Romanian Journal of Morphology and Embryology 2007, 48(1):5965

ORIGINAL PAPER
Morphological changes positive correlates
with oxidative stress in COPD. Preliminary
data of an experimental rat model
study and literature review
ELENA DOINA MURRESCU1), ROXANA IANCU2),
MARIA SULTANA MIHAILOVICI1)
1)
2)

Department of Pathology

Department of Pathophysiology

Gr. T. Popa University of Medicine and Pharmacy, Iassy

Abstract
It is well known that nicotine that is a major toxic component of cigarette smoke induces oxidative stress which is responsible for the lung
damages in COPD and cancer. There have been reported some cases of COPD in never smoking patients exposed to air pollutants.
The aim of our study is to evaluate the morphological pulmonary changes in rats exposed to cigarette smoke respectively to solid
combustible smoke and to establish the relationship between the exposure and the level of oxidative stress measured through serum (s)
and pulmonary tissue (l) MDA in rats (TBARS method). Thirty Wistar rats were divided into three groups (n = 10): (1) the control group (C),
(2) the cigarette smoke group (CS), and (3) the solid combustible smoke group (SC). Apart from the control group, these were treated with
solid combustibles smoke or cigarette smoke for six months. We collected blood for serum determination of MDA and the lungs were
removed for histopathological analysis and to determine the levels of malondialdehyde (MDA). The levels of serum and lung MDA were
significantly higher in CS and SC groups compared with C group, but not significantly differences between CS and SC group were
detected. These findings are positively correlated with histopathological changes (squamous metaplasia and clear cell hyperplasia in the
bronchium epithelium, emphysema) found in pulmonary tissue. Preliminary data of our study confirms that not only the cigarette smoke but
also the environmental pollutants are involved in the major pathways of COPD.
Keywords: COPD, oxidative stress, cigarette smoke, solid combustible smoke, rat experimental model.

 Introduction
COPD is an extremely complex disease. Lots of
studies were made during the last 40 years concerning
COPD
pathogenesis,
physiopathology
and
morphological changes but they did not obtain clear and
unitary conclusions. The main risk factor incriminated
in COPD pathogenesis is cigarette smoking but only
15% of active smokers develop airway flow obstruction
and COPD. On the other hand, epidemiological studies
showed that 512% of COPD patients never smoked.
Only a little is known about the pathophysiology
of the constant airway flow obstruction in these
patients. These data suggest the implication of some
factors, other than cigarette smoking in pathogenesis of
COPD [1].
A major impediment in the study of COPD is that
the fundamental morphological changes that produce
the major pulmonary dysfunction are placed in the
peripheral airways, at bronchiole-alveolar junction, that
are difficult to be explored by optic fiber endoscopies.
That is why the animal models had an important role in
the modern era of research in COPD, after Gross P et al.
[2] discovered that the intratracheal administration of
papain determine emphysema. On the other hand,
Laurell CB and Erikkson SE [3] determined that the

deficit of alfa1-antitrypsin determine a high risk for

emphysema. These two findings are the scientific
fundament for the hypothesis elastaseantielastase in the
pathogenesis of emphysema and even after 40 years
animal models are important tools for the studies of the
pulmonary morphological changes in COPD [4].
It has been lately reported COPD cases in patients
who never smoked. A coherent morphological study on
human tissue is practically impossible because of the
major pulmonary dysfunction of the COPD patients that
do not permit optic fiber endoscopies or surgical
procedures. That is why we have proposed to make a
comparative study on experimental rat model consisting
in rats exposed to cigarette smoke, respectively to solid
combustible burning smoke. The aim of our study is to
analyze the morphological changes induced by smoke in
the central and peripheral airway walls, respectively the
pulmonary parenchyma and to evaluate the oxidative
stress.
 Material and methods
We used 30 Wistar adult male rats, divided in three
groups of 10 rats each. The CS (cigarette smoke) lot
was daily exposed to the smoke resulted from burning
of 10 cigarettes.

