Cell death

A PRESENTATION ON CELL DEATH

1. CELL DEATH PRESENTED BY : SREYA. S M. Pharmacy I. Semester Academic year:2019-2021 DEPARTMENT OF PHARMACOLOGY 1

2. DEFINITION Cell death  is the event of a  biological cell  ceasing to carry out its functions . This may be the result of the natural process of old cells dying and being replaced by new ones, or may result from such factors as disease, localized  injury, or the death of the organism of which the cells are part.  Cell death can occur in following ways- 1- Apoptosis (cell suicide)/ Autophagy 2- Necrosis (cell murder)   Apoptosis or Type I cell-death, and  autophagy  or Type II cell-death are both forms of programmed cell death, while necrosis is a non-physiological process that occurs as a result of infection or injury 2

3. APOPTOSIS The cells of a multicellular organism are members of a highly organized community. The number of cells in this community is tightly regulated—not simply by controlling the rate of cell division, but also by controlling the rate of cell death . If cells are no longer needed, they commit suicide by activating an intracellular death program . This process is therefore called programmed cell death, although it is more commonly called  apoptosis.  For an average human child between the ages of 8 to 14 years old approximately 20 to 30 billion cells die per day Apoptosis is a highly regulated process. Apoptosis can be initiated through one of two pathw.ays. In the intrinsic pathway the cell kills itself because it senses cell stress, while in the extrinsic pathway the cell kills itself because of signals from other cells   3

4. APOPTOSIS: PATHWAYS 4

5. REGULATION There are a variety of factors responsible for regulating apoptosis, both intracellular and extracellular. External signals can include growth factors or specific signals from other cells, whereas internal factors can include DNA damage or failure of cell division. Apoptosis Inducers Apoptosis Inhibitors Withdrawal of growth factors Loss of matrix attachment Glucocorticoids Some viruses Free radicals Ionising radiation DNA damage Ligand binding at ‘death receptors’ Presence of growth factors Extracellular cell matrix Sex steroids Some viral proteins 5

6. CASPASE ACTIVATION The mammalian initiator caspase-9 is activated as a complex with Apaf-1 and cytochrome c in the apoptosome . Caspase-9 then cleaves and activates effector caspases, such as caspase-3. The effector caspases cleave a variety of cell proteins, including nuclear lamins , cytoskeletal proteins, and an inhibitor of DNase , leading to death of the cell. 6

7. THE INTRINSIC PATHWAY OF APOPTOSIS This pathway triggers apoptosis in response to internal stimuli such as biochemical stress, DNA damage and lack of growth factors. This pathway of apoptosis is the result of increased mitochondrial permeability and release of pro-apoptotic molecules (death inducers) into the cytoplasm The release of these mitochondrial proteins (mostly cytochrome c) is controlled by a finely orchestrated balance between pro- and anti-apoptotic members of the Bcl family of proteins. This family is named after Bcl-2, which was identified as an oncogene in a B-cell lymphoma Bcl2 family is classified in to 3 categories   pro-apoptotic : promote apoptosis, inactive on normal cell. Eg Bax , Bak 7

8. anti-apoptotic : inhibit apoptosis ,ensure cell survival -Bcl-2 Bcl -xl , Bcl -w BH3-only proteins : only have a small BH3 domain . inhibit or promote apoptosis eg ; Bid, Bad, Bim , Puma, Noxa The pro-apoptotic molecules cause permeabilization of the outer mitochondrial membrane, leading to efflux of cytochrome c, which binds the adaptor Apaf-1 and the initiator caspase-9 in the cytosol to form the apoptosome complex.  This stimulates caspase-9, which in turn activates the effector caspases. The mitochondrion also releases a protein called Smac /DIABLO into the cytosol . Smac /DIABLO indirectly promotes apoptosis by blocking the effects of a group of anti-apoptotic proteins called inhibitor of apoptosis proteins (IAPs ). 8

9. The anti-apoptotic proteins Bcl-2 and Bcl -XL inhibit cytochrome c release, whereas Bax , Bak , and Bid, all pro-apoptotic proteins, promote its release from mitochondria. Cytochrome c and deoxyadenosine triphosphate ( dATP ) bind to APAF-1 to form a multimeric complex that recruits and activates pro-caspase-9, an apoptosis-mediating executioner protease that in turn activates the caspase cascade, resulting in cell apoptosis.   During this process, caspase-2, caspase-8, caspase-9 and caspase-10 are involved in the initiation of apoptosis. Caspase-3, caspase-6 and caspase-7 are involved in apoptosis. Caspase-3 and caspase-7 regulate the inhibition of DNA repair and start DNA degradation. In addition, caspase-6 regulates the disintegration of the lamina and cytoskeleton. 9

