Cell Injury, Adaptation, and Cell Death PDF

CELL INJURY, ADAPTATION, and CELL DEATH DR. AARON HAIG PATHOLOGY AND LABORATORY MEDICINE, LHSC, UNIVERSITY CAMPUS OBJECTIVES: At the end of this lesson, the student should be able to understand the various ways in which the cell can respond to damaging stimuli and more specifically:...

CELL INJURY, ADAPTATION, and CELL DEATH DR. AARON HAIG PATHOLOGY AND LABORATORY MEDICINE, LHSC, UNIVERSITY CAMPUS OBJECTIVES: At the end of this lesson, the student should be able to understand the various ways in which the cell can respond to damaging stimuli and more specifically: o Be able to list several causes or agents of cell injury. o Define and give appropriate physiological and pathological examples of cellular adaptation, i.e., atrophy, hypertrophy, hyperplasia, and metaplasia. o Define and contrast reversible versus irreversible cell injury. Be able to describe the cellular changes that characterize reversible cell injury (e.g., cellular swelling/hydropic degeneration; fatty change). Be able to describe the cellular changes that take place in irreversible cell injury leading to necrosis (e.g., within the nucleus - pyknosis, karyorrhexis and karyolysis; within the cytoplasm - eosinophilia). o Describe coagulative, liquefactive, fat, caseous, and gangrenous necrosis and be able to give an example of each. o Define and contrast apoptosis and necrosis. Be able to describe the morphological changes that characterize each. o Describe the mechanism of cell injury and/or death in response to decreased oxygen. o Provide examples of physiological (e.g., lipofuscin, melanin) and pathological intracellular accumulations (e.g., fatty change). Recommended readings: "Robbins Basic Pathology" 11th Edition, 2023. Chapter 1 (pg. 1-24). See Introduction and Overview of Cellular Responses; Cellular Adaptations to Stress; Overview of Cell Injury and Cell Death; Reversible Injury; Irreversible Cell Injury and Necrosis; Apoptosis; Mechanisms of Cell Injury; Ischemic and Hypoxic Injury; Intracellular Accumulations (Fatty Change). Clinical and experimental pathology today are still grounded in the beliefs of 19 th century German scientist and statesman Rudolf Virchow that disease arises, not in organs or tissues in general, but primarily in individual cells. Cells must react and adapt to changing internal and external environments in order to survive. When the environmental changes exceed the capacity of the cell to maintain homeostasis, we recognize cell injury. A number of agents can cause injury or present a damaging stimulus to the cell, including: physical agents (trauma, radiation, extremes of temperature, changes in pressure) chemical agents (air pollutants, CO, pesticides, poisons, toxins, drugs) biological agents (microorganisms such as viral or bacterial infections; biological toxins) nutritional or metabolic alterations (nutrient deficiencies or excesses, ischemia, or lack of adequate blood supply; hypoxia or oxygen deficiency) immune reactions (allergens, autoimmune disease) genetic defects (single amino acid substitutions such as HbS (hemoglobin) in sickle cell disease; chromosomal abnormalities such as trisomy 21 or Down syndrome) cellular aging (loss of intrinsic repair mechanisms; repeated healing & repair following external injuries). PATHOLOGY 3500/PATHOLOGY 9535 1 CELL INJURY, ADAPTATION, AND CELL DEATH Reaction of the cell to a stress or damaging stimulus can range from a mild, completely reversible response to a long-term adaptive change in cell growth or to irreversible damage and cell death. The moment when reversible injury becomes irreversible injury, or the ‘point of no return’, cannot be well defined. The cell’s response depends to a large extent on the severity of the stimulus, the time frame over which it is exposed to the stressor (i.e., whether acute or chronic), and the individual cell type and its characteristics (rate of division; presence of protective mechanisms (e.g., free radical scavenging systems); its nutritional or metabolic state; its blood supply). Four cell components are particularly vulnerable. These are cell membranes critical for ionic and osmotic homeostasis; mitochondria and the generation of energy via ATP; protein synthesis; and cellular DNA. CELLULAR ADAPTATION Long term or chronic stimuli result in different responses in the cells. The response to persistent sublethal injury (chemical or physical) reflects