Chapter 1: Cellular Responses to Stress and Toxic Insults:

Adaptation, Injury, and Death

INTRODUCTION during which new but altered steady states are achieved 
PATHOLOGY – is the study (logos) of disease (pathos) allowing the cell to survive and continue to function
 Devoted to the study of the structural, biochemical, and
functional changes in cells, tissues, and organs that underlie
disease
A. GENERAL PATHOLOGY – concerned with the reaction of cells and
tissues to abnormal stimuli and to inherited defects, which are the
main causes of disease
B. SYSTEMIC PATHOLOGY – examines the alterations in specialized
organs and tissues that are responsible for disorders that involve
these organs

FOUR ASPECTS of a DISEASE PROCESS

1. Etiology – cause
2. Pathogenesis – mechanism of development
3. Molecular and Morphologic changes – biochemical and structural
alteration induced in cells and organs of the body
4. Clinical Manifestations – functional consequences of the changes

ETIOLOGY or CAUSE
Two Major Classes of Etiologic Agents
1. Genetic
2. Acquired

PATHOGENESIS
 Sequence of events in the response of cells or tissues to the
 The Adaptive Response may consist of:
etiologic agent, from the initial stimulus to the ultimate
1. Hypertrophy – increase in the size of cells and functional
expression of the disease
activity
 One of the main domains of pathology
2. Hyperplasia – increase in the number of cells
3. Atrophy – decrease in the size and metabolic activity of cells
MOLECULAR and MORPHOLOGIC CHANGES
4. Metaplasia – change in the phenotype of cells
 Structural alterations in cells or tissues that are either
characteristics of a disease or diagnostic of an etiologic agent
CELLULAR RESPONSES TO INJURY
Nature of Injurious Stimuli Cellular Response
FUNCTIONAL DERANGEMENTS and CLINICAL MANIFESTATIONS
 End result of genetic, biochemical, and structural changes in cells ALTERED Physiological Stimuli;
Cellular Adaptations
and tissues are functional abnormalities, which lead to the Some Nonlethal Injurious Stimuli
clinical manifestations (signs and symptoms) of disease, as well as Increased demand, increased
Hyperplasia, hypertrophy
its progress (clinical course and outcome) stimulation
Decreased nutrients, decreased
Atrophy
 Rudolf Virchow – father of Modern Pathology stimulation
 Study of the causes, mechanisms, and morphologic and Chronic irritation (physical or
Metaplasia
biochemical correlates of cell injury chemical)
 Injury to cells and to extracellular matrix ultimately leads to tissue Reduced Oxygen Supply: Chemical
Cellular Injury
and organ injury  determine the morphologic and clinical Injury; Microbial Infection
patterns of disease Acute reversible injury
Acute and transient
Cellular swelling fatty change
OVERVIEW: CELLULAR RESPONSES to STRESS and NOXIOUS STIMULI Irreversible injury  cell death
 The normal cell is confined to a fairly narrow range of function Progressive and severe (including
Necrosis
and structure by its: DNA damage)
Apoptosis
a. State of metabolism, differentiation, and specialization Metabolic alterations, Genetic or Intracellular accumulations;
b. Constraints of neighboring cells Acquired; Chronic Injury Calcification
c. Availability of metabolic substrates Cumulative Sublethal Injury over
 Homeostasis – steady state long life span
Cellular aging
 Adaptations – reversible functional and structural responses to
more severe physiologic stresses and some pathologic stimuli,
 Cell injury – if the limits of adaptive responses are exceeded or if
cells are exposed to injurious agents or stress, deprived of
essential nutrients, or become compromised by mutations that
affect essential cellular constituents
 Reversible up to a certain point
 Irreversible injury – stimulus persists or is severe enough
from the beginning
 Cell death
 Increased workload – most common stimulus for hypertrophy of
muscle  muscle cells synthesize more proteins and the number
of myofilaments increases  increases the amount of force each
myocyte can generate  increases the strength and work
capacity of the muscles
 Physiologic growth of the uterus during pregnancy as a hormone-
induced increase in the size of an organ

Mechanisms of Hypertrophy
 Hypertrophy is the result of increased production of cellular
proteins
 Can be induced by the linked actions of:
a. Mechanical sensors (triggered by increased work load)
b. Growth factors (including TGF-β, insulin-like growth factor-1
[IGF-1], fibroblast growth factor)
 The relationship between normal, adapted, reversibly injured, and
c. Vasoactive agents (α-adrenergic agonist, endothelin-1,
dead myocardial cells
angiotension II)
 Cell death – end result of progressive cell injury
 Two main biochemical pathways involved in muscle hypertrophy
 One of the most crucial events in the evolution of disease in
a. Phosphoinositide 3-kinase / Akt pathway (more important in
any tissue or organ
 Results from diverse causes: physiologic hypertrophy)
b. Signaling downstream of G-protein-coupled receptors (more
a. Ischemia (reduced blood flow)
important in pathologic hypertrophy)
b. Infections
c. Toxins  Hypertrophy – may also be associated with a switch of contractile
 Normal and essential process of embryogenesis, the proteins from adult to fetal or neonatal forms
development of organs, and the maintenance of homeostasis a. α isoform of myosin heavy chain replaced by slower, more
 Two principal pathway of Cell Death energetically economical contraction of β isoform
a. Necrosis b. Atrial Natriuretic Factor (ANF) – peptide hormone that
b. Apoptosis causes salt secretion by the kidney  decreases blood
 Autophagy – an adaptive cellular response triggered by volume and pressure  reduce hemodynamic load
nutrient deprivation; may also culminate in cell death
 Metabolic derangements in cells and sublethal, chronic injury may
be associated with intracellular accumulations of a number of
substances, including proteins, lipids, and carbohydrates
 Calcium – often deposited at sites of cell death  pathologic
calcification
 Aging – accompanied by characteristic morphologic and
functional changes in cells

ADAPTATIONS of CELLULAR GROWTH and DIFFERENTIATION

 Adaptations are reversible changes in the size, number,
phenotype, metabolic activity, or functions of cells in response to
changes in their environment

