenin. Notably, AC-treatment induced apoptotic cell death, which was associated with sub-G1 accumulation, DNA fragmentation, mitochondrial dysfunction, cytochrome c release, caspase-3/-9 activation, PARP degradation, and Bcl-2/Bax dysregulation. Assays for colony formation also confirmed the growth-inhibitory effects of AC. This is the first report confirming the anticancer activity of this potentially beneficial mushroom against human HER-2/neu-overexpressing breast cancers. PMID: 22701509, [PubMed - in process], PMCID: PMC3371823 123. Triterpenoid-Rich Extract from Antrodia camphorata Improves Physical Fatigue and Exercise Performance in Mice. Evid Based Complement Alternat Med. 2012;2012:364741. Epub 2012 Jul 5. Huang CC, Hsu MC, Huang WC, Yang HR, Hou CC. Source: Graduate Institute of Sports Science, National Taiwan Sport University, Taoyuan 33301, Taiwan. Abstract: Antrodia camphorata (AC) is an endemic mushroom that grows in Taiwan. We investigated the fatigue-alleviating effects of AC on endurance capacity in swim-exercised and weight-loading mice. Male Institute of Cancer Research (ICR) strain mice from 3 groups (n = 10 per group in each test) were orally administered AC fruiting body extract for 7 days at 0, 50, and 200mg/kg/day, designated vehicle, AC-50, and AC-200, respectively. Trend analysis revealed that AC treatments increased grip strength. AC dose-dependently increased swim time, blood glucose, and muscular and hepatic glycogen levels and dose-dependently decreased plasma lactate and ammonia levels and creatine kinase activity. The increase in 54

swimming endurance with AC administration was caused by an increase in liver and muscle glycogen deposition. A. camphorata may have potential for use in ergogenic and antifatigue activities. PMID: 22829854, [PubMed - as supplied by publisher] , PMCID: PMC3398672 124. Inhibition of Na(+)/K(+) -ATPase by Antcins, Unique Steroid-Like Compounds in Antrodia camphorate. Am J Chin Med. 2012;40(5):953-65. Chung TY, Li FY, Chang CI, Jinn TR, Tzen JT. Source: Graduate Institute of Biotechnology, National Chung Hsing University, Taichung 40227, Taiwan. Abstract: The inhibition of Na(+)/K(+) -ATPase by versatile steroid-like compounds contributes to the putative therapeutic effects of many Chinese medicinal cardiac products via the same molecular mechanism triggered by cardiac glycosides. Five major steroid-like compounds, antcin A, B, C, H, and K were isolated from Niuchangchih (Antrodia camphorata), a unique Taiwan mushroom, and all inhibited Na(+)/K(+) -ATPase. Antcin A exhibited significantly higher inhibitory potency than the other four antcins, though weaker than ginsenoside Rh2 . In contrast, cortisone (an analogous steroid with anti-inflammatory effects stronger than antcin A) showed no detectable inhibitory potency. Molecular modeling has shown that antcins bind to Na(+)/K(+) -ATPase with the steroidal skeleton structurally upside-down in comparison with ginsenoside Rh2 . The inhibitory potency of antcin A is attributed to steroidal hydrophobic interaction within the binding pocket and the formation of three hydrogen bonds between its carboxyl group and two cationic residues around the cavity entrance of Na(+)/K(+) -ATPase. The presence of an additional carbonyl or hydroxyl group at C7 of the other four antcins leads to severe repulsion in the hydrophobic pocket, and thus significantly reduces inhibitory potency. It is proposed that antcin A is a bi-functional compound that exerts anti-inflammatory effects and that enhances blood circulation via two different molecular mechanisms. PMID: 22928827, [PubMed - in process] 125. An efficient total synthesis of a potent anti-inflammatory agent, benzocamphorin f, and its anti-inflammatory activity. nt J Mol Sci. 2012;13(8):10432-40. Epub 2012 Aug 21. Liao YR, Kuo PC, Liang JW, Shen YC, Wu TS. Source: Department of Chemistry, National Cheng Kung University, Tainan 701, Taiwan; E-Mails: l3892101@mail.ncku.edu.tw (Y.