Cell damage-2

Second lecture
DR-Rami AL-Zagha
Medical/ Dental/Pharmacy students
2011-2012
 Mechanisms of cell injury
What are the general principle?
• The cellular responses to injurious stimuli
depend on the type, duration, and severity of
injury.
• Consequences of an injurious stimulus depend on
the type, status, adaptability and genetic make up
of the cell.
 Mechanisms of cell injury
• The main targets for the injury are:
– Cell membrane, ionic and osmolarity
homeostasis.
– Mitochondrial aerobic respiration (ATP)
– Protein synthesis
 Genetic apparatus (DNA)
 Mechanisms of cell injury
• Regardless of the target that was initially
attacked, the different components are
integrated in a way, that secondary effects
will occur, and other components will be
affected.
 Mechanisms of cell injury
• Chronologically cell injury will follow
– Loss of function (cell is viable), reversible
– Cell death, irreversible.
• Biochemical changes
• Ultrastructural changes
• Light microscopic changes
• Gross “macroscopic” changes
 General biochemical
mechanisms:
1. Mitochondrial damage and ATP depletion :
Mitochondrial damage either directly or
indirectly will lead to loss of ATP
production.
This will lead to loss of the function of cell
organelles which are all dependent on the
supply of the ATP
 Mitochondrial damage.

• Affection of the mitochondrial membrane by Ca,

phospholipases, oxidative stress.
• This will lead to the formation of High-
Conductive –Channels in the inner mitochondrial
membrane. (mitochondrial permeability
transition).
• Loss of the proton gradient across the
mitochondrial membrane, and escape of
cytochrome-C, initiating apoptosis.
Mitochondrial damage
 General biochemical
mechanisms:
2. Oxygen changes:
Whether through oxygen deprivation
(hypoxia/ischemia),
OR
through the formation of free radicals.
 Oxygen changes:
a.Hypoxic/ischemic injury:
– Hypoxia/ischemia will lead to decrease in the
oxygen supply. The first system to be affected
is the oxidative respiration by the mitochondria.
– This will lead to decrease in the ATP
a. Hypoxic/ ischemic injury
a. Oxygen changes:hypoxic/ischemic
injury
• Loss of ATP dependent Na/K pump
• Increase intracellular Na
• Decrease intracellular K
• Increase intracellular osmolarity
• Acute cellular swelling
• Increase anaerobic glycolysis:
• Decrease glycogen
• Accumulation of lactic acid
• Lower pH
• Detachment of ribosomes, with loss of protein
synthesis
 a. Oxygen changes:hypoxic/
ischemic injury
• All of these changes are reversible if the
stimulus is removed.
• The changes will become irreversible if the
stimulus continues.
 b. Ischemia/reperfusion injury
• After restoration of the blood supply, a
paradoxical tissue loss occurs, with more cells
dying.
• This is explained on the basis of more influx of Ca
into the partially damaged cells, inflammatory
cells brought into the site of injury by blood, and
production of free radicals by the partially
damaged mitochondria.
 Oxygen changes
c. Free radical-induced cell injury:
Free radicals:
• Definition: unstable chemical species with a
single unpaired electron in the outer orbital
• Either decay spontaneously or react with
any other molecule to transfer the electron
and change into a more stable form.
 Sources of free radicals:
• produced normally in the cells:
– in normal respiration: Redox reaction
– Cell membrane e.g. NADPH oxidase
– Cytosol: Fenton reaction, xanthine oxidase.
– Peroxisomes: oxidase.
– ER: P450 oxidase.
– Inflammatory cells, in the lysosomes, myeloperoxidase,
NO synthetase.
• absorption of radiation, hydrolysis of water.
• chemicals, CCl4, or drugs, Paracetamol, or O2
toxicity.
 Targets for the free radicals