HYPERTROPHY
 Refers to an increase in the size of cells, resulting in an increase in
the size of the organ
 Has NO NEW cells, just larger cells
 Due to the synthesis of more structural components of the cells
 Nondividing cells increase tissue mass by hypertrophy
 Biochemical Mechanisms of Myocardial Hypertrophy
 Selective hypertrophy – subcellular organelle’s hypertrophy
 Barbiturates  hypertrophy of the smooth endoplasmic
reticulum (SER) in hepatocytes  increases the amount of
enzymes (cytochrome P450 mixed function oxidases) 
detoxify the drugs
HYPERPLASIA
 An increase in the number of cells in an organ or tissue, usually
resulting in increased mass of the organ or tissue
 Can occur together with hypertrophy and may be triggered by the
same external stimulus
 Takes place if the cell population is capable of dividing 
increasing the number of cells
A. Physiologic Hyperplasia
1. Hormonal hyperplasia
 Increases the functional capacity of a tissue when needed
 E.g. proliferation of the glandular epithelium of the female
breast at puberty and during pregnancy, usually accompanied
by enlargement (hypertrophy) of the glandular epithelial cells
2. Compensatory hyperplasia
 Increases tissue mass after damage or partial resection
B. Pathologic Hyperplasia
 Caused by excesses of hormones or growth factors acting on
target cells
 E.g. endometrial hyperplasia as an abnormal hormone-induced
hyperplasia brought about by absolute or relative increase in the
amount of estrogen  hyperplasia of the endometrial glands
 E.g. benign prostatic hyperplasia – hyperplasia in response to
androgens
 Hyperplasia is distinct from cancer, but pathologic hyperplasia
constitutes a fertile soil in which cancerous proliferation may
eventually arise
 A characteristic response to certain viral infection, such as
papillomaviruses, which causes skin warts and several mucosal
lesions composed of masses of hyperplastic epithelium
Mechanisms of Hyperplasia
 Result of growth factor-driven proliferation of mature cells and, in
some cases, by increased output of new cells from tissue stem
cells
ATROPHY
 Reduced size of an organ or tissue resulting from a decrease in
cell size and number
A. Physiologic atrophy
 Common during normal development
 E.g. Notochord and thyroglossal duct, undergo atrophy during
fetal development
 E.g. Uterus decreases in size after parturition
B. Pathologic atrophy
 Depends on the underlying cause and can be local or generalized
Common Causes of Atrophy
 Decreased workload (atrophy of disuse)  initial decrease in cell
size is reversible once activity is resumed
 With more prolonged disuse, skeletal muscle fibers decrease
in number as well as in size
 Loss of Innervation (Denervation atrophy)  normal metabolism
and function of skeletal muscle are dependent on its nerve supply
 Diminished blood supply  a decrease in blood supply (ischemia)
to a tissue as a result of slowly developing arterial occlusive
disease results in atrophy of the tissue
 Senile atrophy – progressive atrophy of the brain because of
reduced blood supply as a result of atherosclerosis
 Inadequate nutrition – profound protein-calorie malnutrition
(marasmus) is associated with the use of skeletal muscle as a
source of energy after other reserves such as adipose stores have
been depleted
 Cachexia – marked muscle wasting; also seen in patients
with chronic inflammatory diseases and cancer
 Loss of endocrine stimulation – many hormone – responsive
tissues are dependent on endocrine stimulation for normal
metabolism and function
 Pressure – tissue compression for any length of time can cause
atrophy
 Result of ischemic changes caused by compromise of the
blood supply by the pressure exerted by the expanding mass
 A. Normal brain of a young adult. B. Atrophy of the brain in an 82
year old male with atherosclerotic cerebrovascular disease,
resulting in reduced blood supply. Note that loss of brain
substance narrows the gyri and widens the sulci. The meninges
have been stripped from the right half of each specimen to reveal
the surface of the brain
 Initial response: Decrease in cell size and organelles  reduce the
metabolic needs of the cell sufficiently to permit its survival
 Atrophic muscle contains FEWER Mitochondria and myofilaments
and a REDUCED amount of Rough ER
 Early in the process atrophic cells may have diminished function,
but they are not dead
Mechanisms of Atrophy
 Atrophy results from decreased protein synthesis and increased
protein degradation in cells
 Reduced metabolic activity  decrease protein synthesis 
nutrient deficiency and disuse  ubiquitin ligases (UBIQUITIN –
PROTEASOME PATHWAY)  attach the small peptide ubiquitin to
cellular proteins and target these proteins for degradation in
proteasomes
 Also responsible for the accelerated proteolysis seen in a
variety of catabolic conditions including cancer cachexia
 Autophagy – also accompanied in atrophy
 Increases the number of autophagic vacuoles
 “self-eating”; the process in which the starved cell eats its
own components in an attempt to find nutrients and survive
 Autophagic vacuoles – membrane bound vacuoles that
contain fragments of cell components
METAPLASIA
 A reversible change in which one differentiated cell type
(epithelial or mesenchymal) is replaced by another cell type
 May represent an adaptive substitution of cells that are sensitive
to stress by cell types better able to withstand the adverse
environment
 Columnar to squamous – most common epithelial metaplasia
 Occurs in the respiratory tract in response to chronic
irritation
 Stones in the excretory ducts of the salivary glands, pancreas,
or bile ducts may also cause replacement of the normal
secretory columnar epithelium by stratified squamous
epithelium
 The influences that predispose to metaplasia, if persistent, may
initiate malignant transformation in metaplasmnic epithelium
 Metaplasia of columnar to squamous epithelium. A. Schematic
diagram. B. Metaplasia of columnar epithelium to squamous
epithelium in a bronchus
 Metaplasia from squamous to columnar type may also occur, as
in Barrett’s esophagus, in which the esophageal squamous
epithelium is replaced by intestinal-like columnar cells under the
influence of refluxed gastric acid  typical glandular
(adeno)carcinoma
 Connective tissue metaplasia – formation of cartilage, bone,
adipose tissue (mesenchymal tissues) in tissues that normally do
not contain these elements  myositis ossificans  bone
formation in muscle which occurs after intramuscular hemorrhage
Mechanisms of Metaplasia
 Result of a reprogramming of stem cells that are known to exist in
normal tissues, or of undifferentiated mesenchymal cells present
in connective tissue
 Differentiation of stem cells generated by cytokines, growth
factors, and extracellular matrix components in the cell’s
environment
OVERVIEW of CELL INJURY and CELL DEATH
 Cell injury results when cells are stressed so severely that they are
no longer able to adapt or when cells are exposed to inherently
damaging agents or suffer from intrinsic abnormalities
a. Reversible cell injury – in early stages or mild forms of injury, the
functional and morphologic changes are REVERSIBLE if the
damaging stimulus is removed
 HALLMARKS of Reversible Injury
1. Reduced oxidative phosphorylation with resultant
depletion of energy stores in the form of ATP
2. Cellular swelling caused by changes in ion
concentrations and water influx
3. Intracellular organelles, mitochondria and the
cytoskeleton show alterations
b. Cell death – with continuing damage the injury becomes
irreversible  cannot recover  dies
1. Necrosis – damage to membranes is severe, lysosomal
enzymes enter the cytoplasm  digest the cell  cellular
contents leak out
 ALWAYS a pathologic process
2. Apoptosis – cell’s DNA or proteins are damaged beyond
repair
 Form of cell death that is characterized by nuclear
dissolution, fragmentation of the cell without complete
loss of membrane integrity, and rapid removal of the
cellular debris
 Serves many normal functions and is not necessarily
associated with cell injury
CAUSES of CELL INJURY
1. Oxygen Deprivation
 Hypoxia – deficiency of oxygen  reduced aerobic oxidative
respiration  cell injury
 Causes of Hypoxia
a. Reduced blood flow (ischemia)
b. Inadequate oxygenation of the blood due to
cardiorespiratory failure
c. Decreased oxygen-carrying capacity of the blood, as in
anemia or carbon monoxide poisoning or after severe blood
loss
2. Physical agents
a. Mechanical trauma
b. Extremes of temperature (burns and deep cold)
c. Sudden changes in atmospheric pressure
d. Radiation
e. Electric shock
3. Chemical agents and drugs
a. Glucose or salt in hypertonic concentrations
b. Oxygen at high concentrations is toxic
c. Poisons
- Arsenic
- Cyanide
- Mercuric salts
d. Environmental and air pollutants, insecticides, & herbicides
e. Industrial and occupational hazards, carbon monoxide and
asbestos
f. Recreational drugs e.g. alcohol
g. Ever-increasing variety of therapeutic drugs
4. Infectious agents Reversible injury is characterized by:
a. Viruses 1. Generalized swelling of the cell and its organelles
b. Ricketsiae 2. Blebbing of the plasma membrane
c. Bacteria 3. Detachment of ribosomes from the ER
d. Fungi 4. Clumping of nuclear chromatin
e. Parasites
f. Large tapeworms Morphologic changes are associated with:
5. Immunologic Reactions 1. Decreased generation of ATP
a. Endogenous self-antigens  autoimmune diseases 2. Loss of cell membrane integrity
b. Immune reactions to many external agents (microbes and 3. Defects in protein synthesis
environmental substances) 4. Cytoskeletal damage
6. Genetic derangements 5. DNA damage
a. Congenital malformations associated with Down syndrome
caused by a chromosomal anomaly
b. Decrease life span of red blood cells caused by a single amino
acid substitution in hemoglobin in sickle cell anemia
c. Deficiency of functional proteins  enzyme defects in inborn
errors of metabolism
d. Accumulation of damaged DNA or misfolded proteins
e. Variations in the genetic makeup  influence the
susceptibility of cells to injury by chemicals and other
environmental insults
7. Nutritional imbalances
a. Protein – calorie deficiencies
b. Vitamin deficiencies
c. Nutritional excesses
- Excess of cholesterol  atherosclerosis
- Obesity  diabetes and cancer
d. Composition of the diet

MORPHOLOGIC ALTERATIONS IN CELL INJURY  Schematic illustration of the morphologic changes in cell injury
culminating in necrosis or apoptosis