an adaptation of the cell to the environment achieving a new steady state and preserving cell viability. Adaptive responses may include regulation of cell receptors (up- or down-regulation) or changes in protein synthesis and turnover. These adaptations include changes in cell size (atrophy or hypertrophy), number (hyperplasia) or organization (metaplasia; dysplasia). Atrophy: Reduced demand leads to atrophy of organs (if you don’t use it, you lose it!). Atrophy is defined as a decrease in mass due to the shrinkage in cell size. Due to diminished blood supply (ischemia) or diminished nutritional or trophic factors, a new steady state is reached in which a smaller cell is able to survive. The degradation of cellular proteins plays a key role in atrophy and two systems are involved: o lysosomes - intracellular organelles that contain powerful digestive enzymes that degrade both molecules from outside the cell (brought into the cell by endocytosis) or inside the cell (autophagy of subcellular components). o ubiquitin-proteasome pathway - cytosolic and nuclear proteins are targeted for degradation by conjugation to a 76 amino acid protein, ubiquitin, and degraded within a large cytoplasmic proteolytic complex - the proteasome. Atrophy can be physiological or pathological. For example, physiological atrophy occurs in normal aging (e.g., shrinkage and loss of brain cells with age). An example of pathological atrophy is disuse atrophy of skeletal muscle in immobilized limbs or denervation atrophy following loss of nerve input to a muscle. Hypertrophy: increase in the size of existing cells, due to the increase in synthesis of cellular protein and structural components and organelles responsible for producing them. Can be physiological or pathological in response to increasing functional demand or specific hormonal stimulation. Hyperplasia: increase in number of cells caused by cell division. Hypertrophy and hyperplasia can occur as a normal response to stimuli and often occur together. For example, hyperplasia and hypertrophy of uterine smooth muscle occur in pregnancy in response to estrogen; hypertrophy of skeletal muscles also occurs as a normal physiological response in weight training. Compensatory hyperplasia occurs when a portion of tissue is removed (e.g., after partial liver resection mitotic activity in remaining cells begins within 12 hours and eventually restores liver to normal weight). However, they may also be pathological responses. Hypertrophy of cardiac muscle fibres occurs in response to increased PATHOLOGY 3500/PATHOLOGY 9535 2 CELL INJURY, ADAPTATION, AND CELL DEATH  workload as a result of systemic hypertension (cardiomegaly). Excessive stimulation of the normal uterus by estrogen may result in endometrial hyperplasia. Metaplasia: If the long-term environment becomes unsuitable for certain types of cells, they may change into a different cell type, a process called metaplasia. For example, cells in the bronchi of smokers change from ciliated columnar cells to squamous cells. Importantly in all cases of growth adaptation, if the stimulus is removed, the tissue reverts to the resting state. This should be contrasted with the autonomous growth in neoplasia (you will hear more about this later in the section on Neoplasia). REVERSIBLE AND IRREVERSIBLE CELL INJURY Cells lose functional activity relatively quickly as a result of biochemical derangements, while the morphological changes of cell injury and death lag far behind. For example, heart cells lose the ability to contract after 1 to 2 minutes of ischemia, but do not die until 20 to 30 minutes of prolonged ischemia. Changes in cell ultrastructure (seen by light or electron microscopy) may not be apparent until several hours later following ischemic injury. At early stages or in mild forms of injury the functional and morphological changes are entirely reversible if the stimulus is removed. At this stage the injury has not progressed to severe membrane or nuclear damage. With continuing damage, cell injury becomes irreversible; the cell cannot recover and dies. There are two types of cell death – necrosis and apoptosis – which differ in their morphology, mechanisms, and cause (whether pathologic or physiologic). REVERSIBLE CELL INJURY Injury caused by a variety of agents (e.g., chemical or biological toxins, viral or bacterial infections, ischemia or hypoxia, excessive heat or cold) produces a characteristic