10. Mitocondrial pathway 10 H 2 O 2 casp9 Bcl2 PI3K Akt BAD Apaf1 Cyt.C ATP casp3 casp3 IAPs Smac/ DIABLO AIF Bax Bax p53 Fas Casp8 Bid Bid Bid DNA damage

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12. THE EXTRINSIC PATHWAY OF APOPTOSIS Apoptosis is known as a physiological process of cell deletion and is also a process of programmed cell death, resulting in morphological change and DNA fragmentation. It is stimulated by external or internal events of cells, one of which is the extrinsic pathway mediated by the death receptor. The death receptors include Fas receptors, tumor necrosis factor (TNF) receptors, and TNF-related apoptosis-inducing ligand (TRAIL) receptors . 12

13. As a surface receptor, for example, TNF receptor-1 (TNF-R1), it will interact with TNF to induce the recruitment of adaptor proteins such as Fas -associated protein with death domain (FADD) and Tumor necrosis factor receptor type 1-associated DEATH domain protein (TRADD), which recruits a series of downstream factors, including Caspase-8, which is a critical mediator of the extrinsic pathway, resulting eventually in cell apoptosis. 13

14. The extrinsic pathway that initiates apoptosis is triggered by a death ligand binding to a death receptor, such as TNF-α to TNFR1 The TNFR family is a large family consisting of 29 transmembrane receptor proteins, organized in homotrimers and activated by binding of the respective ligand (s). They share similar cysteine -rich extracellular domains and have a cytoplasmic domain of about 80 amino acids called the "death domain" (DD). Besides TNFR1, the Fas and DR4/DR5 also involved the pathway as death receptors and bind CD95 and TRAIL, respectively. All of the ligand binding to receptors will lead, with the help of the adapter proteins (FADD/ TRADD) to recruitment, dimerization , and activation of a caspase cascade and eventually cleavage of both cytoplasmic and nuclear substrates. 14

15. 15 Receptor trimerization results in recruitment of several death domains and eventually recruitment and activation of caspase-8 and caspase-10. Active caspase-8 and caspase-10 then either initiate apoptosis  

16. FLIP This pathway of apoptosis can be inhibited by protein called FLIP binds to pro-caspase-8 but cannot cleave and activate the caspase because it lacks a protease domain. Some viruses and normal cells produce FLIP and protect themselves from Fas -mediated apoptosis . 16

17. NECROSI S It is defined as focal death along with protein denaturation & degradation of tissue by hydrolytic enzymes liberated by cells. It is accompanied by inflammatory reaction Necrosis is caused by various agents such as hypoxia, chemical and physical agents, microbial agents and immunological injury Two essential changes bring about irreversible cell injury in necrosis-cell digestion by lytic enzymes and denaturation of proteins of proteins 17

18. Types of Necrosis Cogulative necrosis Liquefactive necrosis Gangrenous necrosis Caseous necrosis Fat necrosis Fibrinoid necrosis 18

19. COAGULATIVE NECROSIS This type of necrosis is caused by irreversible focal injury, mostly from sudden cessation of blood flow ( ischaemia ) and less often from bacterial and chemical agents A form of tissue necrosis in which the component cells are dead but the basic tissue architecture is preserved for at least several days. Presumably the injury denatures not only structural proteins but also enzymes and so blocks the proteolysis of the dead cells; as a result, eosinophilic , anucleate cells may persist for days or weeks. the necrotic cells are removed by phagocytosis of the cellular debris by infiltrating leukocytes and by digestion of the dead cells by the action of lysosomal enzymes of the leukocytes. 19

20. Coagulative necrosis is characteristic of infarcts (areas of ischemic necrosis) in all solid organs except the brain. 20

21. LIQUEFACTIVE NECROSIS This type of necrosis occurs commonly due to ischaemic injury and bacterial or fungal infections It is seen in focal bacterial or, fungal infections, because microbes stimulate the accumulation of inflammatory cells and the enzymes of leukocytes digest ("liquefy") the tissue in to a liquid viscous mass If the process was initiated by acute inflammation, the material is frequently creamy yellow and is called pus. E.g.- hypoxic death of cells within the central nervous system often evokes liquefactive necrosis . 21

22. Gross Appearance: The tissue is in a  liquid form and sometimes creamy yellow because of pus formation. Microscopic: Inflammatory cells with numerous neutrophils . 22

23. CASEOUS NECROSIS Caseous necrosis is encountered most often in foci of tuberculous infection, caused by syphilis, certain fungi The term " caseous " (cheese-like) is derived from the friable yellow-white appearance of the area of necrosis. On microscopic examination, the necrotic focus appears as a collection of fragmented or lysed cells with an amorphous granular appearance. Caseous necrosis is often enclosed within a distinctive inflammatory border; this appearance is characteristic of a focus of inflammation known as a granuloma . 23