FEATURES of NECROSIS and APOPTOSIS

Feature Necrosis Apoptosis
Cell size Enlarged (swelling) Reduced (shrinkage)
Fragmentation into
Pyknosis  Karyorrhexis
Nucleus nucleosome – size
 Karyolysis
fragments
Intact; altered structure,
Plasma
Disrupted especially orientation of
membrane
lipids
Cellular Enzymatic digestion; Intact; may be released
contents may leak out of cell in apoptotic bodies
Adjacent
Frequent No
inflammation
Often physiologic, means
of eliminating unwanted
Physiologic or Invariably pathologic
cells; may be pathologic
pathologic (culmination of
after some forms of cell
 Sequential development of biochemical and morphologic changes role irreversible cell injury)
injury, especially DNA
in cell injury damage
 Cells may become rapidly nonfunctional after the onset of injury,
although they are still viable, with potentially reversible damage REVERSIBLE INJURY
 A longer duration of injury may eventually lead to irreversible Two features of reversible injury:
injury and cell death 1. Cellular swelling – appears whenever cells are incapable of
 Irreversible biochemical alteration may cause cell death, and maintaining ionic and fluid homeostasis and is the result of
typically this precedes ultrastructural, light microscopic, and energy-dependent ion pumps in the plasma membrane
grossly visible morphologic changes 2. Fatty change – occurs in hypoxic injury and various forms of toxic
or metabolic injury
  Manifested by the appearance of lipid vacuoles in the  C. Proximal tubular cell showing late injury  irreversible  swollen
cytoplasm mitochondria containing electron-dense deposits, expected to
 Seen mainly in cells involved in and dependent on fat contain precipitated calcium and proteins
metabolism, such as hepatocytes and myocardial cells
NECROSIS
Morphology  Result of denaturation of intracellular proteins and enzymatic
 Cell swelling – the first manifestation of almost all forms of injury digestion of the lethally injured cell
to cells  Necrotic cell – unable to maintain membrane integrity and their
 When it affects many cells, it causes: contents often leak out  inflammation in the surrounding tissue
a. Pallor  Enzymes that digest the necrotic cell are derived from the
b. Increased turgor lysosomes of of the dying cells themselves and from the
c. Increased in weight of the organ lysosomes of leukocytes that are called in as part of the
 Microscopic: Small clear vacuoles may be seen within the inflammatory reaction
cytoplasm  distended and pinched off segments of the ER   The earliest histologic evidence of myocardial necrosis does not
Hydropic change or vacuolar degeneration become apparent until 4 to 12 hours later
 Ultrastructural changes of reversible cell injury:  Decrease plasma membrane integrity  cardiac enzymes and
1. Plasma membrane alteration – blebbing, blunting, and loss proteins rapidly released from necrotic muscle  can be detected
of microvilli in the blood as early as 2 hours after myocardial cell necrosis
2. Mitochondrial changes – swelling and the appearance of
small amorphous densities Morphology
3. Dilation of the ER – detachment of polysomes;  Increased eosinophilia  loss of cytoplasmic RNA and in part of
intracytoplasmic myelin figures may be present desaturated cytoplasmic proteins
4. Nuclear alterations – disaggregation of granular and fibrillar  More glassy homogenous appearance than do normal cells 
elements result of loss of glycogen particles
 Cytoplasm become vacuolated and appears moth-eaten
 Myelin figures  dead cells replaced by large, whorled
phospholipid masses; derived from damage cell membranes
 Phospholipid precipitates  phagocytosed by other cells 
further degraded into fatty acids  calcification of fatty acid
residues  generation of calcium soaps
 Electron microscopy, necrotic cells are characterized by:
a. Discontinuities in plasma and organelle membranes
b. Marked dilation of mitochondria with the appearance of
large amorphous densities
c. Intracytoplasmic myelin figures
 Morphologic changes in reversible cell injury and necrosis d. Amorphous debris
 A. Normal kidney tubules with viable epithelial cells e. Aggregates of fluffy material probably representing
 B. Early (reversible) ischemic injury showing surface blebs, denatured protein
increased eosinophilia of cytoplasm, and swelling of occasional  Nuclear changes  due to nonspecific breakdown of DNA
cells 1. Basophilia of the chromatin may fade (KARYOLYSIS), a
 C. Necrosis (irreversible injury) of epithelial cells, with loss of change that presumably reflect loss of DNA because of
nuclei, fragmentation of cells, and leakage of contents enzymatic degradation of endonucleases
2. PYKNOSIS  characterized by nuclear shrinkage and
 Ultrastructure features of increased basophilia  chromatin condenses into a solid,
reversible and irreversible cell shrunken basophilic mass
injury (necrosis) in the kidney 3. KARYORRHEXIS  pyknotic nucleus undergoes
 A. Normal epithelial cell of fragmentation  after a day or two, nucleus totally
the proximal kidney tubule  disappears
abundant microvilli lining the
luminal surface COAGULATIVE NECROSIS
 B. Epithelial cell of the  Form of necrosis in which the architecture of dead tissues is
proximal tubule showing early preserved for a span of at least some days
cell injury resulting from  Affected tissues exhibit a firm texture
reperfusion following  Injury denatures not only structural proteins but also enzymes
ischemia  microvilli are lost and so blocks the proteolysis of the dead cells  eosinophilic,
and have been incorporated anucleate cells may persist for days or weeks
in apical cytoplasm  blebs  Necrotic cells are removed by phagocytosis of the cellular debris
have formed and are by infiltrating leukocytes and by digestion of the dead cells by the
extruded in the lumen  action of lysosomal enzymes of the leukocytes
swollen mitochondria during
ischemia
 Ischemia caused by obstruction in a vessel may lead to
coagulative necrosis of the supplied tissue in all organs except the
BRAIN
 INFARCT – localized area of coagulative necrosis

LIQUEFACTIVE NECROSIS
 Characterized by digestion of the dead cells  transformation of
the tissue into a liquid viscous mass
 Seen in focal bacteria, or occasionally, fungal infections 
microbes stimulate the accumulation of leukocytes and the
liberation of enzymes from these cells
 PUS – creamy yellow appearance of the necrotic material due to
dead leukocytes
 Ex: hypoxic death cells within the CNS
FAT NECROSIS
 Focal areas of fat destruction, resulting from release of activated
pancreatic lipases into the substance of the pancreas and the
peritoneal cavity
 Occurs in the calamitous abdominal emergency  ACUTE
PANCREATITIS  pancreatic enzymes leak out of acinar cells and
liquefy the membranes of fat cells in the peritoneum
 Released lipases split the triglyceride esters contained within fat
cells  fatty acids combine with calcium to produce grossly
visible chalky-white areas (fat saponification)  enable the
surgeon and the pathologist to identify the lesions