-R.L.); l36994279@mail.ncku.edu.tw (J.-W.L.). Abstract: A naturally occurring enynyl-benzenoid, benzocamphorin F (1), from the edible fungus Taiwanofungus camphoratus (Antrodiacamphorata) was characterized by comprehensive spectral analysis. It displays anti-inflammatory bioactivity and is valuable for further biological studies. The present study is the first total synthesis of benzocamphorin F and the developed strategy described is a more efficient procedure that allowe the large-scale production of benzocamphorin F for further research of the biological activity both in vitro and in vivo. PMID: 22949872, [PubMed] , PMCID: PMC3431870 126. Taiwanofungus camphorata nitroreductase: cDNA cloning and biochemical characterisation. Food Chem. 2012 Dec 15;135(4):2708-13. doi: 10.1016/j.foodchem.2012.07.024. Epub 2012 Jul 16. Chen CC, Ken CF, Wen L, Chang CF, Lin CT. Source: Institute of Bioscience and Biotechnology and Center of Excellence for Marine Bioenvironment and Biotechnology, National Taiwan Ocean University, Keelung 202, Taiwan. Abstract: Nitroreductases (Nrs) play important roles in redox system via NADPH or NADH as a reductant. A TcNr cDNA encoding a putative Nr was cloned from Taiwanofungus camphorata. A 3-D structural model of the TcNr has been created based on the known structure of BcNr (Bacillus cereus). To characterise the TcNr, the coding region was subcloned into an expression vector and transformed into Escherichia coli. The recombinant His(6)-tagged TcNr was purified by Ni affinity chromatography. The purified enzyme showed a single band at molecular mass of approximately 25kDa on 12% sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The enzyme exhibited Nr activity via ferricyanide assay. The Michaelis constant (K(M)) value for ferricyanide was 0.86mM. The enzyme(')s half-life of deactivation at 45C was 12.3min. The enzyme was most active at pH 6. The enzyme's preferred substrate is 1-chloro-2, 4-dinitrobenzene. Copyright 2012 Elsevier Ltd. All rights reserved. PMID: 22980862, [PubMed - in process] 55

127. Lanostane triterpenoids and sterols from Antrodia camphorata. Phytochemistry. 2012 Sep 18. pii: S0031-9422(12)00378-0. doi: 10.1016/j.phytochem.2012.08.011. [Epub ahead of print] Huang HC, Liaw CC, Yang HL, Hseu YC, Kuo HT, Tsai YC, Chien SC, Amagaya S, Chen YC, Kuo YH. Source: Department of Chinese Pharmaceutical Sciences and Chinese Medicine Resources, China Medical University, Taichung 404, Taiwan. Abstract: Four lanostane triterpenes, 3,7,11-trioxo-5-lanosta-8,24(E)-dien-26-oic acid, methyl 11-3,7-dioxo-5-lanosta-8,24(E)-dien-26-oate, methyl 3,7,11,12,15,23-hexaoxo-5-lanost-8-en-26-oate, and ethyl 3,7,11,12,15,23-hexaoxo-5-lanost-8-en-26-oate, two sterols, (14,22E)-14-hydroxyergosta-7,22-diene-3,6-dione and a steroid named as camphosterol A were isolated from a mixture of fruiting bodies and mycelia of solid cultures of Antrodia camphorata. The (1)H and (13)C NMR spectra of all compounds were fully assigned using a combination of 2D NMR experiments, including COSY, HMQC, HMBC and NOESY sequences. Six compounds were evaluated for cytotoxicity against several human tumor cell lines, all of which has moderate activity. Copyright 2012 Elsevier Ltd. All rights reserved. PMID: 22999074, [PubMed - as supplied by publisher] 128. The Antitumor Activity of Antrodia camphorata in Melanoma Cells: Modulation of Wnt/-Catenin Signaling Pathways. Evid Based Complement Alternat Med. 2012;2012:197309. doi: 10.1155/2012/197309. Epub 2012 Sep 25. Hseu YC, Tsou HT, Kumar KJ, Lin KY, Chang HW, Yang HL. Source: Department of Cosmeceutics, College of Pharmacy, China Medical University, Taichung 40402, Taiwan. Abstract: Antrodia camphorata (AC) is well known in Taiwan as a traditional Chinese medicine. The aim of this study was to investigate whether a fermented culture broth of AC could inhibit melanoma proliferation and progression via suppression of the Wnt/-catenin signaling pathway. In