Cell Damage and Cell Death Flashcards
What are the causes and mechanisms of cell death?
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GENETIC:
- Abnormal chromosomes (deletions/translocatioNs)
- Increased fragility (Fanconi’s anaemia)
- Failure of repair (Xeroderma pigmentosa)
- Inborn errors (Storage disorders ie. Tay Sachs disease)
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INFLAMMATION:
- Trauma
- Thrombo-embolism
- Atherosclerosis
- Vasculitis
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PHYSICAL:
- Irradiation
- Heat
- Cold
- Barotrauma
- TRAUMATIC DAMAGE:
- Interruption of blood supply
- Direct rupture of cells
- Entry of foreign agents
- INFECTION:
- Toxic agents
- Competition for nutrients
- Intracellular replication (viruses/mycobacteria provoking an immune response)
CHEMICAL:
- Acids/corrosives
- Specific actions e.g. enzymes
- Interference with metabolism e.g. alcohol
What are the three mechanisms of cell death?
- Necrosis
- Apoptosis
- Autophagic Cell Death
Describe necrosis
Most common cause of cell death due to stresses such as ischaemia, trauma and chemical injury
Describe apoptosis
Programmed cell death, eliminating unwanted host cells through activation of a co-ordinated, internally programmed series of events effected by a dedicated set of gene products
Describe autophagic cell death
Process whereby cells engulf parts of themselves
(degradation of normal proteins involved in cellular remodelling found during metamorphis, ageing and differentiation and for digestion and removal of abnormal proteins which would accumulate following toxin exposure, cancer or disease)
Example is breast cancer cell death induced through tamoxifen
What is the cause of necrosis?
Lack of blood supply to cells or tissues
due to
- injury
- infection
- cancer
- infarction
- inflammation
Steps in necrosis
Lack of blood supply = no oxygen
- Energy deprivation causes changes (cells unable to produce ATP due to oxygen deprivation)
- ATP is required for ion pumps to work; electrolyte levels are imbalanced causing water to enter the cell and cause swelling
- Sometimes ion pumps will restore the imbalance and return back to a normal state (reversible)
- However, water influx can cause disintegration and bursting, (haphazard destruction) of organelles and nuclear material (irreversible)
- Lysosomes can release their proteases and lipases which can attack nearby cells
- Cellular debris will stimulate an inflammatory response!!
What is the microscopic appearance of necrosis? (Nuclear Changes)
- Nuclear changes
- Chromatin condensation
- Fragmentation of nucleus
- Dissolution of chromatin by DNAse
What is the microscopic appearance of necrosis? (Cytoplasmic changes)
- Opacification: denaturation of proteins with aggregation
- Complete digestion of cells by enzymes causing cell to liquify (liquefactive necrosis)
What is the microscopic appearance of necrosis (chemical changes)?
- Release of enzymes such as creatine kinase or lactate dehydrogenase
- Release of proteins such as myoglobin
- Biochemical changes are useful in the clinic to measure the extent of tissue damage
What is the main function of necrosis?
What can failure of necrosis lead to?
Removal of damaged cells from an organism, failure of necrotic cell debris may lead to chronic inflammation!!!
What type of cells are selected in apoptosis?
- Selective process for the deletion of superfluous, infected or transformed cells
Provide examples of processes in which apoptosis is involved in
- Embryogenesis
- Metamorphis
- Normal tissue turnover
- Endocrine-dependant tissue atrophy
- Cells of the mammary gland atrophy once they are no longer required
- Variety of pathological conditions
Give some examples of apoptosis
- Cell death in hand to form individual fingers
- DNA damage mediated apoptosis
- DNA damage due to chemo/radiation will cause p53 tumour supressor gene to accumulate. Causes cell cycle arrest allowing for the cell to repair damage
- Cell death in tumours causing regression
- Cell death in viral diseases
- Cell death induced by cytotoxic T-cells
- Death of neutrophils during inflammatory response
- Death of immune cells
- T + B lymphocytes after depletion of cytokines as well as death of autoreactive T cells in developing thymus
What are the two types of apoptosis?
Intrinsic and Extrinsic
Describe intrinsic apoptosis
Apoptosis triggered by intracellular signals
- DNA damage -p53 dependant pathway
- Interruption of the cell cycle
- Inhibition of protein synthesis
- Viral infection
- Change in redox
Describe extrinsic apoptosis
Apoptosis triggered by extracellular signals
- Withdrawal of growth factors
- Extracellular signals e.g TNF
- T cells or Natural Killer e.g granzyme
What is the initiator of apoptosis?
Apoptosis is induced by a family of proteases called CASPASES
What is apoptosis mediated by?