Elena Doina Murrescu et al.

60

The SC (solid combustible smoke) was exposed to

the smoke resulted from the burning of 300 g of solid
combustible (wood, coal). The third lot, C (control) was
unexposed. The exposure was made in one 50/50/50 cm
precinct, 60 minutes daily.
The slaughter of the animals was performed two
days after the last exposure, confirm the EC rules.
The rats were anaesthetized with Thiopental and
then sacrificed. Blood and tissues (lungs, heard,
kidneys) were removed. Lung tissue was processed
by usual histopathological technique: 10% formalin
fixed, paraffin embedded, HematoxylinEosin and
Van Gieson stained.
We have considerate malondialdehide (MDA) as a
marker of oxidative stress induced by the cigarette
smoke respectively the solid combustible smoke
exposure. MDA levels were determined using TBARS
method. Unfortunately we did not have the technical
possibility to perform spirometry tests.
 Results
The rats medium weight was at the beginning of the
study 250 g. All the animals were standard feed during
the whole period of experiment. In the end, medium
weight of the C lot was 350g, 289g for CS lot,
respectively 297g for the SC lot. We can see a loss of
weight in smoke exposed rats (Figure 1).
Both the two smoke exposed lots presented
lesions at the large and small, peripheral, airways and
also at the pulmonary parenchyma comparative with
those that are characteristic for COPD. We have found
clear cell hyperplasia (Figures 2 and 3) and squamous
metaplasia (Figures 4 and 5) of the small airway
mucosal epithelium, mononuclear inflammatory
infiltrate in the small airway walls (Figures 6 and 7) and
alveolar walls destructions corresponding for
emphysema (Figures 8 and 9).
The serum level of MDA was 6.29+ 2.76 nmoles/ml
in the C lot; in the pulmonary homogenate the MDA
level was 0.42+ 0.23 nmoles/mg protein.
The serum level of MDA was 8.06+ 1.32 nmoles/ml
in the CS lot; in the pulmonary homogenate the MDA
level was 0.84+ 0.82 nmoles/mg protein.
The serum level of MDA was 7.3+ 1.34 nmoles/ml
in the SC lot; in the pulmonary homogenate the MDA
level was 0.79+ 0.56 nmoles/mg protein.
 Discussions
There is no longitudinally study on COPD
experimental model until 1997, when Wright JL et al.
[5] showed that the pulmonary function of the rat
normally decrease during 12 months, because of rat
aging and the cigarette smoke accelerates this effect
similarly in human, as also demonstrated Gold DR et al.
in 1996 [6].
Wright JL et al. [5] demonstrated that the chronic
cigarette smoke exposure determine airflow obstruction.
The vital capacity (VC) and the forced vital capacity
(FVC) are increased in the smoke exposed animals
comparative with the control, according to the increased
statically pulmonary compliance.