FIBRINOID NECROSIS
GANGRENOUS NECROSIS
 Special form of necrosis usually seen in immune reactions
 Not a specific pattern of cell death
involving blood vessels
 Applied to a limb, generally the lower leg, that has lost its blood
 Occurs when complexes of antigens and antibodies are deposited
supply and has undergone necrosis (coagulative necrosis)
in the walls of arteries
involving multiple tissue planes
 Deposits of these “immune complexes” together with fibrin that
 When bacterial infection is superimposed there is more
has leaked out of vessels  bright pink and amorphous
liquefactive necrosis  actions of degradative enzymes in the
appearance in H&E stains FIBRINOID
bacteria and the attracted leukocytes  WET GANGRENE
 Ex: immunologically mediated vasculitis syndromes
CASEOUS NECROSIS
 Encountered most often in foci of tuberculous infection
 “Caseous” – cheeselike is derived from the friable white
appearance of the area of necrosis
 Collection of fragmented or lysed cells and amorphous granular
debris enclosed within a distinctive inflammatory border
 GRANULOMA – characteristic of a focus of inflammation
 Dystrophic calcification – if necrotic cells and cellular debris are
not promptly destroyed and reabsorbed, they tend to attract
calcium salts and other minerals and to become calcified
MECHANISMS of CELL INJURY
1. The cellular response to injurious stimuli depends on the nature
of the injury, its duration, and its severity
2. The consequences of cell injury depend on the type, state, and
adaptability of the injured cell
 Cell’s nutritional and hormonal status and its metabolic
needs
 Genetic variations
3. Cell injury results from different biochemical mechanisms acting
on several essential cellular components
 Cellular components that are most frequently damaged by
injurious stimuli include:
a. Mitochondria
b. Cell membranes
c. Machinery of protein synthesis and packaging
d. DNA in nuclei
4. Any injurious stimuli may simultaneously trigger multiple
interconnected mechanisms that damage cells
 The principal mechanisms of cell injury, and their biochemical and
functional effects
DEPLETION of ATP
 ATP Depletion and Decreased ATP synthesis are frequently
associated with both hypoxic and chemical (toxic) injury
 Two Ways ATP is Produced:
1. Oxidative phosphorylation of adenosine diphosphate – major
pathway; reaction that results in reduction of oxygen by the
electron transfer system of mitochondria
2. Glycolitic pathway – can generate ATP in the absence of oxygen
using glucose derived either from body fluids or from the
hydrolysis of glycogen
 Major Causes of ATP Depletion
1. Reduced supply of oxygen and nutrients
2. Mitochondrial damage
3. Actions of some toxins (cyanide)
Depletion of ATP to 5% to 10% of normal levels has widespread
effects on many critical cellular systems
a. The activity of the plasma membrane energy-dependent sodium
+ +
pump (ouabain-sensitive Na K ATPase) is reduced  sodium
enters and accumulates inside cells and potassium diffuse out 
CELL SWELLING  dilation of the ER
b. Cellular energy metabolism is altered
 Supply of oxygen to cells is reduced  Ischemia  oxidative
phosphorylation ceases  decrease in cellular ATP and
increase AMP  stimulate phosphofructokinase and
phosphorylase activities  increased rate of ANAEROBIC
glycolysis  glycogen stores are rapidly depleted 
accumulation of LACTIC acid and inorganic phosphates from
hydrolysis of phosphate esters  reduce intracellular pH 
decreased activity of many cellular enzymes
2+ 2+
c. Failure of the Ca pump  influx of Ca  damaging effects on
numerous cellular component
d. With prolonged or worsening depletion of ATP, structural
disruption of the protein synthetic apparatus occurs 
detachment of ribosomes from the rough ER and dissociation of
polysomes  reduction in protein synthesis
e. In cells deprived of oxygen or glucose, proteins may become
misfolded  trigger a cellular reaction (UNFOLDED PROTEIN
RESPONSE)  culminates in cell injury and even death
f. Irreversible damage to mitochondrial and lysosomal membranes
 NECROSIS
 Functional and morphologic consequences of decreased
intracellular ATP during cell injury
 Morphologic changes are indicative of reversible cell injury
 Further depletion of ATP results in cell death, typically by necrosis
MITOCHONDRIAL DAMAGE
 Mitochondria – cell’s suppliers of life-sustaining energy in the
form of ATP
 Are also critical players in cell injury and death
 Can be damaged by:
a. Iincreases of cytosolic Ca
2+
b. Reactive oxygen species
c. Oxygen deprivation
d. All types of injurious stimuli (hypoxia and toxins)
Two Major Consequences of Mitochondrial Damage
1. Mitochondrial damage often results in the formation of a high-
conductance channel in the mitochondrial membrane
(MITOCHONDRIAL PERMEABILITY TRANSITION PORE)
 Opening of the conductance channel  loss of mitochondrial
membrane potential  failure of oxidative phosphorylation
and progressive depletion of ATP  necrosis of the cell
 Cyclophilin D – a protein structural component of the
mitochondrial permeability transition pore  target of the
immunosuppressive drug cyclosporin
2. The mitochondria also sequester between their outer and inner
membranes several proteins that are capable of activating
apoptotic pathways; cytochrome c and proteins that indirectly
activate apoptosis inducing enzymes (CASPASES)
 Increased permeability of the outer mitochondrial
membrane  leakage of proteins into the cytosol  death
by apoptosis
 Consequences of mitochondrial dysfunction, culminating in cell
death by necrosis or apoptosis
INFLUX of CALCIUM and LOSS of CALCIUM HOMEOSTASIS
 Cytosolic free calcium is normally maintained at very low
concentration (-0.1 μmol) compared with extracellular levels of
1.3 mmol
 Most intracellular calcium is sequestered in mitochondria and the
ER
 Ischemia and certain toxins  increase in cytosolic calcium
concentration  release of Ca2+ from the intracellular stores 
increased influx across the plasma membrane  cellular injury
Mechanisms
2+
1. Accumulation of Ca in mitochondria results in opening of the
mitochondrial permeability transition pore  failure of ATP
generation
2+
2. Increased cytosolic Ca activates a number of enzymes:
a. Phospholipases  membrane damage
b. Proteases  break down both membrane and cytoskeletal
proteins
c. Endonucleases  responsible for DNA and chromatin
fragmentation
d. ATPases  hastening ATP depletion
3. Increased intracellular Ca2+ levels also result in the induction of
apoptosis by:
a. Direct activation of caspases
b. Increasing mitochondrial permeability
 Role of increased cytosolic calcium in cell injury
ACCUMULATION OF OXYGEN+DERIVED FREE RADICALS (OXIDATIVE
STRESS)
 Cell injury induced by free radicals, particularly reactive oxygen
species, is an importance mechanism of cell damage in many
pathologic conditions, such as:
1. Chemical and radiation injury
2. Ischemia-reperfusion injury
3. Cellular aging
4. Microbial killing by phagocytes
 FREE RADICALS – chemical species that have a single unpaired
electron in an outer orbit
 Initiate autocatalytic reactions  conversion to free radicals
 propagating chain of damage
 REACTIVE OXYGEN SPECIES (ROS) – type of oxygen derived free
radical whose role in cell injury is well established
 Produced normally in cells during mitochondrial respiration
and energy generation
 Degraded and removed by cellular defense systems
 Also produced in large amounts of leukocytes, particularly
NEUTROPHILS and MACROPHAGES, as mediators for
destroying microbes, dead tissue, and other unwanted
substances
 Increased production of ROS / scavenging systems are ineffective
 excess of free radicals  OXIDATIVE STRESS
 Oxidative stress has been implicated in a wide variety of b. Vitamin E
pathologic processes, including: c. Ascorbic acid
a. Cell injury d. Glutathione
b. Cancer 2. Iron and copper can catalyze the formation of ROS  can be
c. Aging minimized by binding of the ions to storage and transport proteins
d. Degenerative disease (Alzheimer disease) to minimize the formation of ROS
a. Transferrin
PROPERTIES OF THE PRINCIPAL FREE RADICALS b. Ferritin
INVOLVED IN CELL INJURY c. Lactoferrin
-
O2 H2O2 OH ONOO- d. Ceruloplasmin
MECHANISMS of PRODUCTION 3. Enzyme that act a free-radical-scavenging systems and breaks
Produced by down H202 and O2-
Generated - a. Catalase – presents in peroxisomes, decomposes H2O2
Incomplete interaction of O2
from H2O by (2H2O2  O2 + 2H2O)
reduction of O2 and NO
Generated by hydrolysis, e.g.
during oxidative generated by NO b. Superoxide dismutases (SODs) – are found in many cell
SOD from O2- by radiation; - -
phosphorylation;
and by oxidases from H2O2 by
synthase in many types and convert O2 to H2O2 (2O2 + 2H  H2O2 + O2)
by phagocyte
in peroxisomes Fenton
cell types 1. Manganese-SOD  mitochondria
oxidase in (endothelial 2. Copper-Zinc-SOD  cytosol
reaction; from
leukocytes - cells, leukocytes,
O2 c. Glutathione peroxidase – also protects against injury by
neurons, others
catalyzing free radical breakdown
MECHANISMS OF INACTIVATION
Conversion to
H2O and O2 by
Conversion to
catalase Conversion to
Conversion to HNO2 by
(peroxisomes), H2O by
H2O2 and O2 by peroxiredoxins
glutathione glutathione
SOD (cytosol,
peroxidase peroxidase
mitochondria)
(cytosol,
mitochondria)
PATHOLOGIC EFFECTS
Stimulates
production of Can be
Most reactive
degradative converted to
- oxygen-derived
enzymes in OH and OCL ,
free radical;
leukocytes and which destroy
principal ROS Damages lipids,
other cells; may microbes and
responsible for proteins, DNA
directly damage cells; can act
damaging
lipids, proteins, distant from
lipids, proteins,
DNA; acts close site of
and DNA
to site of production
production