this study, we observed that AC treatment resulted in decreased cell viability and disturbed Wnt/-catenin cascade in B16F10 and/or B16F1 melanoma cells. This result was accompanied by a decrease in the expression of Wnt/-catenin transcriptional targets, including c-Myc and survivin. Furthermore, treatment of melanoma cells with AC resulted in a significant increase in apoptosis, which was associated with DNA fragmentation, cytochrome c release, caspase-9 and -3 activation, PARP degradation, Bcl-2/Bax dysregulation, and p53 expression. We also observed that AC caused G(1) phase arrest mediated by a downregulation of cyclin D1 and CDK4 and increased p21 and p27 expression. In addition, we demonstrated that non- and subcytotoxic concentrations of AC markedly inhibited migration and invasion of highly metastatic B16F10 cells. The antimetastatic effect of AC was further confirmed by reductions in the levels of MMP-2, MMP-9, and VEGF expression. These results suggest that Antrodia camphorata may exert antitumor activity by downregulating the Wnt/-catenin pathways. 129. Lanostanoids from Fungi: A Group of Potential Anticancer Compounds. J Nat Prod. 2012 Oct 23. [Epub ahead of print] R JL, Andjar I, Recio MC, Giner RM. os Source: Departament de Farmacologia, Facultat de Farmacia, Universitat de Valencia , Avenida Vicent Andrs Estells s/n, 46100 Burjassot, Valencia, Spain. Abstract: Lanostanes are a group of tetracyclic triterpenoids derived from lanosterol. They have relevant biological and pharmacological properties, such as their cytotoxic effects via induction of apoptosis. This review compiles the most relevant lanostanoids studied from 2000 to 2011, principally those isolated from Ganoderma lucidum and other related fungi, such as Poria cocos, Laetiporus sulphureus, Inonotus obliquus, Antrodia camphorata, Daedalea dickinsii, and Elfvingia applanata, which have great potential as anticancer agents because of their cytotoxic or apoptotic effects. The compounds were selected on the basis of their proapoptotic mechanisms, through their ability to modify transcriptional activities via nuclear factors or genes and the activation or inhibition of pro- or antiapoptotic proteins; studies based only on their cytotoxicity were excluded from this review in the absence of complementary studies on their mechanisms of action. A total of 81 compounds from Ganoderma lucidum and other species from this genus are included, as well as 96 compounds isolated from other fungi, principally Poria cocos. Some of these compounds were found to arrest the cell cycle in the G(1) phase, increase levels of p53 and Bax, or inhibit the phosphorylation of Erk1/2 or the activation of NF-B and AP-1. Other lanostanes have inhibitory effects on the growth of androgen prostate carcinoma through increasing the expression of p21, which activates the tumor suppressor protein p53, while other compounds have been shown to selectively inhibit topo II activity without affecting topo I. General considerations concerning the chemical structure-biological activities of these 56

compounds are also discussed. PMID: 23092389, [PubMed - as supplied by publisher] 130. Antrodia camphorata ATCC 200183 sporulates asexually in submerged culture. Appl Microbiol Biotechnol. 2012 Oct 28. [Epub ahead of print] Geng Y, He Z, Lu ZM, Xu HY, Xu GH, Shi JS, Xu ZH. Source: Laboratory of Pharmaceutical Engineering, School of Medicine and Pharmaceutics, Jiangnan University, 1800 Lihu Avenue, Wuxi, Jiangsu, 214122, People's Republic of China. Abstract: Antrodia camphorata is a well-known Chinese medicinal mushroom that protects against diverse health-related conditions. Submerged fermentation of A. camphorata is an alternative choice for the effective production of bioactive metabolites, but the effects of nutrition and environment on mycelial morphology are largely unknown. In this study, we show that A. camphorata American Type Culture Collection 200183 can form arthrospores