24. FAT NECROSIS In fat necrosis the enzyme lipase release fatty acid from triglycerides, then it form complex with calcium to form soaps This processed is usually triggered by several factors leading to inflammation of the pancreas, otherwise known as pancreatitis, can also occur in breast, salivary glands Causes of acute pancreatitis include alcohol, gall bladder stones, poisoning, and insect bites. Since fat necrosis in the pancreas is triggered by an inadvertent release of enzymes, this process is also referred to as enzymatic fat necrosis. The trigger for necrosis in breast is usually trauma. 24

25. Gross Appearance: Whitish deposits as a result of the formation of calcium soaps. Microscopic: Anucleated adipocytes with deposits of calcium (Seen on H&E as areas of bluish stains ) 25

26. GANGRENOUS NECROSIS A type of tissue death caused by lack of blood supply or infection , also associated with diabeties and long term tobacco smoking Symptoms change in skin colour to red or black Types dry , wet , gas etc Dry gangrene is a form of coagulative necrosis that develop in ischemic tissues ,also due to peripheral artery disease Wet gangrene is characterised by thriving bacteria and poor prognosis.It is infected by saprogenic microorgasnism cause tissue to swell and emit bad smell. Gas gangrene due to bacteria,produce gas within tissue eg clostridium 26

27. FIBRINOID NECROSIS Fibrinoid necrosis is a pattern of cell death characterized by endothelial damage and exudation of plasma proteins This pattern of necrosis is prominent when complexes of antigens and antibodies are deposited in the walls of arteries. Deposits of these "immune complexes," together with fibrin that has leaked out of vessels, result in a bright pink and amorphous appearance in H&E stains, called " fibrinoid " (fibrin-like) by pathologists. The immunologically mediated diseases (e.g., polyarteritis nodosa ) in which this type of necrosis is seen. 27

28. Gross Appearance: Usually not grossly discernible. Microscopic: Deposition of fibrin within blood vessels . 28

29. AUTOPHAGY The term ‘ autophagy ’, derived from the Greek meaning ‘eating of self’, It is the natural, regulated mechanism of the cell that removes unnecessary or dysfunctional components.  It allows the orderly degradation and recycling of cellular components There are 3 types   Macroautophagy Microautophagy Chaperone-mediated autophagy  (CMA) 29

30. initial sequestered closes into a double membrane vesicle the  autophagosome some autophagosomes formed in a PI3P-enriched ( phosphatidylinositol 3-phosphate) membrane compartment dynamically connected to the endoplasmic reticulum an autophagosome fuses with a lysosome Regulated process of the degradation and recycling of organelles and cellular components Resulting in organelle turnover and in the bioenergetics of starvation Could result in cell death through excessive self-digestion and degradation of essential cellular constituents 30

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32. MACROAUTOPHAGY is the main pathway, used primarily to eradicate damaged cell organelles or unused proteins.  First the phagophore engulfs the material that needs to be degraded, which forms a double membrane known as an  autophagosome , around the organelle marked for destruction.  The  autophagosome then travels through the cytoplasm of the cell to a lysosome , and the two organelles fuse .Within the lysosome , the contents of the autophagosome are degraded via acidic lysosomal hydrolase 32

33. MICROAUTOPHAGY It is the direct uptake of soluble or particulate cellular constituents into  lysosomes . It translocates cytoplasmic substances into the lysosomes for degradation via direct  invagination , protrusion, or septation of the lysosomal limiting membrane 33

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35. CHAPERONE-MEDIATED AUTOPHAGY It is a very complex and specific pathway, which involves the recognition by the hsc70-containing complex. This means that a protein must contain the recognition site for this hsc70 complex which will allow it to bind to this chaperone, forming the CMA- substrate/chaperone complex. This complex then moves to the lysosomal membrane-bound protein that will recognise and bind with the CMA receptor. Upon recognition, the substrate protein gets unfolded and it is translocated across the lysosome membrane with the assistance of the lysosomal hsc70 chaperone. CMA is significantly different from other types of autophagy because it translocates protein material in a one by one manner, and it is extremely selective about what material crosses the lysosomal barrier. 35

36. REFERENCES  Apoptosis an overview. www.sciencedirect.com. ScienceDirect Topics. A poptosis." Reproductive and Cardiovascular Disease Research Group, St. George's, University of London Renehen , Andrew G., Catherine Booth, Christopher S. Potten . "What is apoptosis, and why is it important?" British Medical Journal Levine B, Kroemer G. Autophagy in the pathogenesis of disease. Cell 2008;132:27–42. [ PubMed : 18191218] Mizushima N. Autophagy : process and function. Genes Dev 2007;21:2861–2873. [ PubMed : 18006683] Molecular Biology of the Cell 4th ed. - IV. Internal Organization of the Cell Chapter 17. The Cell Cycle and Programmed Cell Death "necrosis" Molecular Biology of the Cell -Cell- A molecular Approach 36

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