Cellular responses to stress (Adaptations, injury and death) (4 of 5) Ali Al Khader, MD Faculty of Medicine Al-Balqa’ Applied University Email: ali.alkhader@bau.edu.jo

**Apoptosis** The cell here decides to die by activating enzymes that degrade its own DNA and proteins The name means: “falling off” because the fragments of the cell appear falling off Cell membrane is intact but it is altered that the cell and its fragments appear good targets for phagocytosis The dead cells and its fragments are cleared rapidly before contents leaked out…inflammation doesn’t occur

Apoptosis Necrosis Membranes are intact but they are altered and are avid for phagocytosis Membrane integrity is lost The cell is shrunken The cell is swollen Dead cells and fragments are cleared rapidly before inflammation can occur Contents are leaked out inducing surrounding inflammation Can be physiologic…mention 2 examples Always pathological (always preceded by irreversible injury)

Apoptosis in physiologic situations Embryogenesis Involution of hormone-dependent tissues upon hormone deprivation …mention 2 examples Cell loss in proliferating cell populations, such as intestinal crypt epithelia…tissue renewal Elimination of cells that have served their useful purpose…like neutrophils after end of inflammation (because there are no signals now) Elimination of self-reactive lymphocytes

Apoptosis in pathologic conditions DNA damage: -radiation -cytotoxic anticancer drugs -extremes of temperatures -…etc …even causes of necrosis can also cause apoptosis especially when the main damage is to the DNA or proteins Accumulation of misfolded proteins: due to -gene mutations (DNA damage) or -direct misfolding of proteins such as by free radicals …these misfolded proteins accumulate in the ER causing ER stress which culminates in apoptosis

Apoptosis in pathologic conditions, cont’d Certain infections, esp., viruses: -apoptosis is induced by the virus…examples: adenovirus and HIV or -apoptosis is induced by the immune response…cytotoxic T cells kill virus-infected cells…example: hepatitis viruses ….cytotoxic T cells also kill neoplastic cells Death of parenchymal cells due to obstruction of organ ducts…e.g., pancreas, parotid, and kidney

Morphology of apoptosis Nucleus: chromatin condensation fragmentation (karyorrhexis) into nucleosome-sized pieces Cell shrinkage Formation of cytoplasmic buds these buds fragment into apoptotic bodies …these fragments are rapidly phagocytosed so: no inflammation and the whole process may be invisible on H&E

Apoptosis in normal colonic mucosa

Mechanisms of apoptosis 2 pathways: 1- The Mitochondrial (Intrinsic) Pathway of Apoptosis…most situations 2- The Death Receptor (Extrinsic) Pathway of Apoptosis Both pathways activate caspases that are the main players here they are cysteine proteases that cleave proteins after aspartic residues

= BH3 proteins = Caspase 9 in the mitochondrial pathway = CD95 Synthesized when there are survival signals (like growth factors) = CD95 Also inhibit antiapoptotic proteins (Bcl-2 and Bcl-XL) These receptors have death domain in their cytoplasmic part = Caspase 9 in the mitochondrial pathway Some also inhibit caspase antagonists = BH3 proteins = Pro-apoptotc proteins..they dimerize and form channel in mit. membrane..so cytochrome c and others escape to cytosol

Its ligand is called: Fas ligand…present on activated T lymphocytes: for killing self-reactive lymphocytes and killing target cells by cytotoxic T cells Caspase 8 in this pathway These activate caspase 8 after they bound to the death domain of the receptor that bound to the ligand May activate pro-apoptotic member of BcL-2 family called Bid which activates mitochondrial pathway *Caspase antagonist called FLIP block activation of caspases in this pathway…some viruses produce homologues to FLIP to keep the infected cell alive May also secrete granzyme which directly activates caspases (not mitochondrial or death-receptor pathway)

Caspases The executioner caspases when activated activate nucleases degrade nuclear matrix and cytoskeleton fragmentation Degrade DNA and nucleoproteins

Clearance of apoptotic cells Apoptotic cells produce “eat me” signals Phosphatidyl serine is normally present on the inner leaflet of plasma membrane…it becomes on the outer leaflet in apoptotic cell…recognized by macrophages Also secreted factors by dying cells induce phagocytosis Some apoptotic bodies express adhesive glycoproteins that are recognized by macrophages

Necrosis and apoptosis may coexist DNA damage (seen in apoptosis) activates an enzyme called poly- ADP(ribose) polymerase, which depletes cellular supplies of nicotinamide adenine dinucleotide, leading to a fall in ATP levels and ultimately necrosis Even in common situations such as ischemia, it has been suggested that early cell death can be partly attributed to apoptosis, with necrosis supervening later as ischemia worsens Apoptosis induced by some pathologic stimuli may progress to necrosis…like some stimuli of DNA damage

Examples of Apoptosis Growth Factor Deprivation: -hormone-sensitive cells deprived of the hormone, -lymphocytes that are not stimulated by antigens and cytokines -neurons deprived of nerve growth factor *In all these situations, apoptosis is triggered by the mitochondrial pathway: -activation of pro-apoptotic members of the Bcl-2 family -decreased synthesis of Bcl-2 and Bcl-xL

Examples of Apoptosis, cont’d DNA Damage: When DNA is damaged, the p53 protein arrests the cell cycle (at the G1 phase). Why? …if the damage is too great to be repaired successfully, p53 triggers apoptosis (mitochondrial pathway) …When p53 is mutated or absent, cells with damaged DNA survive, and in this case What may happen?

Examples of Apoptosis, cont’d Accumulation of Misfolded Proteins: -First: unfolded protein response increased synthesis of chaperones in ER (which control proper folding) -Then: if the accumulation is more severe ER stress activates mitochondrial pathway **Cell death by this mechanism is a feature of many neurodegenerative diseases: -Alzheimer -Huntington -Parkinson diseases -..etc