GENERATION of FREE RADICALS

1. The reduction-oxidation reactions that occur during normal
metabolic processes
2. Absorption of radiant energy (ultraviolet light, x-rays)
3. Rapid burst of ROS are produced in activated leukocytes during
inflammation
4. Enzymatic metabolism of exogenous chemicals or drugs can
generate free radicals that are not ROS but have similar effects
(CCl4  CCl3)
5. Transition metals such as irons and copper donate or accept free
electrons during intracellular reactions and catalyze free radical
2+ 3+
formation as in the Fenton Reaction (H2O2 + Fe  Fe + OH +
-
OH )
6. Nitric oxide (NO), an important chemical mediator generated by
endothelias cells, macrophages, neurons, and other cell types, can
act as a free radical and can also be converted to a highly reactive
peroxynitrite anion (ONOO-) as well as NO2 and NO3-
REMOVAL OF FREE RADICALS
1. Antioxidants either block the initiation of free radical formation
or inactivate free radicals
a. Vitamin A
PATHOLOGIC EFFECTS of FREE RADICALS
1. Lipid peroxidation in membranes
 Oxidative damage is initiated when the double bonds in
unsaturated fatty acids of membrane lipids are attacked by O2
derived free radicals, particularly by OH
 Lipid – free radical interactions yield peroxides, unstable and
reactive  autocatalytic chain reaction (PROPAGATION) 
extensive membrane damage
2. Oxidative modification of proteins – free radicals promote:
a. Oxidation of amino acid side chains
b. Formation of protein – protein cross-linkages
c. Oxidation of the protein backbone
Oxidative modification of proteins may:
a. Damage the active sites of enzymes
b. Disrupt the conformation of structural proteins
c. Enhance proteasomal degradation of unfolded or misfolded
proteins
3. Lesions in DNA
 Free radicals are capable of causing:
a. Single- and double-strand breaks in DNA
b. Cross-linking of DNA strand
c. Formation of adducts
DEFECTS in MEMBRANE PERMEABILITY
 Consistent feature of most forms of cell injury (except apoptosis)
 Membrane damage may affect the functions and integrity of all
cellular membranes
Mechanisms of Membrane Damage
 Ischemic cells membrane defects may be the result of ATP
depletion and calcium-mediated activation of phospholipases
 Can be damaged directly by various bacterial toxins, viral proteins,
lytic complement components, and a variety of physical and
chemical agents
1. Reactive Oxygen Species – cause injury to cell membranes by
LIPID peroxidation
2. Decreased phospholipid synthesis – as a consequence of
defective mitochondrial function or hypoxia  decrease the
production of ATP  affect energy – dependent enzymatic
activities
 May affect ALL cellular membranes, including the
mitochondria themselves
3. Increased phospholipid breakdown – due to activation of
endogenous phospholipases by increased levels of cytosolic and
2+
mitochondrial Ca
 Leads to accumulation of lipid breakdown products,
including unesterified free fatty acids, acyl carnitine, and
lysophospholipids  detergent effect on membranes
 Can also insert into the lipid bilayer of the membrane or
exchange with membrane phospholipids, potentially causing
changes in permeability and electrophysiologic alterations
4. Cytoskeletal abnormalities – cytoskeletal filaments serve as
anchors connecting the plasma membrane to the cell interior
 Activation of proteases by increased cytosolic calcium 
damage to elements of the cytoskeleton
 Cell swelling  damage  detachment of the cell
membrane from the cytoskeleton  susceptible to
stretching and rupture
 Mechanisms of membrane damage in cell injury
 Decrease O2 and increase cytosolic Ca2+ are typically seen in
ischemia but may accompany other forms of cell injury
 Reactive oxygen species (ROS) which are produced on reperfusion
of ischemic tissues, also cause membrane damage
CONSEQUENCES of MEMBRANE DAMAGE
1. Mitochondrial membrane damage – results in opening of the
mitochondrial permeability transition pore  decreased ATP 
release of proteins  apoptotic death
2. Plasma membrane damage – results in loss of osmotic balance
and influx of fluids and ions and loss of cellular contents  cells
may leak metabolites that are vital for the reconstitution of ATP
 depleting energy stores
3. Injury to lysosomal membranes – results in leakage of their
enzymes into the cytoplasm and activation of the acid hydrolases
in the acidic intracellular pH of the injured cell. Contains:
a. RNases
b. DNases
c. Proteases activation of these enzymes leads to
d. Phosphatases enzymatic digestion of proteins, RNA,
e. Glucosidases DNA, and glycogen  cells die by
f. Cathepsins necrosis
DAMAGE to DNA and PROTEINS
 “Point of No Return” – the damage becomes irreversible
Two Phenomena consistently characterize irreversibility
1. The ability to reverse mitochondrial dysfunction (lack of oxidative
phosphorylation and ATP generation) even after resolution of the
original injury
2. Profound disturbances in membrane function
 Leakage of intracellular proteins through the damaged cell
membrane and ultimately into the circulation provides a means of
detecting tissue-specific cellular injury and necrosis using blood
serum samples
 Cardiac muscle  Creatine kinase and Troponin
 Liver  (bile duct epithelium) Alkaline phosphatase
 Hepatocytes  Transaminases
Clinico-Pathologic Correlations:
Selected Examples of Cell Injury and Necrosis
ISCHEMIC and HYPOXIC INJURY
 Most common type of cell injury in clinical medicine
 Hypoxia – reduced oxygen availability
 Ischemia – supply of oxygen and nutrients is decreased most
often because of reduced blood flow as a consequence of a
mechanical obstruction in the arterial system
 Also be caused by reduced venous drainage
 Compromises the delivery of substrates for glycolysis
 Not only is AEROBIC metabolism compromised but
ANAEROBIC energy generation also stops after glycolytic
substrates are exhausted
 Glycolysis is inhibited by the accumulation of metabolites
that would have been removed otherwise by blood flow
 Ischemia tends to cause more rapid and severe cell and
tissue injury than does hypoxia in the absence of ischemia
Mechanisms of Ischemic Cell Injury
 Oxygen tension within the cell decreases  loss of oxidative
phosphorylation and decrease generation of ATP  failure of the
sodium pump with loss of potassium, influx of sodium and water,
2+
influx of Ca  cell swelling
 Progressive loss of glycogen and decreased protein synthesis
 Cytoskeleton disperses  loss of ultrastructural features such as
microvilli and the formation of “blebs” at the cell surface
 “Myelin figures” – derived from degenerating cellular
membranes, may be seen within the cytoplasm (autophagic
vacuoles) or extracellularly
 Mitochondria – usually swollen  result of loss of volume control
in these organelles
 ER – remains dilated
 Entire cell – markedly swollen
 Increased concentrations of water, sodium, and chloride and a
decreased concentration of potassium
 If oxygen is restored, all of these disturbances are reversible
 If ischemia persist, IRREVERSIBLE injury and necrosis ensue
 Irreversible injury is associated morphologically with:
a. Severe swelling of mitochondria
b. Extensive damage to plasma membranes (myelin figures)
c. Swelling of lysosomes
 Dead cells may become replaced by large masses composed of
phospholipids in the form of myelin figures  either
phagocytosed by leukocytes or degraded further into fatty acids
 Calcification of fatty acid residues may occur  formation of
calcium soaps
Protective Responses to Hypoxic Stress
1. Induction of a transcription factor called hypoxia-inducible factor-
1  promotes new blood vessel formation  stimulates cell
survival pathways  enhances anaerobic glycolysis
2. Ischemic (and traumatic) brain and spinal cord injury  transient
induction of hypothermia (reducing the core body temperature to
92°F)
a. reduces the metabolic demands of the stressed cells
b. decreases cell swelling
c. suppresses the formation of free radicals
d. inhibits the host inflammatory response
ISCHEMIA-REPERFUSION INJURY
 Under certain circumstances, when blood flow is restores to cells
that have been ischemic but have not died, injury is paradoxically
exacerbated and proceeds at an accelerated pace
 Reperfused tissues may sustain loss of cells in addition to the
cells that are irreversibly damaged at the end of ischemia 
ischemia reperfusion injury
Mechanisms of Ischemia-Reperfusion Injury
1. New damage may be initiated during reoxygenation by increased
generation of reactive oxygen and nitrogen species from
parenchymal and endothelial cells and from infiltrating leukocytes
 May be produced in reperfused tissue as a result of
mitochondrial damage  incomplete reduction of oxygen or
because of the action of:
a. Oxidases in leukocytes
b. Endothelial cells
c. Parenchymal cells
 Calcium may also enter reperfused cells  damaging various
organelles, including mitochondria  increasing the
production of free radicals
2. Ischemic injury is associated with inflammation as a result of the
production of cytokines and increased expression of adhesion
molecules by hypoxic parenchymal and endothelial cells  recruit
circulating neutrophils to reperfused tissue
 Treatment with antibodies that block cytokines or adhesion
molecules  reduce the extent of the injury
3. Activation of the complement system may contribute to ischemia-
reperfusion injury
 Involved in host defense and is an important mechanism of
immune injury
 IgM antibodies  deposit in ischemic tissues  blood flow
resumed  complement proteins bind to the deposited
antibodies  activation  more cell injury and inflammation
CHEMICAL (TOXIC) INJURY
 Major limitation to drug therapy
 Liver – frequent target of drug toxicity
 Toxic Liver Injury – most frequent reason for terminating the
therapeutic use or development of a drug
Chemicals induce cell injury by one of two general mechanisms:
1. Some chemicals can injure cells directly by combining with critical