Usually, the vital capacity is decreased in human

COPD patients. This paradox in animals is explained by
transpulmonary positive pressure of the air income
rather than active inspire [7].
The forced expiratory volume in one second (FEV1)
physiologically decrease during the 4th and the 12th
month, but the decrease is more significant in the
exposed group than in the unexposed one. FEV1/FVC
decreased significantly in the smoke exposed group in
the 4th8th month period meanwhile the aging
decreasing of the airflow was produced in the 8th12th
month period.
It is evident that is a net discrepancy between the
smoking effects on the pulmonary volumes and
compliance and, respectively on the airflow, pulmonary
volumes and compliance being precocious influenced
comparative with airflow possible because of the
damaged pulmonary matrix.
Some authors suggested in their studies that the
chronic cigarette smoke exposure determine a dynamic
alteration of the lung collagen with secondary
pulmonary dysfunction [8, 9]. Snider GL et al. have
demonstrated using biochemical methods, increased
levels of the pulmonary collagen that positive correlated
with the decrease of the FEV1 in experimental model
emphysema [10].
Wright JL et al. (1997) sustain that the alteration of
the pulmonary volumes followed by decreasing of the
airflow indicates a remodeling process of the matrix in
rats [5]. It is well known that the matrix is damaged
during lifetime [11]; changes observed in cigarette
smoke exposed rats are analogue to the rapidly aging
process like in smoking humans. [6]
In their study, Wright and Churg showed that rats
were used in lot of COPD studies [6]. Huber GL et al.
[12] and Heckman CA and Dalbey WE [13] have
exposed rats for 6 to 30 months. Both of them determine
a small but significant raise of the aerial space volume
at a 10 cigarettes per day exposure level and
demonstrated the decrease of the elastic recoil.
Heckman CA and Dalbey WE have noticed that the
volumetric proportion of the aerial space was not
significantly increased when the exposure level
decreased at seven cigarettes per day, even at prolonged
period [13].
Ofulue AF et al. found a relatively stabile increase
of the aerial space (130%) after six months of 10 per
day cigarette smoke exposure [14].
Rubio ML et al. exposed rats to the two cigarettes
smoke per day for 10 weeks and morphometrically
demonstrated the thickening of the bronchiolar walls in
bronchioles less than 100 m in diameter that was
associated with an abnormal N2 elimination curve [15].
Huber GL et al. [12] and Heckman CA and
Dalbey WE [13] also demonstrated the existence of an
inflammatory
mononuclear
infiltrate
with
peribronchiolar distribution. Huber GL et al. [12] found
an increased metaplastic secretor cells in the tracheal
epithelium. Also, Hayashi M et al. showed increased
mucous glands volume after 30 consecutive days of
smoke exposure, data that sustain the results of the
Jones and all study from 1973 [16, 17].

Morphological changes positive correlates with oxidative stress in COPD. Preliminary data of an experimental rat model

61

Figure 1 Comparative evolution of the weight

of the three groups

Figure 2 Pulmonary morphological changes in CS group.

Clear cell hyperplasia

Figure 3 Pulmonary morphological changes in SC group.

Clear cell hyperplasia

Figure 4 Pulmonary morphological changes in CS group.

Squamous metaplasia of the small airway epithelium

Figure 5 Pulmonary morphological changes in SC group.

Squamous metaplasia of the small airway epithelium

Figure 6 Pulmonary morphological changes in CS group.

Mononucleate inflammatory infiltrate in the
small airway wall

62

Elena Doina Murrescu et al.

Figure 7 Pulmonary morphological changes in SC group.

Mononucleate inflammatory infiltrate in the
small airway wall

Figure 8 Pulmonary morphological changes in CS group.

Emphysema

9
8
7
6
5
4
3
2
1
0

6.29

8.06

7.3
nm oles/m L

nmoles/mL

Figure 9 Pulmonary morphological changes in SC group.

Emphysema

CS

SC

0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
0.1
0

0.84

0.79

CS

SC

0.42
C

GROUPS

GROUPS

Figure 10 Serum values of MDA are significantly lower

in control group comparing the CS and SC groups.
There are no significant differences for serum
MDA between CS and SC group

Figure 11 MDA levels in pulmonary tissue are significantly

higher in both SC and CS groups comparing the control
group. The insignificantly difference between SC and CS
group appears also in pulmonary tissue as well as in serum

Morphological changes positive correlates with oxidative stress in COPD. Preliminary data of an experimental rat model

