1. CELL INJURY ( PART 1) Sufia Husain Assistant Prof & Consultant KKUH, Riyadh. September 2014

2. CELL INJURY When the cell is exposed to an injurious agent or stress, a sequence of events follows that is loosely termed cell injury. Cell injury is reversible up to a certain point, but if the stimulus persists or is severe enough from the beginning, the cell reaches a point of no return and suffers irreversible cell injury and ultimately cell death. Cell death, is the ultimate result of cell injury

3. CELL DEATH There are two principal patterns of cell death, necrosis and apoptosis. Necrosis is the type of cell death that occurs after ischemia and chemical injury, and it is always pathologic. Apoptosis occurs when a cell dies through activation of an internally controlled suicide program.

4. C AUSES OF C ELL I NJURY Causes of both reversible and irreversible injury are the same 1) Oxygen Deprivation (Hypoxic cell injury). It common cause of cell injury and cell death. Hypoxia can be due to a) Ischemia/hypoxia (obstruction of arterial blood flow), the most common cause seen in myocardial infarction and atherosclerosis. b) Inadequate oxygenation of the blood e.g. lung disease and carbon monoxide poisoning c) Decreased oxygen-carrying capacity of the blood e.g. anemia d) Inadequate tissue perfusion due to cardiorespiratory failure, hypotension, shock etc Depending on the severity of the hypoxic state, cells may adapt, undergo injury, or die. Also some cell types are more vulnerable to hypoxic injury then others e.g. neurons are most susceptible followed by cardiac muscle, hepatocytes and then skeletal muscles.

5. C AUSES OF C ELL I NJURY CONT. 2) Physical Agents e.g. mechanical trauma, burns and deep cold, sudden changes in atmospheric pressure, radiation, and electric shock 3)Chemical Agents and Drugs e.g. oxygen in high concentrations, poisons, pollutants, insecticides, industrial and occupational hazards, alcohol and narcotic drugs and therapeutic drugs. 4)Infectious Agents 5) Immunologic agents e.g. thyroid damage caused by autoantibodies. 6) Genetic Derangements eg sickle cell anemia. 7) Nutritional Imbalances

6. MECHANISM OF CELL INJURY 1. Depletion of ATP 2. Cell membrane damage/defects in membrane permeability: Membrane damage may be  indirect via hypoxia, ATP depletion and activation of phospholipases.  direct by certain bacterial toxins, viral proteins, complement mediated lysis via membrane attack complex (MAC) and by free radicals (reactive oxygen species). 3. Mitochondrial damage:  It is seen specially in hypoxic injury and cyanide poisoning.

7. MECHANISM OF CELL INJURY CONT. 4. Ribosomal damage:  It is seen in alcohol associated damage of liver cells and with antibiotic use bacterial infection. 5. Nuclear and DNA damage: 6. Influx of intracellular calcium and loss of normal calcium balance (homeostasis): ischemia causes an increase in intracellular calcium concentration. Increased Ca2+ in turn activates a number of enzymes which cause damage.

8. MECHANISM OF CELL INJURY CONT. 7. Accumulation of oxygen-derived free radicals (oxidative stress): Free radicals are highly reactive and harmful atoms that have single unpaired electron in an outer orbit. They are referred to as reactive oxygen species/free radicals. The production of free radicals are initiated within cells in several ways called as the free radical generating systems. They are produced via: i. Normal metabolism/ respiration: Small amounts of harmful reactive oxygen forms are produced as a bi-product of mitochondrial respiration (metabolic reduction-oxidation reactions that occur during normal metabolic processes). During normal respiration, small amounts of free radicals are produced. ii. Via ionizing radiation injury (absorption of radiant energy e.g. ultraviolet light, x-rays or any other type of radiation). iii. Chemical toxicity: enzymatic metabolism of exogenous chemicals or drugs. iv. Oxygen therapy and reperfusion injury v. Immune response or inflammation (neutrophil oxidative burst) vi. Transition metals such as iron and copper can trigger production.  The common free radicals are  superoxide anion radical (O2-),  hydrogen peroxide (H2O2),  and hydroxyl ions (OH).  Nitric oxide (NO) is an important chemical mediator generated by various cells and it can also act as a free radical.