molecular components
Example:
a. Mercuric chloride poisoning  mercury binds to the
sulfhydryl groups of cell membrane proteins  causing
increased membrane permeability  inhibition of ion
transport  cells of the GI tract and kidney damage
b. Cyanide poisons mitochondrial cytochrome oxidase 
inhibits oxidative phosphorylation
c. Antineoplastic chemotherapeutic agents and antibiotic drugs
 direct cytotoxic effects  cell damage
2. Most toxic chemical are not biologically active in their native form
but must be converted to reactive toxic metabolites  acts on
target molecules
a. Cytochrome P450 mixed-function oxidases in the smooth ER
of the liver and other organs
b. Formation of free radicals  lipid peroxidation 
membrane damage & cell injury
c. Direct covalent binding to membrane proteins and lipids also
contribute
d. CCl4  CCl3 (highly reactive free radical)  lipid peroxidation
 damaged many cellular structures
e. Acetaminophen, analgesic drug  converted to a toxic
product during detoxification in the liver  cell injury
Apoptosis
 A pathway of cell death that is induced by a tightly regulated
suicide program in which cells destined to die activate enzymes
that degrade the cells’ own nuclear DNA and nuclear and
cytoplasmic proteins
 Apoptotic bodies – fragments of apoptotic cells that break up;
contain portions of the cytoplasm and nucleus
 “Falling off” – morphologic appearance of membrane-bound
fragments derived from cells
Characteristics of Apoptosis
1. Loss of membrane integrity
2. Enzymatic digestion of cells
3. Leakage of cellular contents
4. A host reaction
 Apoptosis and necrosis sometimes coexist  apoptosis may
progress to necrosis
CAUSES of APOPTOSIS  Fragmentation into membrane-bound apoptotic bodies composed
Apoptosis in Physiologic Situations of cytoplasm and tightly packed organelles, with or without
 Death by apoptosis is a normal phenomenon that serves to nuclear fragments
eliminate cells that are no longer needed, and to maintain steady Phagocytosis of apoptotic cells or cell bodies, usually by macrophages
number of various cell populations in tissues  Rapidly ingested by phagocytes and degraded by the phagocyte’s
1. The programmed destruction of cells during embryogenesis, lysosomal enzymes
including:
a. Implantation
b. Organogenesis
c. Developmental involution
d. Metamorphosis
 “Programmed cell death” – denote death of specific cell types at
defined times during the development of an organism
2. Involution of hormone-dependent tissues upon hormone
withdrawal
a. Endometrial cell breakdown during menstrual cycle
b. Ovarian follicular atresia in menopause
c. The regression of the lactating breast after weaning
d. Prostatic atrophy after castration
3. Cell loss in proliferating cell populations
a. Immature lymphocytes in the bone marrow and thymus that
fail to express useful antigen receptors
b. B lymphocytes in germinal centers
c. Epithelial cells in intestinal crypts  HOMEOSTASIS
4. Elimination of potentially harmful self-reactive lymphocytes  A. Reduced in size of cell and contains brightly eosinophilic
prevent reactions against one’s own tissues cytoplasm and a condensed nucleus
5. Death of host cells that have served their useful purpose B. Nuclei with peripheral crescents of compacted chromatin and
a. Neutrophils in an ACUTE INFLAMMATORY RESPONSE others that are uniformly dense or fragmented
b. Lymphocytes at the end of an IMMUNE RESPONSE C. 1. Blebbing and formation of apoptotic bodies
2. Nuclear fragmentation
Apoptosis in Pathologic Conditions 3. Activation of caspase-3
 Apoptosis eliminates cells that are injured beyond repair without
eliciting a host reaction  limiting collateral tissue damage  On histologic examination, apoptotic cell appears as a round or
1. DNA Damage oval mass of intensely eosinophilic cytoplasm with fragments of
a. Radiation dense nuclear chromatin
b. Cytotoxic anticancer drugs  Apoptosis does NOT elicit INFLAMMATION, making it more
c. Hypoxia difficult to detect histologically
2. Accumulation of misfolded proteins – due to mutations in the
genes encoding proteins or because of extrinsic factors Biochemical Features of Apoptosis
 ER Stress – accumulation of the misfolded proteins in the ER Activation of Caspases
and culminates in apoptotic cell death  Based on two properties of this family of enzymes:
3. Cell death in certain infections a. “c” = cysteine protease
4. Pathologic atrophy in parenchymal organs after duct obstruction b. “aspase” = unique ability of these enzymes to cleave after
a. Pancreas aspactic acid residues
b. Parotid gland 1. Initiator caspases
c. Kidney  Caspase-8
 Caspase-9
MORPHOLOGIC and BIOCHEMICAL CHANGES in APOPTOSIS 2. Executioner caspase
Morphology  Caspase-3
Cell Shrinkage  Caspase-6
 Cell is smaller in size
 Cytoplasm is dense DNA and Protein Breakdown
 Organelles though relatively normal, are more tightly packed  Apoptotic cells exhibit a characteristic breakdown of DNA into
Chromatin condensation large 50- to 300-kilobase pieces
 Most characteristic feature of apoptosis  Cleavage of DNA by Ca2+ and Mg2+ dependent endonucleases into
 Chromatin aggregates peripherally, under the nuclear membrane, fragments whose sizes are multiples of 180 to 200 base pairs,
into dense masses of various shapes and sizes reflecting cleavage between nucleosomal subunits
 Nucleus may break up  producing two or more fragments
Formation of cytoplasmic blebs and apoptotic bodies Membrane Alterations and Recognition by Phagocytes
 Extensive surface blebbing  Movement of some phospholipids (notably phosphatidylserine)
from the inner leaflet to the outer leaflet of the membrane,
 where they are recognized by a number of receptors on
phagocytes
 These lipids are also detectable by binding of a protein called
annexin V  commonly used staining to identify apoptotic cells
MECHANISM of APOPTOSIS
 The genes and proteins that control the process and the sequence
of events are conserved in all multicellular organism
1. INITIATION Phase – some caspases become catalytically active
2. EXECUTION Phase – other caspases trigger the degradation of
critical cellular components
 Initiation of apoptosis occurs principally by signals from two
distinct pathways:
1. Intrinsic (Mitochondrial) Pathway
 Involves the action of sensors and effectors of the Bcl-2
family, which induce leakage of mitochondrial proteins
 With some of the anti-apoptotic proteins (“regulators”) that
inhibit mitochondrial leakiness and cytochrome c-dependent
caspase activation in the mitochondrial pathway
2. Extrinsic (Death Receptor-Initiated) Pathway
 Engagement of death receptors leads directly to caspase
activation
 The two pathways of apoptosis differ in their induction and
regulation, and both culminate in the activation of “Executioner”
caspases
The Intrinsic (Mitochondrial) Pathway of Apoptosis
 Major mechanism of apoptosis in all mammalian cells, and its role
in a variety of physiologic and pathologic processes is well
established
 Result of increased mitochondrial permeability and release of pro-
apoptotic molecules (death inducers) into the cytoplasm
 Release of these mitochondrial proteins is controlled by a finely
orchestrated balance between pro- and anti-apoptotic members
of the Bcl family of proteins
 Growth factors and other survival signals stimulate production of
anti-apoptotic proteins, the main ones being Bcl-2, Bcl-x, band
Mcl-1  deprived of survival signals or their DNA is damaged, or
misfolded proteins induce ER stress  sensors of damage or
stress are activated
 Sensors: Bim, Bid, and Bad that contains a single “Bcl-2 homology
domain: and are called “BH3-only proteins”  activate two
critical (proapoptotic) effectors: Bax and Bak
 Cytochrome c  released into the cytosol  binds to protein
Apaf-1 (apoptosis-activating factor 1) forms a wheel-like
hexamer  APOPTOSOME  bind caspase-9 molecules  auto-
amplification process
 Mitochondrial proteins (Smac/DIABLO) enter the cytoplasm 
bind to and neutralize cytoplasmic proteins that function as
physiologic inhibitors of apoptosis (IAPs)  block the activation of
caspases (executioners like caspase-3)  keep cells alive
The Extrinsic (Death Receptor-Initiated) Pathway of Apoptosis
 Initiated by engagement of plasma membrane death receptors on
a variety of cells
 Death receptors – members of the TNF receptor family that
contain a cytoplasmic domain involved in protein-protein
interactions  DEATH DOMAIN  essential for delivering
apoptotic signals
Types of Death Receptors
1. Type 1 TNF receptor (TNFR 1)
2. Fas (CD95)
 The ligand for Fas is called Fas ligand (FasL)  expressed on:
a. T cells that recognize self-antigens and functions to eliminate
self-reactive lymphocytes
b. Cytotoxic T lymphocytes – kill virus infected and tumor cells
 FasL binds to Fas  3 or more molecules of Fas are brought
together  cytoplasmic death domains form a binding site for an
adapter protein that also contains a death domain  FADD (Fas-
associated death domain)
 FADD  attached to death receptors  binds an inactive form of
caspase-8 (caspase-10 in humans) via death domain  multiple
pro-caspase molecules brought into proximity  cleave one
another to generate active caspase-8  triggers a cascade of
caspase activation by cleaving  activating other pro-caspases 
active enzymes mediate the execution phase of apoptosis
 Can be inhibited by a protein called FLIP  binds to pro-caspase-8
but cannot cleave and activate the caspase because it lacks a
protease domain
 2. DNA Damage
a. Exposure of cells to radiation or chemotherapeutic agents 
induces apoptosis initiated by DNA damage (genotoxic
stress) which involves the tumor-suppressor gene p53
 p53 – accumulates in cells when DNA is damaged  arrest the
cell cycle (at the G1 phase)  allow time for repair
 Too great damage  p53 tirggers apoptosis
 p53 – serves as a critical “life or death” switch following genotoxic
stress