Shore S et al. [18] and Finlay GA et al. [19] reported

COPD attributes emphysema or bronchitis, on rat
experimental models. The emphysema or chronic
bronchitis in rats was generally obtained using chemical
substances and they do not reproduce exactly the
biological and pathological aspects of the human COPD
as chronic inflammation. Kodavanti UP et al. [20]
obtained emphysema using elastase instillation and the
persistent chronic inflammation of bronchi with sulfur
dioxide. Finally, the lesions and clinical manifestations
were similar with those of human COPD.
Lots of irritant substances were used to induce
COPD in experimental models: lipopolysaccharides,
cadmium chloride, O3, silica dust. Of course, the
cigarette smoke exposure is the one who determine the
complex pulmonary changes in COPD. Lots of species
were exposed to cigarette smoke during these years:
dog, rabbit, Guineea pig and rodents [21].
Like in human patients with COPD, the weight of
the animals with chronic exposure to smoke is lower
than of the unexposed during the whole period of
experiment [7, 22, 23].
In our study, rats that were exposed in the same
conditions to the cigarette smoke respectively to solid
combustible smoke had a similar body weight evolution.
It is unanimous accepted the loss of the muscular mass
in COPD, probably through apoptotic mechanisms.
Recently, the developing of COPD is associated
with intense oxidative stress, respectively with reduced
antioxidant resources [24, 25]. Important from this point
of view are the oxidant-antioxidant balance
disequilibrium and the role of the oxidative metabolism
in the systemic effects of COPD. Systemic and
pulmonary oxidative metabolism disturbances that
determine loss of muscular mass and carcinogenesis are
obviously in COPD patients [26].
The oxidative stress determines increasing in
proteolysis susceptibility through amino acid chains
changes resulting in protein aggregates and peptide
connections cleavage. Human plasmatic proteins
contain carbonil groups and loose sulfhidril groups as
result of the action of the saturated and unsaturated
aldehides contained in the cigarette smoke after
cigarette smoke exposure [27, 2830].
The plasmatic proteins can be degraded by the
reactive oxygen species (RSA) through nitration and
oxidation. The smokers have higher serum levels of
nitric proteins as fibrinogen, transferyn, ceruloplasmin
and plasminogen respectively oxydated proteins
comparing with non-smokers [31].
Cigarette smoke is a rich source of oxidant agents.
Cigarette smoke tar (particles that lay down of the filter)
contains 1018 spin/gram tar. Generates radicals are
sufficiently stable to permit their detection through ESR
(electron spin resonance). Oxidant agents from the
cigarette smoke are semiquinone, carbon and nitric
reactive species, reactive olefins and dienes, NO.
Oxidative stress may be quantified through
measurement of some biomarkers.
The most convincing method in the direct
measurement of the oxygen radicals in the pulmonary
tissue and in the exhaled air but this method is also the

63

most difficult because of the high reactivity of the short

life species free oxygen radicals.
On the other hand, it is difficult to use direct
techniques and ESR on the lung tissue. The alternative
for these direct methods is the measurement of the
oxygen radical effects on the variety of pulmonary bio
molecules, lipids, proteins or DNA [32].
Free radicals generate lipid peroxidation resulting in
lipidic radicals who react with oxygen in aerobic cells
resulting peroxyl radical. The peroxyl radical starts up a
reaction chain through which the polyunsaturated fat
acids (free or belonging to lipids) are transformed in
lipidic hydroperoxids. Lipid peroxidation determine
membrane dysfunctions, inactivates membrane sites of
the receptors and enzymes, increase the membrane
permeability [33].
The reaction between the lipid hydroperoxids with
the antioxidants (-tocopherol), iron or copper ions, or
haemoglobin, results in gaseous hydrocarbonated and
unsaturated aldehydes (malondialdehide) [34].
Lipid peroxidation products have increased
plasmatic and pulmonary lavage liquid levels in healthy
smokers and in emphysema, chronic bronchitis and
asthma patients. The high level of the lipid peroxidation
products positive correlates with the small airway
obstruction degree in COPD patients [35].
COPD is considerate lately a systemic disease. One
of the systemic manifestations of COPD is the presence
in COPD patients blood of oxidative stress markers.
This is morphological reflected in high sequestration of
neutrophils in pulmonary microcirculation during
smoking and COPD exacerbation periods, oxidant
mediated phenomenon.
Cell membrane polyunsaturated lipids and the fatty
acids are major targets for the free radicals attack
resulting in lipid peroxidation, a process that continues
as a chain reaction and generates peroxides and
aldehydes. Lipid peroxidation reaction products may be
measured in the body fluids as substances that react
with the thiobarbituric acid (TBARS thiobarbituric
acid reactive substances).
Plasmatic level of TBARS is significantly increased
in healthy smokers and in COPD patients comparing
with healthy nonsmokers. Oxidative stress measured as
TBARS plasmatic level inversely correlated with the
percentage of the predicted VEMS indicating the fact
that the lipid peroxidation is associated with airflow
limitation [36].
The data analyze showed that the serum MDA levels
are significantly lower in the C lot than in the CS and
SC lots. There are no significant differences between
the two smoke exposed lots (Figure 10).
Concerning the pulmonary tissue MDA level, the
value in the C lot is again significantly lower than in the
two smoke exposed lots. The differences between the
two smoke exposed lots are insignificant (Figure 11).
The comparative increased of serum and lung tissue
MDA levels in the two chronic smoke exposed lots,
significantly higher than those stand out in the M lot,
indicates a high level of the oxidative stress induced by
the environmental pollutants. The increased serum
MDA suggests the whole body implication in COPD.