• lipid peroxidation of membranes, double bonds in

the membrane polyunsaturated lipids are the
targets.
• DNA fragmentation, due to reaction with thymine.
• Cross linking of proteins and fragmentation, due
to sulfhydryl-mediated protein cross-linking.
 Protection mechanisms
• Spontaneous decay.
• Superoxide dismutase (SOD), in the
cytoplasm and mitochondria
– Catalyses the reaction 2O2+2H SOD H2O2+O2
– It also enhances spontaneous decay
 Protection mechanisms:
• Glutathione peroxidase, in the cytoplasm
and mitochondria
– 2OH +2GSH 2H2O+GSSG (glutathione
homodimer)
– GSH: GSSG is a reflection of the oxidative
status of the cell.
 Protection mechanisms:
• Catalase, in the peroxisomes
– 2H2O2 Catalase O2+2H2O
• Transport proteins, Ferritin, ceruloplasmin.
• Antioxidants: vitamin E/C/A
– They either block the formation, or savage the
ones that are formed
Summary of free radicals
 Cell injury
3.defects in plasma membrane permeability:
• Whether direct from the injurious stimulus
or indirect, like 2ry effect after ATP
depletion, or influx of calcium into cells.
• This will lead to break of the concentration
gradient of the metabolites between the
intra and extracellular environment.
 Cell injury
4. loss of calcium homeostasis:
• Free intracellular Ca concentration is 10000
less than extracellular concentration; this is
achieved through an ATP-dependent pump
with sequestration of calcium within the
mitochondria and ER, and by binding to Ca-
buffering proteins.
 Cell injury
• With injury, influx of Ca into the cell, increase Ca
concentration and activation of enzymes.
• Calcium is very important, since it acts as
secondary messenger for the activation of different
enzymes within the cell cytoplasm, like protein
kinase, leading to loss of the integration of the
cell.
• Calcium is NOT essential in causing cell death
Calcium homeostasis
 Cell injury
5.Chemical injury:
Two mechanisms:
• Direct combination with a critical molecule
of cell component.
• Activation of an inactive chemical.
Direct combination with a critical
molecule of cell component
• The cells that are affected are the ones that use,
absorb, excrete, or concentrate the compound.
• HgCl2: Hg binds directly to sulfhydryl group of
cell membrane and other membranes. This will
affect the ATPase transport channels, leading to
increased permeability and death of the cells.
• Chemotherapy and antibiotics.
 Activation of an inactive
chemical
• through the P-450 in the SER in the liver.
This is followed by changing into free
radicals
• CCl4 P-450/liver CCl3 free radical. This
leads to membrane peroxidation, decrease
protein synthesis, accumulation of fat, with
fatty liver resulting.
 Types of cell injury
Reversible cell injury/sublethal cell damage
• Morphology:
Remember the sequence of events; loss of
function, ultrastructural, microscopic,
macroscopic changes
 Reversible cell injury
Ultrastructural features:
• plasma membrane alternation:
– blebs, loss of microvilli, loosing of intracellular
attachment
• mitochondrial alternation
– earliest manifestation of sublethal injury
– swelling, appearance of phospholipid rich
amorphous densities
 Reversible cell injury
• dilation of endoplasmic reticulum
– loss of the ribosomes, and dissociation of
polysomes
• nuclear alternation
 Reversible cell changes
Light microscopic changes: Fig 3.7,
pathology, page-28.
• cell swelling (hydropic changes or vacuolar
degeneration):
– universal to all cell types
– loss of ionic and fluid homoeostasis
– distended and punched out segments of ER
– cells show clear vacuoles in the cytoplasm
 Reversible cell changes
• Fatty changes:
– Specific to cells dealing with fat metabolism,
liver and heart
– Lipid vacuoles in the cytoplasm
 Reversible cell changes
Macroscopic changes; gross features:
– increase in the weight of the organ
– pallor of the organ
 Types of cell injury
Irreversible cell injury: “lethal damage”
• General pathways:
– excessive damage to all membranes, cytosolic
and organelles
– Calcium is a potential mediator of irreversible
cell death. Increase intracellular Ca content
from the intracellular compartments, with
activation of the different enzymes
 Irreversible cell injury
- One of the earliest manifestations of irreversible cell
injury is the vacuolization of the mitochondria, and
the accumulation of amorphous, calcium-rich
densities in mitochondrial matrix
- Affection of the oxidative Phosphorylation, decrease
ATP.
- Leakage of proteins and cellular constituents. This
forms the basis for blood testing.
 Irreversible cell injury
– Leakage of digestive enzymes from lysosomes.
With the low pH, hydrolases are activated.
They begin to digest the cell components.
• Autolysis: if the cells are digested by their own
enzymes
• Heterolysis: if the cells are digested by lysosomes
from other cells
 Irreversible cell injury
– Nuclear changes:
• pyknosis, karyolysis, karyorrhexis