in the end of liquid fermentation. Different morphologies of A. camphorata in submerged culture were analyzed using scanning electron microscopy. The optimal carbon and nitrogen sources for sporulation were soluble starch and yeast extract. We found that a carbon-to-nitrogen ratio (C/N) of 40:1, MgSO(4) (0.5 g/l), KH(2)PO(4) (3.0 g/l), an initial pH5.0, and an inoculum size of 1.510(5) spores/ml led to maximum production of arthroconidia. Our results will be useful in the regulation and optimization of A. camphorata cultures for efficient production of arthroconidia in submerged culture, which can be used as inocula in subsequent fermentation processes. PMID: 23104644, [PubMed - as supplied by publisher] 131. Quality of bread supplemented with mushroom mycelia. Food Chem. 2013 May 1;138(1):70-6. doi: 10.1016/j.foodchem.2012.10.051. Epub 2012 Nov 8. Ulziijargal E, Yang JH, Lin LY, Chen CP, Mau JL. Source: Department of Food Science and Biotechnology, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan, ROC; NCHU-UCD Plant and Food Biotechnology Program, Biotechnology Center, National Chung Hsing University, 250 Kuokuang Road, Taichung 40227, Taiwan, ROC. Abstract: Mushroom mycelia of Antrodia camphorata, Agaricus blazei, Hericium erinaceus and Phellinus linteus were used to substitute 5% of wheat flour to make bread. Bread quality, including specific volume, colour property, equivalent umami concentration (EUC), texture profile analysis, sensory evaluation and functional components, was analysed. Mycelium-supplemented bread was smaller in loaf volume and coloured, and had lower lightness and white index values. White bread contained the lowest amounts of free umami amino acids and umami 5'-nucleotides and showed the lowest EUC value. Incorporating 5% mushroom mycelia into the bread formula did not adversely affect the texture profile of the bread. However, incorporating 5% mushroom mycelia into the bread formula did lower bread's acceptability. After baking, mycelium-supplemented bread still contained substantial amounts of -aminobutyric acid and ergothioneine (0.23-0.86 and 0.79-2.10mg/g dry matter, respectively). Overall, mushroom mycelium could be incorporated into bread to provide its beneficial health effects. Copyright 2012 Elsevier Ltd. All rights reserved. PMID: 23265457, [PubMed - in process] 132. The in vivo antitumor effects on human COLO 205 cancer cells of the 4,7-dimethoxy-5-(2-propen-1-yl)-1,3-benzodioxole (apiole) derivative of 5-substituted 4,7-dimethoxy-5-methyl-l,3-benzodioxole (SY-1) isolated from the fruiting body of Antrodia camphorate. J Cancer Res Ther. 2012 Oct;8(4):532-6. doi: 10.4103/0973-1482.106529. Wei PL, Tu SH, Lien HM, Chen LC, Chen CS, Wu CH, Huang CS, Chang HW, Chang CH, Tseng H, Ho YS. Source: Center of Excellence for Cancer Research; Department of Laboratory Medicine; School of Medical Laboratory Science and Biotechnology, College of Medicine, Taipei Medical University, Taipei, Taiwan. Abstract: Context: The compound 4,7-dimethoxy-5-(2-propen-1-yl)-1,3-benzodioxole (apiole) has been isolated from several different plant species, including Petroselinum sativum. Our recent study found that apiole is a chemical derivative of 4,7-dimethoxy-5-methyl-l,3-benzodioxole (SY-1), which has been isolated from dried Antrodia camphorata (AC ) fruiting bodies, a traditional Chinese medicine with antitumor properties. Aims: Our previous in vitro study demonstrated that apiole inhibits the growth of human colon (COLO 205) cancer cells through the arrest of the cell cycle in G0/G1 phase. The in vivo antitumor effects of apiole were evaluated in this study. Setting and Design: Apiole was administered to mice at 1-30 mg/kg body weight through intraperitoneal (I.P.) injection three times per week (defined as a dosage of 1 -30). 57