9. MECHANISM OF CELL INJURY CONT.  Free radicals cause damage to lipids, proteins, and nucleic acids. The main effects of these reactive species/ free radicals are: 1. Lipid peroxidation of membranes: leads to damage of membranes, organelles etc. 2. Oxidative modification of proteins: leads to protein fragmentation. 3. DNA damage: can lead to cell aging and malignant transformation of cells.  Certain substances in the cells remove/ inactivate the free radicals in order to minimize injury caused by them. They are called as the free radical scavenging system. They are:  Antioxidants: vitamins E, A and C (ascorbic acid).  Enzymes: which break down hydrogen peroxide and superoxide anion e.g. Catalase, Superoxide dismutases, Glutathione peroxidase and mannitol.  Any imbalance between free radical-generating and radical-scavenging systems results in oxidative stress causing cell injury.

10. Cellular and biochemical sites of damage in cell injury. Downloaded from: Robbins & Cotran Pathologic Basis of Disease (on 4 September 2005 02:13 PM) © 2005 Elsevier

11. M ECHANISM IN HYPOXIC CELL INJURY Hypoxia Decreased oxidative phosphorylation Decreased ATP Membrane damage and proton pump failure Loss of ion gradient Anaerobic respiration leading to excess lactic acid production Membrane, organelle and nuclear damage Release of Calcium from mitochondria Calcium influx into cell from extracellular space which leads to further injury

12. R EVERSIBLE C ELL I NJURY The type of injury, the time duration of injury and the severity of injury will determine the extent of cell damage i.e. whether the injury is reversible or irreversible. Earliest changes associated with cell injury are reversible. They are: 1. Swelling of cell cytoplasm with vacuolization of cytoplasm called hydropic or vacuolar degeneration. 2. Mitochondrial swelling. 3. Fatty change. 4. Plasma membrane blebbing. 5. Dilation and degranulation of the rough endoplasmic reticulum leading to loss of protien synthesis. 6. Eosinophilia (due to decreased cytoplasmic RNA) Within limits, the cell can compensate for these derangements and, if the injurious stimulus is removed the damage can be reversed.

13. I RREVERSIBLE C ELL I NJURY Persistent or excessive injury, however, causes cells to pass the threshold into irreversible injury. Irreversible injury is marked by 1. severe mitochondrial dilatation and damage with the appearance large, amorphous densities in mitochondria. 2. extensive damage and disruption of plasma/cell membrane 3. increased eosinophilia 4. Numerous myelin figures 5. swelling and rupture of lysosomes leakage and enzymatic digestion of cellular contents 6. Nuclear damage: i. pyknosis (shrinkage), ii. karyolysis (dissolution) iii. karyorrhexis (break down)

14. n Downloaded from: Robbins & Cotran Pathologic Basis of Disease (on 4 September 2005 10:51 AM) © 2005 Elsevier

15. NECROSIS. Necrosis is changes that follow cell death in living tissue, due to enzymatic digestion and denaturation of intracellular protein in the injured cell. It occurs in irreversible injury. It is usually associated with inflammation in the surrounding tissue. It involves the death of a group of cells in one area. Necrosis can result in:  Cessation of function of the involved tissue or organ  Release of certain cellular enzymes that can be detected in blood. The level of these enzymes can be used as markers to diagnose the injury and also can help determine the time and the extent of injury eg. Cardiac enzymes in myocardial infarction (heart attack).  An inflammatory response

16. The enzymes used in this degradation come from either the lysosomes of the dying cell itself (referred to as autolysis) or from lysosomes of neighbouring leukocytes (referred to as heterolysis). Autolysis is the death/disintegration of cells or tissues by it’s own enzymes. Autolysis is also seen in cells after death/ post mortem. Autolysis is also seen in some pathologic conditions in living organisms.

17. T YPES OF NECROSIS There are 5 types of necrosis:  coagulative necrosis  liquefactive necrosis  caseous necrosis  fat necrosis  fibrinoid necrosis

18. C OAGULATIVE NECROSIS : Coagulative necrosis is characteristically seen due to blood loss leading to ischemic/hypoxic death of cells in all tissues except brain. e.g. heart (myocardial infraction), kidney( renal cortical necrosis/ infarct), spleen (infarct) etc. Coagulative necrosis is not seen in the brain. Gross: The affected organ looks pale and firm/solid. It looks liked cooked meat or boiled egg. Microscopy: In it there is preservation of the general tissue architecture and initially the basic ghost outline of the coagulated cell remains preserved for a few days. The cell cytoplasm is eosinophilic with loss of nucleus. Ultimately, the necrotic cells are removed by phagocytosis by the macrophages.