cellular or hydropic swelling when seen under the microscope. Hydropic swelling or hydropic change is an increase in cell volume characterized by a large, pale and vacuolated cytoplasm and a normally located nucleus. This cellular swelling results from impairment of the process that controls ion (i.e., sodium) concentrations in the cytoplasm. Injurious agents can impair the energy-dependent Na+-K+ ATPase plasma membrane pump, PATHOLOGY 3500/PATHOLOGY 9535 3 CELL INJURY, ADAPTATION, AND CELL DEATH leading to an accumulation of sodium in the cell. This leads to an increase in water in the cell to maintain iso-osmotic conditions and the cell swells. Mitochondria may also swell, the cisternae of the endoplasmic reticulum also become dilated, and blebs may form on the plasma membrane. With removal of the stimulus, these changes are reversible, and the cell reverts to its normal state. Another form of reversible adaptation to cell stressors is fatty change or the abnormal accumulation of triglycerides within cells (see section on “Intracellular Accumulations”). IRREVERSIBLE CELL INJURY & CELL DEATH If overwhelming injury occurs or occurs at a rate at which the cell cannot adapt, necrosis or cell death is the result. Necrosis is characterized by certain structural changes; these include intense eosinophilia (pinkness) of the cytoplasm and pyknosis (shrinkage), karyorrhexis (the pyknotic nucleus fragments) and karyolysis (dissolution) of the nucleus. Several types of necrosis are seen. These are associated with different types of cell damage. Coagulative: The most common form of necrosis - microscopically all of the changes described above are seen (eosinophilia, pyknosis, karyorrhexis and karyolysis). Cells appear like ‘ghosts’ of themselves in which the basic structural outline of the coagulated cell persists for a number of days. Typical of ischemia, e.g., in heart (myocardial) cells. Liquefactive: rapid loss of tissue architecture and digestion of the dead cells. Most often seen in CNS. Typical of bacterial damage. Fat: specific to fat (adipose) tissue. Released enzymes digest fat that complexes with calcium to form chalky-white deposits, e.g., pancreatitis; damage to breast tissue. Caseous: soft, friable, ‘cheesy’ material. Characteristic of tuberculosis. Note: the term “gangrenous necrosis” or “wet gangrene” is used to refer to coagulative necrosis (most frequently of a limb) when there is superimposed infection with a liquefactive component. If the necrotic tissue dries out (with no infectious component) it becomes dark black and mummified and is called “dry gangrene”. PROGRAMMED CELL DEATH - APOPTOSIS Apoptosis is the morphologic manifestation of programmed cell death and is distinct from necrosis, the uncontrolled process of cell death in response to overwhelming injury. Apoptosis is an energy-dependent process specifically designed to switch off unneeded or damaged cells and eliminate them. Therefore, apoptosis can occur under either physiologic (e.g., during embryogenesis in shaping of fingers and toes; the physiological involution of thymus during development or endometrium during the menstrual cycle; removal of an infected or damaged cell) or pathologic (e.g., following radiation injury, in some cancers) conditions. Apoptotic cells initiate their own death by the activation of proteases known as caspases and endogenous endonucleases that breakdown the cell nucleus and cytoskeleton. The cell nucleus collapses, the cell shrinks and is cleaved into membrane-bound clumps enclosing organelles (apoptotic bodies). The membrane bound material is recognized and quickly engulfed by phagocytic cells. The magnitude and type of injurious stimulus can determine whether a cell undergoes apoptosis or necrosis. Severe damaging stimuli tend to result in necrosis and lower-grade stimuli and immune-mediated damage tend to cause apoptosis. A critical factor seems to be how much PATHOLOGY 3500/PATHOLOGY 9535 4 CELL INJURY, ADAPTATION, AND CELL DEATH cellular ATP is available after cell damage (remember apoptosis is an energy-dependent process!). Where there is severe depletion of ATP the necrotic pathway is followed. COMPARISON OF CELL DEATH BY APOPTOSIS AND NECROSIS FEATURE APOPTOSIS NECROSIS Physiological or pathological Due to pathological injury (e.g., Induction stimuli hypoxia, toxins) Extent Single cells Cell groups Gene activation; Energy- Impairment or cessation of ion dependent