Cell Death Lecture ATBU Bauchi Sep 2024 PDF

CELL DEATH DR. RAYMOND VHRITERHIRE ATBU BAUCHI 9TH SEPTEMBER 2024  CELL DEATH Cell death is the natural endpoint of normal cell physiology which result in the irreversible termination of cellular functions such as growth, division and homeostasis. Cell death...

CELL DEATH DR. RAYMOND VHRITERHIRE ATBU BAUCHI 9TH SEPTEMBER 2024  CELL DEATH Cell death is the natural endpoint of normal cell physiology which result in the irreversible termination of cellular functions such as growth, division and homeostasis. Cell death is essential to maintain cell physiology and function, and in some situations dysfunctional, worn- out or damaged cells are removed from the system through the process of cell death. Cell death, survival, proliferation and differentiation are fundamental and essential processes of life. Thus cell death may occur as a component of a physiological process or a response to a pathological condition.  CELL DEATH In this regard, a homeostatic mechanisms maintain cells in a constant state of equilibrium, within a narrow metabolic window allowing minimal deviation. However, stressors beyond homeostatic mechanisms lead to reversible injury initially and later to irreversible injury when the stress is either severe or prolonged. Irreversible injury may lead to cell death.  TYPES OF CELL DEATH 1. Apoptosis [type 1 6. Anoikis cell death] 7. Entosis 2. Autophagy [type II 8. Ferroptosis cell death] 9. Pyroptosis 3. Necrosis [type II cell death] 10. Paraptosis 4. Necroptosis 11. Parthanatos 5. Lysosome 12. Mitoptosis dependent cell 13. NETosis death Apoptosis and necrosis are the two principal types of cell death  TYPES OF CELL DEATH Feature Necrosis Apoptosis Autophagy Mode of cell Accidental Programmed Programmed death Inflammatory Proinflammatory No inflammation No inflammation response Initiators Toxins, infections, TNFa, FasL, TRAIL, Nutrient trauma, inflammation infections deprivation, hypoxia, infection Cell size Enlarged, swelling Reduced, shrinkage Reduced Nucleus Pyknosis, karyorrhexis, Fragmentation karyolysis Plasma Disrupted Intact membrane Cell contents Enzyme digestion Intact Physiologic or Usually pathological Pathological and Physiological, pathological physiological pathological  NECROSIS Necrosis is the pathological endpoint of severe injury which is characterized by characterized by denaturation of proteins, leakage of cellular contents through damaged, membranes, local inflammation, and enzymatic digestion of the lethally injured cell. Severe injury lead to cell membrane damage which lead to leakage of cell contents into the extracellular space.  NECROSIS Damage associated molecular patterns which may elicit inflammation include the following: 1. HMBG-1 (high motility group box 1 protein) stimulates movement of macrophages, dendritic cells and neutrophils to the site of injury. 2. Heat shock proteins, Hsp 70, HSP 90 and Uric acid both of which act on TLR2 and TLR4 to trigger inflammation. 3. IL-6 4. DNA, mRNA, ATP and nucleosides to immunostimulate macrophages and dendritic cells.  NECROSIS Morphology Necrotic cells show increased eosinophilia, glassy homogeneous appearance, vacuolated cytoplasm, myelin figures. Nuclear changes include karyolysis, karyorrhxis, pyknosis. Morphological patterns of Tissue Necrosis 1. Coagulative Necrosis 4. Caseous necrosis 2. Liquefactive Necrosis 5. Fat necrosis 3. Gangrenous necrosis 6. Fibrinoid necrosis  NECROSIS Coagulative necrosis This is a form of necrosis in the architecture of dead tissue is preserved. There is denaturation of both structural proteins and enzymes. This blocks proteolysis and the shape and structure of the cell is preserved. This is the type of cell death that is caused by ischaemia. A localized area of coagulative necrosis is called an infarct.  NECROSIS Liquefactive necrosis This is characterized by digestion of the dead cells, resulting in transformation of the tissue into a viscous liquid. This is found in bacterial and fungal infections. The yellow digested material is called pus. Pus consists of leukocytes and necrotic cellular debris. Liquefactive necrosis is the only type of necrosis which occur in the brain.  NECROSIS Gangrenous necrosis is not a specific pattern of cell death. Dry gangrene if it is coagulative and wet gangrene if liquefactive due to bacterial infection Caseous Necrosis is seen in tuberculosis infection and it appears white and like cheese. Histologically consists of a granuloma with central area of necrosis surrounded by chronic inflammatory cells and fibrosis.  