Elena Doina Murrescu et al.

64

Actually, COPD appears to be a general disease not an

only one organ one.
Although the majority of information about the
pathogenesis of COPD is focused on the emphysema
development, is well-known that COPD includes also
chronic bronchitis and small airway lesions. There are
not so many informations concerning these aspects but,
presumably, factors that initiate the inflammation and
the effects of the proteolitic and oxidant aggression are
also implicated. Emphysema animal model elastaseinduced is associated with airway epithelium clear cell
hyperplasia and mucous hyper secretion that is
characteristic for chronic bronchitis [36]. The consistent
alveolar wall destructions in rats were a surprise for us
considering that the literature data suggests the rat
seems to be the most resistant to emphysema [21].
Histopathology aspects positive correlate with the
serum and tissue levels of MDA.
 Conclusions
These preliminary data of our study strongly suggest
the fact that the environmental pollutants could also be
implicated in pathogenesis of COPD. This kind of
aggression could be an explication for the cases of
COPD in patients that never smoked.
It becomes more obviously that COPD is a complex
disease that implicates the whole body. The very
numerous studies made during the last 50 years in their
tendency of considering COPD as a pulmonary disease
and the lack of a unitary vision determined the existence
still of a lot of obscure aspects concerning COPD
pathogenesis and, secondarily, the inefficiency of the
treatment.
References
[1] BIRRING S. S., BRIGHTLING C. E., BRADDING P.,
ENTWISLE J. J., VARA D. D., GRIGG J., WARDLAW A. J.,
PAVORD I. D., Clinical, radiologic, and induced sputum
features of chronic obstructive pulmonary disease in
nonsmokers. a descriptive study, Am J Respir Crit Care
Med, 2002, 166(8):10781083.
[2] GROSS P., PFITZER E., TOLKER M., BABYAK M. A.,
KASCHAK M., Experimental emphysema: its production with
papain in normal and silicotic rats, Arch Environ Health,
1965, 11:5058.
[3] LAURELL C. B., ERIKSSON S. E., The electrophoretic alphaglobulin pattern of serum in alpha-antitrypsin deficiency,
Scand J Clin Invest, 1963, 15:132140.
[4] SHAPIRO S. D., Animal models for COPD, Chest, 2000,
117(5 Suppl 1):223S227S.
[5] WRIGHT J. L., SUN J. P., VEDAL S., A longitudinal analysis of
pulmonary function in rats during a 12 month cigarette
smoke exposure, Eur Respir J, 1997, 10(5):11151119.
[6] GOLD D. R., WANG X., WYPIJ D., SPEIZER F. E., WARE J. H.,
DOCKERY D. W., Effects of cigarette smoking on lung
function in adolescent boys and girls, N Engl J Med, 1996,
335(13):931937.
[7] WRIGHT J. L., CHURG A., Cigarette smoke causes
physiologic and morphologic changes of emphysema
in the guinea-pig, Am Rev Respir Dis, 1990,
142(6 Pt 1):14221428.
[8] LAROS C. D., KUYPER C. M., The pathogenesis
of pulmonary emphysema (II), Respiration, 1976,
33(5):325348.
[9] MACKLEM P. T., EIDELMAN D., Re-examination of the elastic
properties of emphysematous lungs, Respiration, 1990,
57(3):187192.