fragmentation of DNA homeostasis; depletion of ATP; influx by endogenous endonucleases Biochemical events; of sodium, water and calcium; cell and cleaved into regular mechanisms membrane damage; DNA fragmented nucleosomal fragments irregularly; may be mediated by free (laddering); breakdown of radicals cytoskeleton by proteases Cell membrane Maintained Lost integrity Cell shrinkage; fragmentation to Cell swelling & lysis; swelling & Morphology form apoptotic bodies with dense disruption or organelles; nucleus - chromatin pyknosis, karyorrhexis, karyolysis Inflammatory None Yes response Ingested (phagocytosed) by Ingested (phagocytosed) by Fate of dead cells macrophages neutrophils & macrophages MECHANISMS OF CELL INJURY Cell injury, whether reversible or irreversible, depends on both the type of injury (its duration and severity) and the injured cell (rate of mitotic activity, nutritional or hormonal status, blood supply, its genetic makeup). GENERATION OF REACTIVE OXYGEN SPECIES (ROS) Partially reduced or reactive oxygen species (e.g., superoxide anion, hydroxyl radical and hydrogen peroxide) are identified as a likely cause of cell injury. These highly reactive species can be formed via the action of ionizing radiation (hydrolysis of water produces hydroxyl radicals resulting in DNA damage and apoptosis in rapidly dividing cells); during reperfusion of tissues following a period of ischemia; in the presence of excess oxygen; and normally in inflammatory cells. All of these molecules possess a free electron which makes them highly reactive with a number of key cellular elements. They can initiate lipid peroxidation leading to a loss of membrane integrity, cross-link essential proteins, damage DNA or form secondary damaging ROS (e.g., lipid peroxide radicals, peroxynitrite). Because these molecules are also formed during normal metabolism and oxygen respiration the body has several mechanisms to detoxify these potentially damaging substances including spontaneous decay (e.g., superoxide breaks down in the presence of water into oxygen and hydrogen peroxide), superoxide dismutase (converts superoxide to hydrogen peroxide and oxygen), catalase (converts hydrogen peroxide to water and oxygen) or glutathione peroxidase PATHOLOGY 3500/PATHOLOGY 9535 5 CELL INJURY, ADAPTATION, AND CELL DEATH (reduction of hydrogen peroxide to water). Endogenous or exogenous (ingested) antioxidants such as vitamins A, C and E can also offer some protection against these free radical species. When these endogenous systems become overwhelmed damage can occur. ISCHEMIC AND HYPOXIC CELL INJURY The reduction or interruption of blood flow in ischemia is the most common type of cell injury and an important cause of coagulative necrosis. Ischemia can injure tissues faster than reduced levels of oxygen alone (hypoxia) since both oxygen and substrates for glycolysis and continued ATP generation disappear. Decreased oxygen compromises respiration in mitochondria damaging the ability to produce ATP. Decreased ATP impairs the ability of the cells to pump ions and water (via Na-K ATPase) with the subsequent accumulation of intracellular sodium and the diffusion of potassium out of the cell. The net gain of sodium is accompanied by an iso-osmotic gain of water resulting in acute cellular swelling and in the swelling of cellular components with damage and rupture to the cell plasma membranes. Decreased ATP also results in increased anaerobic glycolysis and the depletion of glycogen stores with a build-up of lactic acid in the cell (and hence a decrease in intracellular pH or a more acidic environment). Decreased ATP and pH levels cause ribosomes to detach from rough endoplasmic reticulum (RER) with a reduction in protein synthesis. DIAGRAM OF REVERSIBLE CELL INJURY, POINT OF NO RETURN AND CELL DEATH. Taken from: Oxford Textbook of Pathology, Volume 1, Principles of Pathology, (McGee, Isaacson and Wright), p148, 1992. If oxygen is restored all of the above disturbances are reversible; however, if ischemia or hypoxia is not relieved, worsening mitochondrial function and increasing membrane permeability cause further deterioration with irreversible cell injury and: The release of proteolytic enzymes from their normal compartments in the cell which induces widespread damage. PATHOLOGY 3500/PATHOLOGY 9535 6 CELL INJURY, ADAPTATION, AND CELL DEATH  In addition, calcium increases in cells due to pump failure (Ca-ATPase). Increase in intracellular calcium activates many enzyme systems inappropriately leading to further cell damage. Some of the enzyme systems attack cellular skeletal proteins which cause cells to become abnormally fragile; membrane damage is critical to the development of lethal cell injury. Leakage of cell proteins across the degraded cell membrane into the peripheral circulation provides a means of detecting tissue-specific cell injury and death by measuring their levels in blood serum samples (e.g., heart muscle cells contain a specific isoform of the enzyme creatine kinase and of the contractile protein troponin which become elevated in blood following a ‘heart attack’ or myocardial infarction and death of cardiac cells). ISCHEMIA/REPERFUSION INJURY Restoration of blood flow can result in cell recovery; however, under some circumstances, the restoration of blood flow can result paradoxically in additional cell injury. Thus, cells are lost during the ischemic episode and afterwards due to what is called ischemia/reperfusion injury. Reperfusion of ischemic tissues may cause further damage by: Restoration of blood flow bathes cells in high concentrations of calcium when they may not be able to fully regulate its intracellular level, activating destructive enzymatic pathways. Damaged mitochondria in compromised cells may yield increased production of reactive oxygen species which promote additional damage. Local increase of inflammatory cells that release high levels of ROS which promote additional membrane and mitochondrial damage. Injured cells may have compromised antioxidant defense mechanisms. CHEMICAL OR DRUG-RELATED CELL INJURY Chemicals can cause cell injury and damage directly (e.g., heavy metals; chemotherapeutic agents) or indirectly following metabolism to active metabolites. For example, carbon tetrachloride (CCl4) is metabolized by the liver cytochrome P450 enzyme complex to a highly reactive trichloromethyl free radical intermediate which begins a cascade of lipid peroxidation and cell membrane damage. This same P450 mixed-function oxidase system is responsible for the metabolism of a variety of drugs and chemicals others of which also form active, injurious metabolites. INTRACELLULAR ACCUMULATIONS Under normal circumstances, cells store fats, glycogen, vitamins and minerals for use in general cell metabolism. Cells also store the products of turnover as endogenous pigments such as degraded phospholipids as the golden-brown granules of the “wear-and-tear” pigment, lipofuscin, which increases with age, particularly in heart, lung and brain; melanin, an insoluble brown-black pigment, found in the skin and in certain brain cells; or hemosiderin, the iron-rich brown pigment derived from the breakdown of red blood cells. In hemosiderosis, the excess iron storage in skin, pancreas, heart, kidneys, and endocrine glands can damage vital organs. The most common exogenous pigment is carbon (from air pollution) which is inhaled and deposited in tracheobronchial lymph nodes and lung tissue. Many inherited disorders of metabolism may also lead to the abnormal accumulation of metabolites in cells, e.g., glycogen storage diseases; complex lipids; iron (hemochromatosis, an abnormality in iron absorption). Fatty change or steatosis is linked to the intracellular accumulation of fat either because of PATHOLOGY 3500/PATHOLOGY 9535 7 CELL INJURY, ADAPTATION, AND CELL DEATH increased delivery of fat to the cell (e.g., starvation, diabetes); an impairment of fat metabolism within the cell (e.g., in liver cells in alcoholism); or decreased synthesis of apolipoproteins for transport out of the cell (e.g., protein malnutrition, CCl4 toxicity). Small vacuoles of fat appear throughout the cytoplasm or may coalesce to form one large vacuole that displaces the nucleus. The liver, where most fats are stored and metabolized, is particularly susceptible to fatty change, but it may also occur in the heart, kidney, skeletal muscle, and other organs as a result of toxin exposure (e.g., alcohol, carbon tetrachloride), protein malnutrition or starvation, diabetes, obesity and anoxia. In Canada, the most common cause of fatty change in the liver is due to alcohol abuse. Fatty change is entirely reversible if the stimulus is removed. PATHOLOGY 3500/PATHOLOGY 9535 8 CELL INJURY, ADAPTATION, AND CELL DEATH STUDENT NOTES: PATHOLOGY 3500/PATHOLOGY 9535 9 CELL INJURY, ADAPTATION, AND CELL DEATH