NECROSIS Fat necrosis occurs when there is damage to adipose tissue such as digestion which occurs in pancreatitis. Histologically may consist of ghost adipocytes, inflammatory cells and calcification. Fibrinoid necrosis occurs when there is vascular damage with deposition of immune complexes and plasma proteins (fibrin). There is a bright pink appearance of the wall of the involved arteries.  APOPTOSIS Apoptosis is a synchronized, genetically controlled programmed cell death that allows the body to self- regulate, eliminate, and dispose of unwanted and dysfunctional cells, without spillage of its contents into the surrounding environment. Apoptosis may have physiological or pathological causes. Physiological causes of apoptosis: Removal of supranumerary cells during embryonic development. E.g. lack of apoptosis can lead to webbed fingers (syndactyly) (fusion of two or more digits). Involution of hormone-dependent tissues on hormone withdrawal  APOPTOSIS Cell turnover in proliferating cell populations, such as immature lymphocytes in the bone marrow. Elimination of potentially harmful self-reactive lymphocytes to prevent autoimmunity. Death of host cells that have served their useful purpose, such as neutrophils in an acute inflammatory response, and lymphocytes at the end of an immune response. Pathological causes of apoptosis include the following: 1. DNA damage 2. Accumulation of misfolded proteins 3. Infections especially by viral agents.  APOPTOSIS 4. Immune responses in viral infections, tumours, transplant rejection. 5. Atrophy caused by obstruction of the duct of an organ Morphological changes in apoptosis 1. Cell shrinkage 2. Chromatin condensation (pyknosis), nuclear fragmentation (karyorrhexis), and plasma membrane blebbing. 3. Formation of apoptotic bodies which are intensely eosinophilic, 4. Phagocytosis of apoptotic bodies with intact membranes. 5. Absence of inflammation.  APOPTOSIS Identification of cells undergoing apoptosis 1. Microscopy (as above) 2. Flow cytometry with staining for Annexin V 3. DNA fragmentation assays Mechanisms of Apoptosis 1. Initiation phase 2. Execution phase  APOPTOSIS Initiation involve two pathways mitochondrial pathway and the death receptor pathway. 1.Mitochondrial Pathway of Apoptosis – Intrinsic Pathway This is the most commonly used pathway in both physiologic and pathological conditions. This is initiated by the loss of Cytochrome c from the mitochondrion into the cytoplasm of the cell.  APOPTOSIS Intrinsic Pathway is regulated by these factors. 1. Anti-apoptotic proteins: BCL2, BCL-XL, and MCL1 2. Pro-apoptotic: BAX, BAK, Bok 3. Regulated apoptosis initiators: Bad, Bim, Bid, Bik, Bmf, Hrk, Puma, and Noxa. In the cytosol, cytochrome c binds to APAF-1 (apoptosis-activating factor-1), forming a multimeric structure called the apoptosome. The apoptosome binds to Caspace – 9 which triggers a cascade of caspaces leading to formation of caspace -3.  APOPTOSIS EXTRINSIC PATHWAY OF APOPTOSIS This involve the use of death receptors present on the plasma membrane of many cells. Death receptors are members of the TNF family which have cytoplasmic death domain. Best known is the type 1 TNF receptor with related protein Fas(CD95). The ligand which binds to Fas is expressed on many T cells that recognise self antigens and cytotoxicT lymphocytes.  APOPTOSIS EXTRINSIC PATHWAY OF APOPTOSIS Fas ligand[FasL] binds to Fas. Fas then binds to FAAD (Fas-associated death domain) FAAD is an adaptor protein which also has a death domain. FAAD then activates pro-caspase-8 to form active autocatalytic caspase-8. Caspase-10 is the form of this caspase found in humans. This is the initiator caspasefor the extrinsic pathway.  APOPTOSIS EXECUTION PHASE OF APOPTOSIS Both intrinsic and extrinsic pathways converge on the execution pathway. Caspase-9 from intrinsic and Caspase-8 (caspase-10) from extrinsic. Each activate caspase 3 or caspase 6 which are the executioner caspases.  APOPTOSIS EXECUTION PHASE OF APOPTOSIS Some actions of executioner caspases 1. Cleave cytoskeletal and nuclear matrix proteins. 2. Disrupt the cytoskeleton 3. Nuclear breakdown 4. Cleave proteins involved in transcription, DNA replication, and DNA repair.  