[10] SNIDER G. L., LUCEY E. C., FARIS B., JUNG-LEGG Y.,

STONE P. J., FRANZBLAU C., Cadmium-chloride-induced airspace enlargement with interstitial pulmonary fibrosis
is not associated with destruction of lung elastin.
Implications for the pathogenesis of human emphysema,
Am Rev Respir Dis, 1988, 137(4):918923.
[11] TURINO G. M., The lung parenchyma: a dynamic matrix,
Am Rev Respir Dis, 1985, 132(6):13241334.
[12] HUBER G. L., DAVIES P., ZWILLING G. R., POCHAY V. E.,
HINDS W. C., NICHOLAS H. A., MAHAJAN V. K., HAYASHI M.,
FIRST M. W., A morphologic and physiologic bioassay for
quantifying alterations in the lung following experimental
chronic inhalation of tobacco smoke, Bull Eur Physiopathol
Respir, 1981, 17(2):269327.
[13] HECKMAN C. A., DALBEY W. E., Pathogenesis of lesions
induced in rat lung by chronic tobacco smoke inhalation,
J Natl Cancer Inst, 1982, 69(1):117129.
[14] OFULUE A. F., KO M., ABBOUD R. T., Time course of
neutrophil and macrophage elastinolytic activities in
cigarette smoke induced emphysema, Am J Physiol, 1998,
275(6 Pt 1):L1134L1144.
[15] RUBIO M. L., SANCHEZ-CIFUENTES M. V., ORTEGA M.,
PECES-BARBA G., ESCOLAR J. D., VERBANCK S., PAIVA M.,
GONZALEZ MANGADO N., N-acetylcysteine prevents cigarette
smoke induced small airways alterations in rats,
Eur Respir J, 2000, 15(3):505511.
SORNBERGER
G. C.,
HUBER
G. L.,
[16] HAYASHI M.,
Morphometric analysis of tracheal gland secretion and
hypertrophy in male and female rats after experimental
exposure to tobacco smoke, Am Rev Respir Dis, 1979,
119(1):6773.
[17] WRIGHT J. L., CHURG A., A model of tobacco smokeinduced airflow obstruction in the guinea pig, Chest, 2002,
121(5 Suppl):188S191S.
[18] SHORE S., KOBZIK L., LONG N. C., SKORNIK W.,
VAN STADEN C. J., BOULET L., RODGER I. W., PON D. J.,
Increased airway responsiveness to inhaled methacholine
in a rat model of chronic bronchitis, Am J Respir Crit
Care Med, 1995, 151(6):19311938.
[19] FINLAY G. A., ODONNELL M. D., OCONNOR C. M.,
HAYES J. P., FITZGERALD M. X., Elastin and collagen
remodeling in emphysema: a scanning electron microscopy
study, Am J Pathol, 1996, 149(4):14051415.
[20] KODAVANTI U. P., JACKSON M. C., LEDBETTER A. D.,
STARCHER B. C., EVANSKY P. A., HAREWOOD A.,
WINSETT D. W., COSTA D. L., The combination of elastase
and sulfur dioxide exposure causes COPD-like lesions in
the rat, Chest, 2000, 117(5 Suppl 1):299S302S.
[21] DAWKINS P. A., STOCKLEY R. A., Animal models of
chronic obstructive pulmonary disease, Thorax, 2001,
56(12):972977.
[22] WRIGHT J. L., CHURG A., Smoke-induced emphysema in
guinea pigs is associated with morphometric evidence
of collagen breakdown and repair, Am J Physiol, 1995,
268(1 Pt 1):L17L20.
[23] WALDUM H. L., Nilsen O. G., NILSEN T., RORVIK H.,
SYVERSEN V., SANVIK A. K., HAUGEN O. A., TORP S. H.,
BRENNA E., Long-term effects of inhaled nicotine, Life Sci,
1996, 58(16):13391346.
[24] WHITE C. W., REPINE J. E., Pulmonary antioxidant defense
mechanisms, Exp Lung Res, 1985, 8(23):8196.
[25] HEFFNER J. E., REPINE J. E., Pulmonary strategies
of antioxidant defense, Am Rev Respir Dis, 1989,
140(2):531554.
[26] BOOTS A. W., HAENEN G. R. M. M., BAST A., Oxidant
metabolism in chronic obstructive pulmonary disease,
Eur Respir J, 2003, 22(46 Suppl):14S27S.
[27] STADTMAN E. R., Metal ion-catalyzed oxidation of proteins:
biochemical mechanism and biological consequences,
Free Radic Biol Med, 1990, 9(4):315325.
[28] REZNICK A. Z., CROSS C. E., HU M. L., SUZUKI Y. J.,
KHWAJA S., SAFADI A., MOTCHNIK P. A., PACKER L.,
HALLIWELL B., Modification of plasma proteins by cigarette
smoke as measured by protein carbonyl formation,
Biochem J, 1992, 286(Pt 2):607611.