Materials and Methods: The in vivo antitumor effects of apiole were evaluated in mice with xenografts of COLO 205 cells. Statistical Analysis: All of the data are reported as the means S.E. Comparisons were performed with a one-way analysis of variance (ANOVA) followed by a Fisher's least significant difference test. Significance was defined as P < 0.05. Results: Apiole (> 1) markedly decreased the growth of COLO 205 human colon cancer cell tumor xenografts in an athymic nude mouse model system through the up-regulation of cell cycle regulators, such as p53, p21/Cip1, and p27/Kip1. The apiole-induced increase in G0/G1 phase cell cycle regulators was also associated with a significant decrease in the expression of cyclins D1 and D3. Surprisingly, statistically significantly higher tumor volumes were observed in mice that received 5 apiole compared with 30 apiole-treated mice (P < 0.05). No gross signs of toxicity were observed (e.g., body weight changes, general appearance, or individual organ effects) in any group. Conclusions: Our results show, for the first time, the promising antitumor effects of apiole against colon tumors in an in vivo xenograft model. PMID: 23361270, [PubMed - in process] 133. Inhibition of Helicobacter pylori CagA-Induced Pathogenesis by Methylantcinate B from Antrodia camphorata. Evid Based Complement Alternat Med. 2013;2013:682418. doi: 10.1155/2013/682418. Epub 2013 Jan 8. Lin CJ, Rao YK, Hung CL, Feng CL, Lane HY, Tzeng DT, Hsu PN, Lai CH, Tzeng YM. Source: Graduate Institute of Clinical Medical Science, China Medical University, Taichung 40402, Taiwan. Abstract: The bacterial pathogen Helicobacter pylori (Hp) is the leading risk factor for the development of gastric cancer. Hp virulence factor, cytotoxin-associated gene A (CagA) interacted with cholesterol-enriched microdomains and leads to induction of inflammation in gastric epithelial cells (AGS). In this study, we identified a triterpenoid methylantcinate B (MAB) from the medicinal mushroom Antrodia camphorata which inhibited the translocation and phosphorylation of CagA and caused a reduction in hummingbird phenotype in HP-infected AGS cells. Additionally, MAB suppressed the Hp-induced inflammatory response by attenuation of NF-B activation, translocation of p65 NF-B, and phosphorylation of IB-, indicating that MAB modulates CagA-mediated signaling pathway. Additionally, MAB also suppressed the IL-8 luciferase activity and its secretion in HP-infected AGS cells. On the other hand, molecular structure simulations revealed that MAB interacts with CagA similarly to that of cholesterol. Moreover, binding of cholesterol to the immobilized CagA was inhibited by increased levels of MAB. Our results demonstrate that MAB is the first natural triterpenoid which competes with cholesterol bound to CagA leading to attenuation of Hp-induced pathogenesis of epithelial cells. Thus, this study indicates that MAB may have a scope to develop as a therapeutic candidate against Hp CagA-induced inflammation. 134. New Anti-Inflammatory Aromatic Components from Antrodia camphorata. Int J Mol Sci. 2013 Feb 26;14(3):4629-39. doi: 10.3390/ijms14034629. Chen YC, Chiu HL, Chao CY, Lin WH, Chao LK, Huang GJ, Kuo YH. Source: Department of Chinese Pharmaceutical Sciences and Chinese Medicine Resources, College of Pharmacy, China Medical University, Taichung 404, Taiwan. kuoyh@mail.cmu.edu.tw. Abstract: Three new benzenoids, 3-isopropenyl-2-methoxy-6-methyl-4,5-methylenedioxy- phenol (1), 2-hydroxy-4,4'-dimethoxy-3,3'-dimethyl-5,6,5',6'-bimethylenedioxybiphenyl (2), 4,4'-dihydroxy-3,3'-dimethoxy-2,2'-dimethyl-5,6,5',6'-bimethylenedioxybiphenyl (3), together with two known benzenoids, 2,3,6-trimethoxy-5-methylphenol (4) and 2,3-methylenedioxy- 4-methoxy-5-methylphenol (5), were isolated from Antrodia camphorata. Our results support that compounds 1-5 potently inhibited LPS (lipopolysaccharide)-induced nitric oxide (NO) production in a dose-dependent manner. The IC(50) values of compounds 1, 3 and 5 were 1.8 0.2, 18.8 0.6 and 0.8 0.3 g/mL, respectively.

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