INTRACELLULAR PROTEOLYTIC CASCADE
- Inactive procaspase Y is activated by active caspase X by cleavage N-terminal and C-terminal cleavage sites
- As a result of the cleavage, two parts of the procaspase Y will dimerise forming active caspase Y
- Active caspase Y (initiator caspase 8 or 9) will go on to degrade further substrates
What happens once the active caspase 8 or 9 has been activated?
- The active initiator caspases (8/9) will go to activate other procaspases which will cause cleavage of cytosolic proteins containing cysteine or aspartate
- E.g actin cytosolic skeleton breaks down = cell collapse
- Further caspases will be activated (caspase 1, 3, 6, 7) which will cause amplification of proteolysis
- For example cleavage of nuclear lamin protein essential for nuclear envelope
How can you asses whether cells have been undergone apoptosis or necrosis via gel electrophoresis?
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Apoptotic Cell DNA = clear laddering
- Nucleosomes will remain intact BUT DNAses will cleave around nucleosome = laddering
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Necrotic Cell DNA = no laddering
- DNA fragmentation = random due
- There will be no nucleosomes as they will be denatured due to release of enzymes from lysosomes
- DNA fragmentation = random due
What are the key molecules in the extrinsic apoptotic pathway?
Transmembrane receptor = two domains, extracellular ligand binding domain + intracellular death domain
Death adaptor protein = Two domains death domain and death effector domain. Death domain will form dimers with the death domain of transmembrane receptor
Procaspase 8 = Two domains, a protease domain and death effector domain. Death effector domain can interact with death effector domain on death adaptor protein
How is the initiator caspase in the extrinsic pathway activated?
Tumour necrosis factor-TNF (ligand) induces formation of death-inducing signalling complex (DISC).
- After the interactions between the domains and their close proximity due to the binding of TNF
- Procaspase proteins will undergo autoproteolysis will become active caspase-8 proteins (initiator caspase)
= ACTIVATES CASPASE CASCADE
What are the key molecules in the cytochrome C induced intrinsic apoptosis pathway?
- Cytochrome C (found exclusively in mitochondria)
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APAF-1 protein → 3 domains
- Cytochrome C binding site
- APAF domain
- Caspase recruitment domain (CARD)
- Procaspase-9 has a CARD domain which binds APAF-1 protein
How is the initiator caspase in the intrinsic pathway activated?
- Cytochrome C induces formation of a death inducing complex
- A few cytochrome C molecules will bring together APAF-1 molecules
- These in turn bring together procaspase-9 molecules
- Close proximity of procaspase 9 induces autoproteolysis,
- Activating procaspase-9 → caspase 9
CASPASE CASCADE ACTIVATED
Expand on the expression of anti-apoptotic proteins and pro-apoptotic proteins
- Bcl-2 is part of a multigene family in mammals whcih contain both anti-apoptotic and pro-apoptotic proteins
- Bcl-2 will form dimers (hetero/homo)
- The expression will determine the fate of the cell triggering cell death or promoting life
Where is Bax located?
- Located on the mitochondrial membrane
- Pro-apoptotic protein
What is the function of Bax?
Pro-apoptotic
- Will form homodimers on the mitochondrial membrane
- Pore is formed allowing cytochrome C to be released from the mitochondria → cytosol
- Cytochrome C activates APAF and caspase-9 to initiate apoptosis
Function of bcl-2
Anti-apoptotic protein which prevents normal healthy cells from undergoing apoptosis:
- bcl-2 forms a heterodimer with Bax and blocks pore
- Cytochrome C will not be released from mitochondria
What is the effect of correct survival signals on apoptosis?
If cells are recieving the right survival signals
- Protein kinase B/AKT is activated and phosphorylates Bad (pro-apoptotic) to terminate its actions
Effect is inappropriate survival signals on apoptosis (survival factor withdrawal)
If cells don’t recieve appropriate signals (surival factor withdrawal)
- Protein kinase B (AKT) is not activated
- Bad is therefore in de-phosphorylated form
- Bad will compete with Bcl-2 for binding to Bax complex
- Bad will form a heterodimer with Bcl-2 (anti-apoptotic)
- The block on Bax complex pore is therefore removed = permits release of cytochrome C into cytosol, initiates apoptosis
Effects on intracellular stress (DNA damage) on apoptosis
- During intracellular stress (DNA damage)
- p53 transcription factor activated
- p53 initially activates transcription of cell cycle inhibitor p21 to try and stop cell cycle to repair DNA
- If DNA repair does not occur, p53 will activate expression of pro-apoptotic proteins e.g Bax
- More Bax complexes inserted on mitochondrial membrane
- More pores created permitting release of cytochrome C into cytosol = initiates apoptosis