3. Protein misfolding
 Chaperones in the ER – control the proper folding of newly
synthesized protein
 Misfolded polypeptides are ubiquitinated and targeted for
proteolysis in proteasomes
 Unfolded Protein Response – unfolded or misfolded proteins that
accumulate in the ER
a. Activates signaling pathways that increase the production of
chaperones
b. Enhance proteosomal degradation of abnormal proteins
c. Slow protein translation  reducing the load of misfolded
proteins in the cell
Execution Phase of Apoptosis  ER Stress – if the cytoprotective response is unable to cope with
 The two initiating pathways converge to a cascade of caspase the accumulation of misfolded proteins  cell activates caspases
activation  mediates the final phase of apoptosis and induces apoptosis
 Mitochondrial pathway leads to activation of the initiator caspase-
9, and the death receptor pathway to the initiators caspase-8 and
-10
 Executioner caspases (caspase 3, and -6) – act on many cellular
components
a. Cleave an inhibitor of a cytoplasmic DNase  make the
DNase enzymatically active
b. Induces the characteristic cleavage of DNA into nucleosome-
sized pieces
c. Degrade structural components of the nuclear matrix 
promote fragmentation of nuclei

Removal of Dead Cells

 Cells that are dying by apoptosis secrete soluble factors that
recruit phagocytes
 Thrombospondin – an adhesive glycoprotein that is recognized by
phagocytes
 Macrophages – may produce proteins that bind to apoptotic cells
 target the dead cells for engulfment
 May also become coated with natural antibodies and proteins of
the complement system (C1q)  recognize by phagocytes
4. Apoptosis Induced by the TNF Receptor Family
CLINICO-PATHOLOGIC CORRELATION:  FasL on T cells binds to Fas on the same neighboring lymphocytes
APOPTOSIS IN HEALTH AND DISEASE  elimination of lymphocytes that recognize self-antigens
 Cytokine TNF – an important mediator of the inflammatory
Examples of Apoptosis reaction  capable also of inducing apoptosis  induces
1. Growth Factor Deprivation thrombosis of tumor blood vessels  ischemic death of the tumor
a. Hormone – sensitive cells deprived of the relevant hormone
b. Lymphocytes that are not stimulated by antigens and 5. Cytotoxic T Lymphocyte – Mediated Apoptosis
cytokines  Cytotoxic T Lymphocyte (CTLs) – recognize foreign antigens
c. Neurons deprived of nerve growth factor presented on the surface of infected host cells  secrete
 Triggered by intrinsic (mitochondrial) pathways perforin, a transmembrane pore-forming molecule  promotes
 Due to decreases synthesis of the Bcl-2 and Bcl-x and activation of entry of the CTL granule serine proteases called granzyme
Bim and other pro-apoptotic members of the Bcl family  Granzymes –has the ability to cleave proteins at aspartate
residues  activate a variety of cellular caspases
6. Disorders Associated with Dysregulated Apoptosis  Diseases caused by genetic defects in enzymes involved in
 Dysregulated Apoptosis (“too little or too much”) the metabolism of lipid and carbohydrates  deposition into
1. Disorders associated with defective apoptosis and increased cell lysosomes
survival 4. An abnormal exogenous substance is deposited and accumulates
 Cancer – cells not only fail to die but are susceptible to the because the cell has neither the enzymatic machinery to degrade
accumulation of mutations because of defective DNA repair the substance nor the ability to transport it to other sites
 Autoimmune disorder – failure to eliminate dead cells   Accumulation of carbon particles and nonmetabolizable
potential source of self-antigens chemicals such as silica
2. Disorders associated with increase apoptosis and excessive cell LIPIDS
death  All major classes of lipids can accumulate in cells:
a. Neurodegenerative diseases – loss of specific sets of neurons a. Triglycerides
b. Ischemic injury – myocardial infarction and stroke b. Cholesterol / cholesterol esters
c. Death of virus-infected cells – viral infections c. Phospholipids – components of myelin figures found in
necrotic cells
AUTOPHAGY
 The process in which a cell eats its own contents Steatosis (Fatty Change)
 Survival mechanism in times of nutrient deprivation  Abnormal accumulations of triglycerides within parenchymal cells
 Intracellular organelles and portions of cytosol are first  Often seen in the liver because it is the major organ involved in fat
sequestered from the cytoplasm in an autophagic vacuole  metabolism
fuses with lysosomes  autophagolysosome  cellular  May also occurs in heart, muscle, and kidney
components are digested by lysosomal enzymes
 Regulated by a defined set of “autophagy genes” (Atgs) Causes
a. Toxins
b. Protein malnutrition
c. Diabetes mellitus
d. Obesity
e. Anoxia
f. Alcohol abuse in developed nations, assoc.
g. Nonalcoholic fatty liver disease with DM and obesity