Morphological changes positive correlates with oxidative stress in COPD. Preliminary data of an experimental rat model
[29] ONEILL C. A., HALLIWELL B., VAN DER VLIET A., DAVIS P. A.,
PACKER L., TRITSCHLER H., STROHMAN W. J., RIELAND T.,
CROSS
C. E.,
REZNICK
A. Z.,
Aldehyde-induced
protein modifications in human plasma: protection by
glutathione and dihydrolipoic acid, J Lab Clin Med, 1994,
124(3):359370.
[30] NAGLER R., LISCHINSKY S., DIAMOND E., DRIGUES N.,
KLEIN I., REZNICK A. Z., Effect of cigarette smoke on salivary
proteins and enzyme activities, Arch Biochem Biophys,
2000, 379(2):229236.
[31] PIGNATELLI B., LI C. Q., BOFFETTA P., CHEN Q., AHRENS W.,
NYBERG F., MUKERIA A., BRUSKE-HOHLFELD I., FORTES C.,
CONSTANTINESCU V., ISCHIROPOULOS H., OHSHIMA H.,
Nitrated and oxidized plasma proteins in smokers and lung
cancer patients, Cancer Res, 2001, 61(2):778784.
[32] REPINE J. E., BAST A., LANKORST I., Oxidative stress in
chronic obstructive pulmonary disease. Oxidative
Stress Study Group, Am J Respir Crit Care Med, 1997,
156(2 Pt 1):341357.

65

[33] HALLIWELL B., CHIRICO S., Lipid peroxidation: its

mechanism, measurement and significance, Am J Clin Nutr,
1993, 57(5 Suppl):715S725S.
[34] HAGEMAN J. J., BAST A., VERMEULEN N. P., Monitoring of
oxidative free radical damage in vivo: analytical aspects,
Chem Biol Interact, 1992, 82(3):243293.
[35] PETRUZZELLI S., HIETANEN E., BARTSCH H., CAMUS A. M.,
MUSSI A., ANGELETTI C. A., SARACCI R., GIUNTINI C.,
Pulmonary lipid peroxidation in cigarette smokers and lung
cancer patients, Chest, 1990, 98(4):930935.
[36] MACNEE W., Oxidative stress and lung inflammation in
airways disease, Eur J Pharmacol, 2001, 429(13):195207.

Corresponding author
Elena Doina Murrescu, Assistant, MD, Department of Pathology, Gr. T. Popa University of
Medicine and Pharmacy, 16 University Street, 700 115 Iassy, Romania; Phone +40745638 427,
E-mail: doinamurarescu@yahoo.com

Received: January 15th, 2007

Accepted: March 20th, 2007