APOPTOSIS Recognition and removal of dead cells Phosphatidylserineare exposed on apoptotic cells and are recognised by receptors on the surface of macrophages. Some apoptotic cells secrete thrombospondin recognised by macrophage and phagocytes. Apoptotic bodies may be coated by complement. Annexin V stain is used to identify apoptotic cells The process of apoptotic cell phagocytosis is called efferocytosis.  AUTOPHAGY Autophagy is the self-digestive process that delivers cytoplasmic material of endogenous or exogenous origin to the lysosome for degradation. Autophagy as a means of cell killing was first advanced by Clark’s phenotypic description of “type II autophagic cell death” in 1990. Autophagy is initiated upon cellular stress as a protective response. Once the cellular stress is irreversible, the cell will be committed to death through excessive levels of autophagy. Cell death may also be directly due to the severity of the stressor itself.  AUTOPHAGY Forms of Autophagy There are three main forms of autophagy 1. Macro-autophagy, 2. Micro-autophagy, 3. Chaperone mediated autophagy (CMA) Macro-autophagy This is the most widely described form which occur in the following steps. Nucleation and formation of isolation membrane known as phagophore.  AUTOPHAGY Formation of a vesicle, called the autophagosome, from the isolation membrane. Sequestration of intracellular organelles inside the autophagosome. Fusion of autophagosomes with lysosomes and degradation of its contents by the lysosomal enzymes. Microautophagy Cytosolic contents are directly transferred through the membrane infoldings into the lysosome, where acidic hydrolases further mediate the degradation.  AUTOPHAGY Microautophagy involve the following steps 1. Membrane invagination and autophagic tubes formation. 2. Vesicle formation 3. Vesicle expansion and scission 4. Vesicle degradation and recycling Chaperone Mediated Autophagy CMA is the selective transfer of proteins containing KFERQ-like sequences (Lys-Phe-Glu-Arg-Gln) directly across the lysosomal membrane. These proteins can be recognized by chaperones, are subsequently hijacked into lysosomes, and eventually degraded.  AUTOPHAGY Selective autophagy Various nomenclature depict selective autophagy forms, depending on the organelles eliminated: 1. Mitophagy (for mitochondrion), 2. Ribophagy (for ribosomes), 3. Reticulophagy (for endoplasmic reticulum), 4. Lysophagy (for lysosomes), 5. Pexophagy (for peroxisomes), 6. Lipophagy (for lipid drops), 7. Glycophagy (for glycogen), aggrephagy (misfolded proteins), 8. Xenophagy (foreign pathogens)  AUTOPHAGY In certain circumstances, autophagy can lead to cell death. autophagy-dependent cell death (ADCD) can be defined as the regulated cell death (RCD) that depends on autophagy machinery and does not involve alternate death pathways. ADCD comprises three types: 1. Excessive autophagy, 2. Excessive mitophagy, 3. Autosis  ANOIKIS Anoikis is a form of programmed cell death which occur in the absence of any attachment to extracellular matrix (ECM) or upon any inappropriate cell adhesion. The attachment of cells to the extracellular matrix is essential for cell survival. Integrin receptors which act as a mediator of cell–ECM interaction, provide physical links with the cytoskeleton and transduce signals from the ECM to the cell. Integrins suppress apoptosis in attached cells by eliciting antiapoptotic and pro-survival signals from the ECM.  ANOIKIS Anoikis ensures tissue homeostasis by preventing detached cells from re-adhesion to new matrices in incorrect locations and their dysplastic growth or proliferating at ectopic sites where the ECM proteins are different from the original ones. Different pathways execute the initiation and execution of anoikis. These pathways activate caspases and downstream molecular pathways like activation of endonucleases, DNA fragmentation, and cell death. The induction of the anoikis program occurs through the interplay of two apoptotic pathways, the intrinsic pathway and the extrinsic pathway ANOIKIS  NECROPTOSIS Necroptosis can be defined as the regulated form of necrosis facilitated by death receptors. Morphologically, and to some extent biochemically, it resembles necrosis, as both are characterized by loss of ATP, swelling of the cell and organelles, generation of reactive oxygen species (ROS), release of lysosomal enzymes, and ultimately rupture of the plasma membrane. Necroptosis initiation involve factors of the exstrinsic pathway.  