INTRACELLULAR ACCUMULATIONS  Free fatty acids from adipose tissue or ingested food 
1. Normal cellular constituent – water, lipids, proteins, and transported into hepatocytes  esterified to triglycerides 
carbohydrates  accumulates in excess coverted into cholesterol or phospholipids or oxidized to ketone
2. Abnormal substance bodies and some are synthesized from acetate as well
a. Exogenous – mineral or products of infectious agents  Release of triglycerides from the hepatocytes requires association
b. Endogenous – product of abnormal synthesis or metabolism with apoproteins to form lipoproteins  transported from the
blood into the tissues
Four Types of Abnormalities in Intracellular Accumulations  Excess accumulation of triglycerides within the liver may result
1. Normal endogenous substance is from excessive entry or defective metabolism and export of lipids
produced at a normal or increased  Alcohol – hapatotoxin, alters mitochondrial and microsomal
rate, but the rate of metabolism is functions  increased synthesis and reduced breakdown of lipids
inadequate to remove it  CCl4 and protein malnutrition – cause fatty change by reducing
 fatty change in liver synthesis of apoproteins
 reabsorption proteins droplets in  Hypoxia – inhibits fatty acid oxidation
the tubules of the kidneys  Starvation – increases fatty acid mobilization from peripheral
2. An abnormal endogenous stores
substance, typically the product of
a mutated gene, accumulates
because of defects in protein
folding and transport and an
inability to degrade the abnormal
protein efficiently
 Accumulation of mutated α-1 anti-
trypsin in the liver cells
 Mutated proteins in degenerative
disorders of the CNS
3. A normal endogenous substance
accumulates because of defects,
usually inherited, in enzymes that
are required for the metabolism of
the substance
Morphology
 Fatty change – often seen in the liver and heart
 Appears as clear vacuoles within parenchymal cells
 Intracellular accumulations of water or polysaccharides may also
produce clear vacuoles
a. Liver
 Mild fatty change may not affect the gross appearance
 Progressive accumulation, organ enlarges  becomes
increasingly yellow  may weigh two to four times normal
 bright yellow, soft, greasy organ
 Fatty change begins with the development of minute,
membrane-bound inclusions (liposome) closely applied to
the ER  small vacuoles in the cytoplasm around the nucleus
 vacuoles coalesce  cleared spaces that displace the
nucleus to the periphery of the cell  FATTY CYSTS
b. Heart
 Lipid in the form of small droplets
1. Prolonged moderate hypoxia  profound anemia 
intracellular deposits of fat  bands of yellowed
myocardium alternating with bands of darker, red-
brown, uninvolved myocardium (TIGERED EFFECT)
2. Produced by profound hypoxia or by some forms of
myocarditis  uniformly affected myocytes
Cholesterol and Cholesterol Esters
1. Atherosclerosis – smooth muscle cells and macrophages within
the intimal layer of the aorta and large arteries are filled with lipid
vacuoles
 Have a foamy appearance (foam cells)
 Yellow cholesterol – laden atheromas in the intima
 May rupture  releasing lipids into the extracellular space
2. Xanthomas – intracellular accumulation of cholesterol within
MACROPHAGES
 Clusters of foamy cells that are found in the subepithelial
connective tissue of the skin and in tendons producing
timorous masses
3. Cholesterolosis – focal accumulations of cholesterol – laden
macrophages in the lamina propria of the gallbladder
4. Niemann-Pick disease, Type C – lysosomal storage disease caused
by mutations affecting an enzyme involved in cholesterol
trafficking  cholesterol accumulation in multiple organs
PROTEINS
 Intracellular accumulations appear as rounded, eosinophilic
droplets, vacuoles, or aggregates in the cytoplasm
 Can be amorphous, fibrillar, or crystalline in appearance in
electron microscope
 Amyloidosis – abnormal proteins deposit in extracellular spaces
1. Reabsorption droplets in proximal renal tubules – seen in renal
diseases associated with protein loss in the urine (proteinuria)
 Proteins appear as pink hyaline droplets within the cytoplasm of
the tubular cell
 Reversible process; proteinuria diminishes  protein droplets are
metabolized and disappear
2. The proteins that accumulate may be normal secreted proteins
that are produced in excessive amounts, as occurs in certain
plasma cells engaged in active synthesis of immunoglobulins
 ER becomes hugely distended  large, homogenous
eosinophilic inclusions  RUSSELL BODIES
3. Defective intracellular transport and secretion of critical proteins
 α1-antitrypsin deficiency – mutations in the protein
significantly slow folding  build up of partially folded
intermediates  aggregate in the ER of the liver and not
secreted  deficiency of the circulating enzyme 
EMPHYSEMA
4. Accumulation of cytoskeletal proteins
a. Microtubules – 20 – 25 nm in diameter
b. Thin actin filaments – 6 – 8 nm
c. Thick myosin filaments – 25 nm
d. Intermediate filaments – 10 nm
 Provide a flexible intracellular scaffold that organizes the
cytoplasm and resists forces applied to the cell
1. Keratin filaments (epithelial cells) associated with
2. Neurofilaments (neurons) cell injury
3. Desmin filaments (muscle cells)
4. Vimentin filaments (connective tissue cells)
5. Glial filaments (astrocytes)
 Alcoholic hyaline – eosinophilic cytoplasmic inclusion in liver
cells that is characteristic of alcohol liver disease, composed
of predominantly keratin intermediate filaments
 Neurofibrillary tangle – found in the brain of Alzheimer
disease contains neurofilaments and other proteins
5. Aggregation of abnormal proteins – may deposit in tissues and
interfere with normal functions
 Deposits can be intracellular, extracellular, or both
 Aggregates may be either directly or indirectly cause the
pathologic changes
 Amyloidosis – a.k.a proteinopathies or protein-aggregation
diseases
HYALINE CHANGE
 Alteration within cells or in the extracellular space that gives a
homogenous, glassy, pink appearance in routine histologic
sections stained with hematoxylin and eosin
 Produced by a variety of alterations and does not represent a
specific pattern of accumulation
1. Extracellular Hyaline - collagenous fibrous tissue in old scars may
appear hyalinized
 HPN and DM  walls of arterioles (kidneys) become
hyalinized  from extravasated plasma protein and
deposition of basement membrane material
GLYCOGEN
 Readily available energy source stored in the cytoplasm of healthy
cells
 Excessive intracellular deposits of glycogen are seen in patients
with an abnormality in either glucose or glycogen metabolism
 Appear as clear vacuoles within the cytoplasm
 Dissolves in aqueous fixatives
 Diabetes mellitus – prime example of glucose metabolism
disorder
 Glycogen found in renal tubular epithelial cells, β cells of the
islets of Langerhans, and heart muscle cells
 Glycogen storage diseases or glycogenoses – genetic disorders of
glycogen accumulations within the cells
PIGMENTS
 Colored substances
A. Exogenous Pigments
 Coming from outside the body
1. Carbon (coal dust) – most common exogenous pigment, an
ubiquitous pollutant of urban life
 Inhaled  picked up by macrophages within the alveoli 
transported through lymphatic channels to the regional
lymph nodes in the tracheobronchial region  accumulation
 blacken the tissues of the lungs (anthracosis)
 Coal Worker’s Pneumoconiosis – serious lung disease of coal
miners where aggregates of carbon dust may induce
fibroblastic reaction or even emphysema
 Tattooing – form of localized, exogenous pigmentation of the
skin. Pigments inoculated are phagocytosed by dermal
macrophages  reside for the remainder of the life of the
embellished
B. Endogenous Pigments
 Synthesize within the body itself
 Lipofuscin – insoluble pigment
 Also known as lipochrome or wear-and-teat pigment
 Composed of polymers of lipids and phospholipids in complex
with protein  derived through lipid peroxidation of
polyunsaturated lipids of subcellular membranes
 Not injurious to the cell or its functions
 A telltale sign of free radical injury and lipid peroxidation
 Latin (fuscus – brown) brown lipid
 Appears as a yellow – brown, finely granular cytoplasmic, often
perinuclear pigment
 Melanin – derived from the Greek (melas – black)
 An endogenous, non-hemoglobin derived, brown-black
pigment formed when the enzyme tyrosinase catalyzes the
oxidation of tyrosine to dihydroxyphenylalanine in melanocytes
 Only endogenous brown-black pigment
 Homogentisic acid – a black pigment that occurs with
alkaptonuria  pigment is deposited in the skin, connective
tissue, and cartilage  OCHRONOSIS
 Hemosiderin – hemoglobin – derived, golden yellow-to-brown,
granular or crystalline pigment that serves as one of the major
storage forms of iron
 Transferrins – transport proteins that carries iron
 Apoferritin – storage form in the cells with proteins
 Ferritin – forms hemosiderin granules
 Seen in the mononuclear phagocytes of the bone marrow,
spleen, and liver  engaged in red cell breakdown
 Local or systemic excesses of iron cause hemosiderin to
accumulate within cells
 BRUISE: Extravasated red cells at the site of injury are
phagoctosed over several days by macrophages  break dwon
hemoglobin and recover the iron
 Removal of the iron  heme moiety is converted to biliverdin
(“green bile”)  bilirubin (“red bile”)
 Iron released from heme  incorporated to ferritin 
hemosiderin
 Red-blue  green-blue  golden yellow  resolved
 Systemic overload of iron hemosiderin  may be deposited in
many organs and tissues  HEMOSIDEROSIS
Causes of Hemosiderosis
1. Increased absorption of dietary iron
2. Hemolytic anemias  abnormal quantities of iron are released
from erythrocytes
3. Repeated blood transfusion because the transfused red cells
constitute an exogenous load of iron
Morphology
 Iron pigment appears as a coarse, golden, granular pigment lying
within the cell’s cytoplasm
 Localized breakdown of red cells: hemosiderin is found initially in
the phagocytes in the area
 Systemic hemosiderosis: found at first in the mononuclear
phagocytes of the liver, bone marrow, spleen, and lymph nodes
and in scattered macrophages throughout other organs such as
skin, pancreas, and kidneys
 Hemochromatosis – an inherited disease seen with more extreme
accumulation of iron and is associated with liver, heart, and
pancreatic damage  liver fibrosis, heart failure, and diabetes
mellitus
 Bilirubin – normal major pigment found in bile
 Derived from hemoglobin but contains no iron
 Jaundice is a common clinical disorder caused by excesses of
this pigment within cells and tissues
Pathologic Calcification
 Abnormal tissue deposition of calcium salts, together with smaller
amounts of iron, magnesium, and other mineral salts
a. Dystrophic Calcification – deposition occurs locally in dying
tissues
 Occurs despite normal serum levels of calcium and in
the absence of derangements in calcium metabolism
b. Metastatic calcification – the deposition of calcium salts in
otherwise normal tissues
 Almost always results from hypercalcemia secondary to
some disturbance in calcium metabolism
DYSTROPHIC CALCIFICATION
 Encountered in areas of necrosis, whether they are of coagulative,
caseous, or liquefactive type, and in foci of enzymatic necrosis of
fat
 Almost always present in the atheromas of advanced
atherosclerosis
 Commonly develops in aging or damaged heart valves 
hampering their function
 Appears as fine, white granules or clumps, often felt as gritty
deposits
 Dystrophic calcification of the aortic valve
Morphology
 Calcium salts have a basophilic, amorphous granular, sometimes
clumped appearance
 Can be intracellular, extracellular, or in both locations
 Heterotopic bone may be formed in the focus of calcification
 Single necrotic cells may constitute seed crystals that become
encrusted by the mineral deposits
 Psammoma bodies – lamellated configurations due to progressive
acquisition of outer layers  e.g. Papillary cancers
Pathogenesis
 Final common pathway: formation of crystalline calcium
phosphate mineral in the form of an apatite similar to the
hydroxyapatite of bone
 Calcium is concentrated in membrane-bound vesicles in cells by a
process that is initiated by membrane damage and:
1. Calcium ion binds to the phospholipids present in the vesicle
membrane
2. Phosphatases associated with the membrane generate phosphate
groups  bind to the calcium
3. The cycle of calcium and phosphate binding is repeated  raising
the local concentrations and producing a deposit near the
membrane
4. Structural change occurs in the arrangement of calcium and
phosphate groups  generating a microcrystal  propagate and
lead to more calcium deposition
METASTATIC CALCIFICATION
 May occur in normal tissues whenever there is hypercalcemia
Four Principal Causes of Hypercalcemia
1. Increased secretion of Parathyroid hormone (PTH) with
subsequent bone resorption
a. Hyperparathyroidism due to parathyroid tumors
b. Ectopic secretion of PTH – related protein by malignant
tumors
2. Destruction of bone tissue, secondary to:
a. Primary tumors of bone marrow (multiple myeloma,
leukemia)
b. Diffuse skeletal metastasis (breast cancer)
c. Accelerated bone turnover (Paget disease)
d. Immobilization
3. Vitamin – D related disorders (abnormal sensitivity to Vitamin D
a. Vitamin D intoxication
b. Sarcoidosis – macrophages activate a vitamin D precursors
c. Idiopathic hypercalcemia of infancy (William’s syndrome)
4. Renal failure  retention of phosphate  secondary
hyperparathyroidism
a. Aluminum intoxication – patients on chronic renal dialysis
b. Milk-alkali syndrome – due to excessive ingestion of calcium
and absorbable antacids such as milk or calcium carbonate
 May occur widely throughout the body but principally affects the
interstitial tissues of the gastric mucosa, kidneys, lungs, systemic
arteries, and pulmonary veins
 Nephrocalcinosis – massive deposits of mineral salts in the kidney
CELLULAR AGING
 Result of a progressive decline in cellular function and viability
caused by genetic abnormalities and the accumulation of cellular
and molecular damage due to the effects of exposure to
exogenous influences
Changes that Contribute to Cellular Aging
1. Decreased cellular replication – “SENESCENCE”
 with each cell division there is incomplete replication of
chromosome ends (telomere shortening)  cell cycle arresr
 Telomeres – short repeated sequences of DNA (TTAGGG)
present at the linear ends of chromosomes that are
important for ensuring the complete replication of
chromosomal ends and for protecting chromosomal termini
from fusion and degradation
 Telomerase – enzyme that maintained telomere length; a
specialized RNA-protein complex that uses its own RNA as a
template for adding nucleotides to the ends of chromosomes
 Werner Syndrome – a rare disease characterized by
symptom of premature aging, are defective in DNA
replication and have a markedly reduced capacity to divide
 Replicative senescence – can also be induced by increased
expression of the cell cycle inhibitor p16INK4a and by DNA
damage
2. Accumulation of metabolic and genetic damage
 Cellular life span is determined by a balance between damage
resulting from metabolic events occurring within the cell and
counteracting molecular responses that can impair the damage
 Reactive Oxygen Species – by-products of oxidative
phosphorylation cause covalent modifications of proteins, lipids,
and nucleic acids
1. Variation in longevity among different species is inversely
correlated with the rates of mitochondrial generation of O2- anion
radical
2. Overexpression of the antioxidative enzymes SOD and catalase
extends life span in transgenic forms of Drosophilia
 Ataxia-telangiectasia – patients display some of the
manifestations of aging at an increased rate  mutated gene
encodes a protein involved in repairing double-strand breaks in
DNA

 Calorie restrictions – most effective way of prolonging life span

 May be mediated by a family of proteins called SIRTUINS
 SIRTUINS – have histone deacetylase activity; promote the
expression of several genes whose products increase
longevity
a. Increase metabolic activity
b. Reduce apoptosis
c. Stimulate protein folding
d. Inhibit the harmful effects of oxygen free radicals
e. Increase insulin sensitivity and glucose metabolism

 Growth factors (Insulin-like growth factor) and intracellular

signaling pathways triggered by these hormones also
influence life span