19. K IDNEY : COAGULATIVE NECROSIS

20. Kidney: coagulative necrosis Gross: tissue is firm Micro: Cell outlines are preserved (cells look ghostly), and everything looks red

21. L IVER COAGULATIVE NECROSIS

22. L IQUEFACTIVE NECROSIS is a type of necrosis which results in transformation of the tissue into a liquid viscous mass. Is characteristically seen in hypoxic cell death in the central nervous system/brain and in suppurative infections (pus or abscesses) especially bacterial. The affected tissue is softened/liquefied by the action of hydrolytic enzymes released from the lysosomes in the brain cells or in case of an abscess due to the enzymes released from the neutrophils. Ultimately, in a living patient most necrotic cells are phagacytosed. The affected area is soft with liquefied creamy yellow centre containing necrotic cells, and neutrophils and is called pus/abscess.

23. L IQUEFACTIVE NECROSIS

24. L IQUEFACTIVE NECROSIS ( CENTER LABELED ONE IS NECROSIS AND SURROUNDING IS NEUTROPHILS.

25. C ASEOUS NECROSIS is a type of coagulative necrosis classically seen in tuberculous infection. Gross: White, soft, curd like, cheesy-looking (“caseous”) material. On microscopic examination, the necrotic area appears as amorphous pink granular debris surrounded by a collar of epitheloid cells (modified macrophages), lymphocytes and giant cells. This is known as granuloma. Here the tissue architecture is completely obliterated.

26. T UBERCULOUS LUNG WITH A LARGE AREA OF CASEOUS NECROSIS. T HE CASEOUS DEBRIS IS YELLOW - WHITE AND CHEESY

27. F AT NECROSIS Is focal areas of fat destruction/ necrosis due to enzymatic action of lipase (which is released from injured pancreatic tissue) into the surrounding fat in the abdominal cavity. It is typically seen in acute pancreatitis. Damaged cells release lipases, which breakdown the fat cells into glycerol and free fatty acids. The produced fatty acids combine with calcium in the blood to produce calcium soaps (called as fat saponification) which looks like chalky white spots in the necrotic fat. The outlines of necrotic/dead fat cells can be seen. Inflammation is minimal. Fat necrosis can also be seen in breast fat and other fatty areas due to traumatic injury.

28. Figure 1-21 Foci of fat necrosis with saponification in the mesentery. The areas of white chalky deposits represent calcium soap formation at sites of lipid breakdown. Downloaded from: Robbins & Cotran Pathologic Basis of Disease (on 4 September 2005 10:51 AM) © 2005 Elsevier

29. Normal adipose tissue Fat necrosis

30. F IBRINOID NECROSIS Is necrosis in the blood vessels (arteries, arterioles and capillaries) There is deposition of fibrin material in the arterial walls, which appears smudgy and acidophilic/eosinophilic. It is seen in immune mediated diseases (autoimmune diseases) and also seen in malignant hypertension.

31. F IBRINOID NECROSIS

32. G ANGRENOUS NECROSIS Gangrenous necrosis is term commonly used in clinical practice by surgeons. It is usually seen in limbs, generally the lower leg, that has lost its blood supply and has undergone coagulative necrosis, called dry gangrene/mummification. It is the non-infected ischemic necrosis accompanied with drying of tissue. It is seen as a complication in atherosclerosis and diabetes mellitus. When there is superadded bacterial (putrefactive) infection coagulative necrosis is modified by the action of the bacteria into liquefactive necrosis, called wet gangrene. So, initially there is coagulative necrosis and then there is superadded liquefactive necrosis. The bacteria is usually gram-positive Clostridia. Clostridia lives in the gut or the soil and it can thrive in low oxygen states. The infection can spread to the rest of the body and be life threatening, so the limb has to be amputated. The limb becomes foul smelling and black and starts decomposing.