NECROPTOSIS Immune ligands, such as Fas, TNF-α, and lipopolysaccharide (LPS), initiate the process of necroptosis, leading to RIPK-3 (receptor-interacting protein kinase-3) activation followed by MLKL (mixed lineage kinase domain-like protein) phosphorylation. This phosphorylated MLKL travels towards the plasma membrane’s inner compartments and disrupts cell integrity. Necroptosis leads to increased cytokine and decreased DAMPs secretion  NECROPTOSIS Immune ligands, such as Fas, TNF-α, and lipopolysaccharide (LPS), initiate the process of necroptosis, leading to RIPK-3 (receptor-interacting protein kinase-3) activation followed by MLKL (mixed lineage kinase domain-like protein) phosphorylation. This phosphorylated MLKL travels towards the plasma membrane’s inner compartments and disrupts cell integrity. Necroptosis leads to increased cytokine and decreased DAMPs secretion. Necroptosis occur in physiologic and disease states eg CMV infection, ischemic-reperfusion injury.  OTHER CELL DEATH TYPES Pyroptosisis a form of apoptosis that is accompanied bythe release of the fever-inducing cytokine IL-1. Ferroptosisis a distinct form of cell death that is triggered when excessive intracellular levels of iron or reactive oxygen species overwhelm the glutathione-dependent antioxidant defenses to cause unchecked membrane lipid peroxidation. Entosis (or cannibalism), described in 2007, can be defined as the cell-to-cell internalization, thereby forming a “cell-in-cell” structure or signet ring structure that appears as a “bird’s eye”.  OTHER CELL DEATH TYPES Upon internalization, the entotic cells remain viable for a short period followed by lysosome-mediated degradation and non-apoptotic cell death, while a fraction of the internalized cells can also extricate themselves or are expelled from the host cell. Paraptosis First described by Sperandio et al. in 2000. Paraptosis is a caspace independent programmed cell death characterised by the presence of single layered vacuoles in the mitochondrion. This has been observed in many neurodegenerative diseases. There is no formation of apoptotic bodies.  OTHER CELL DEATH TYPES Parthanatos represents a mitochondrial-linked but caspase-independent cell death and is characterized by the hyperactivation of PARP. PARP mediates the synthesis of poly(ADP-ribose) (PAR), which further shuttles from the nucleus to the cytoplasm and binds to specific mitochondrial proteins followed by apoptosis-inducing factor (AIF) release. No formation of apoptotic bodies.  OTHER CELL DEATH TYPES NETosis Neutrophil extracellular trap-associated cell death (NETosis), a unique form of cell death, is initiated by pathogens or components and mainly occurs in immune cells, particularly neutrophils. The process is promoted through superoxide generated by NADPH oxidase 4 (NOX4). Autosis Autosis was described in 2013 by Beth Levine et al.. Autosis is a non-apoptotic, non-necrotic, autophagic and Na+/K+-ATPase dependent cell death.  OTHER CELL DEATH TYPES Methuosis This is a form of programmed cell death characterised by the displacement of the cytoplasm by large fluid filled vacuoles derived from macropinosomes. There is loss of metabolic capacity and membrane integrity like in necrosis but there is no shrinkage of the cell and fragmentation of the nucleus which occur in apoptosis.  OTHER CELL DEATH TYPES Cuproptosis This was first described and the term was initially defined by Tsvetkov, P. et al. in 2019. Cuproptosis is a form of programmed cell death triggered by copper (Cu). Cuproptosis has been implicated in several pathological conditions, such as Wilson’s disease.  OTHER CELL DEATH TYPES Oxeiptosis The term “oxeiptosis” was coined by Holze et al. in 2018 to describe a caspase-independent, ROS- sensitive, and noninflammatory cell death pathway that protects against inflammation induced by ROS or ROS-generating agents, such as viral pathogens.  OTHER CELL DEATH TYPES Erebosis In 2022, Sa Kan Yoo et al. reported a novel form of cell death called erebosis during the natural turnover of gut enterocytes. The term “erebosis” is derived from the Greek word “erebos,” meaning complete darkness. Erebotic cells undergo membrane blebbing and actin cytoskeleton changes, eventually leading to cell disintegration. This process is marked by the loss of cell adhesion, organelles, and fluorescence emitted from labeled proteins and the accumulation of angiotensin- converting enzyme (ACE). Timeline of the discovery of cell death. This timeline depicts the important discoveries and advancements in cell death research, including the recognition of multiple forms of cell death.