Dead cells are eventually replaced by “myelin

figures”. These are either phagocytosed or
degraded into fatty acids, with deposition of
calcium salts on top.
Mechanisms of irreversible injury
2 phenomena characterize irreversibility
• Inability to reverse mitochondrial
dysfunction
– Development of profound disturbances in the
membrane
– This is the VITAL step at which “No Return”
issues
 Irreversible cell injury
• Cell membrane damage:
• Progressive loss of membrane phospholipids. This caused by
activation of phospholipases by Ca, and decrease synthesis of
proteins
• Cytoskeletal abnormalities. Loss of the anchoring filaments
between the cytoplasm and the cell membrane. So the cell
membrane becomes loose and susceptible to rupture.
• Toxic oxygen radicals. This leads to peroxidation of the
membranes.
• Lipid breakdown products. These act as detergents on the
membranes.
 Morphology of irreversible cell
injury:
Necrosis:
• The morphological correlate of irreversible
cell injury
• Defined as the sequence of morphological
changes that follows cell death in living
tissue
• These morphological changes require hours
to develop. They can’t be detected instantly
 Morphology of irreversible cell
injury:
• This morphological appearance is the
sequence of 2 concurrent processes
– enzymatic digestion of proteins
– denaturation of proteins
 Ultrastructural morphological
changes:
• Defects in cell membrane
• Mitochondrial swelling and large densities
• Swelling of the ER and detachment of
ribosomes
• Appearance of myelin figures
• Rupture of lysosomes
 Light microscopic features

• changes in the cytoplasm

– acidophilia of the cytoplasm (pink staining)
• binding of eosins to denatured proteins
• loss of basophilia from the loss of RNA
– glassy cytoplasm: loss of glycogen
– vacuolated cytoplasm: degeneration of organelles
– calcification of cells
 Light microscopic features
• changes in the nucleus :Fig 3-9, cellular events in
necrosis
– Pyknosis: increased basophilia due to shrinkage of the
nucleus
– Karyorrhexis: fragmentation of the pyknotic nucleus by
nucleases
– Karyolysis: loss of the basophilia of the chromatin, 2ry
to DNAase activity.
1-2 days following death, the nucleus disappears
completely
 Light microscopic features:
fig 3-10
• Coagulative necrosis:
– Defined as death of cells with preservation of
the basic structural outlines of the coagulated
cells, with preservation of the general tissue
architecture
 Coagulative necrosis
– The most common type of necrosis
– Protein denaturation overcomes enzymatic
digestion
– Seen in most organs after hypoxia/ischemia
except brain
– Gangrenous necrosis: ischemic Coagulative
necrosis of limb
 Light microscopic changes
• Liquefactive necrosis:
– Defined as necrosis with complete digestion of
the cells with obliteration of the normal
architecture
– Enzymatic digestion overcomes the
denaturation
 Liquefactive necrosis
– 2 situations
• hypoxic/ischemic injury of the brain
• bacterial/fungal infection, with accumulation of
WBCs and release of enzymes
– Wet gangrenous necrosis: ischemic Coagulative
necrosis with superimposed bacterial infection
 Light microscopic changes
• Caseous necrosis:
– Special type of necrosis, seen with tuberculus
infection
– Grossly: cheesy white appearance to the
necrotic focus
– Microscopically: the necrotic focus is
composed of structureless amorphous granular
debris
 Light microscopic changes
• Fatty necrosis
– Special type of necrosis with focal areas of fat
destruction
– Seen with acute pancreatitis, due to release of
enzymes from the injured pancreas
– Grossly: fat saponification: visible whitw
chalky areas
– Microscopically: shadows of cells with
calcification