cell cycle and apoptosis

programmed cell death is a

necessary biological function

programmed death plays a fundamental role in

animal development and tissue homeostasis

apoptosis

a form of programmed cell death and is a key aspect of cellular homeostasis

in cancer

the processes and signals that promote apoptosis are inhibited allowing cancer cells to survive and proliferate in a dysregulated manner

apoptosis plays a major role from

embryonic development to senescence

1800s

numerous observations of cell death

1908

mechnikov wins nobel prize (phagocytosis)

1930-40

studies of metamorphosis

1948-49

cell death in chick lamb and exploration of NGF

1955

beginning of studies of lysomes

1964-66

necrosis & PCD described

1971

term apoptosis coined

1977

cell death genes in C. elegans

1980-82

DNA ladder observed & ced-3 identified

1989-91

apoptosis genes identified, including bcl-2, fas/apo1 & p53, ced-3 sequenced

apoptosis - evolutionary conserved

occurs in all multicellular animals studies (plants too)

stages and genes conserved from nematodes (worms) and flies to mice and humans

cells are balanced between life and death

damage and physiological death signals → death signal (proapoptotic and antiapoptotic proteins)→ death

two types of cell death

necrosis and apoptosis

necrosis - type of cell death

results from damage by an external agent (infection or injury)

causes rupture of cells and leakage of their contents into surrounding tissues

as a result, the non-specific immune response occurs and leads to inflammation

loss of functional tissue

impaired organ function, transient or permanent

apoptosis - type of cell death

a continuous, controlled process

in adult apoptosis maintains cell numbers, balancing the production of new cells by mitosis and the death of older cells

cells are destroyed in a way that does not cause leakage of their contents into surrounding tissues

so the non-specific immune response does not occur, and there is no inflammation

removal of damaged or unnecessary cells

necrosis

cellular swelling

membranes are broken

ATP is depleted

cell lyses, eliciting an inflammatory reaction

DNA fragmentation is random or smeared

in vivo whole areas of the tissue are affected

apoptosis

cellular condensation

membrane remain intact

requires ATP

cell is phagocytosed no tissue reaction

ladder like DNA fragmentation

in vivo individual cells appear affected

why do we need apoptosis?

the balance between cell proliferation and cell death is linked to human health

2 examples of why we need apoptosis

cancerous cells have mutations that stop them from reacting to chemical signals that normally start apoptosis as a result they over proliferate

in neurodegenerative diseases (alzheimer’s disease) apoptosis causes the progressive loss of neurons

programmed death occurs in

several different areas of the body

which cells have a mutation in their regulation of apoptosis

cancer cells (become immortal)

t or f apoptosis is needed for proper development

true

apoptosis is needed to destroy what cells

cells infected with viruses

cells of the immune system

cells with DNA damage

cancer cells

examples of how apoptosis is needed for proper development

resorption of the tadpole tail

formation of the fingers and toes of the fetus

sloughing off of the inner lining of the uterus

formation of the proper connections between neurons in the brain

an example of apoptosis

life cycle of frog

apoptosis is invovled in

the growth, homeostasis, and maintenance of healthy cells

a deeper understanding of cell death pathways in cancer is essential for

the development of precisely targeted treatments or synergistic treatment combinations

the cell death mechanism is involved during

arrangement of cells during morphogenesis

regulation of cell number during homeostasis

elimination of abnormal cells during pathogenesis

embryological development

in the fifth week, our hands and feet develop as paddle-like structures

ridges develop in these paddle like structures which become our fingers and toes

apoptosis causes the death of cells between these ridges allowing our individual fingers and toes to separate

in childhood

between 8 and 14 the average child loses between 20 to 30 billion cells each day as a result of apoptosis

in adulthood

the average adult loses between 50 - 70 billion cells each day as a result of apoptosis

morphogenesis

eliminates excess cells

selection

eliminates non-functional cells

immunity

eliminates dangerous cells

organ size

eliminates excess cells

tissue remodeling

eliminates cells no longer needed - lactation

steroid immunisuppressants

kill lymphocytes by apoptosis

lymphocytes are

posied to die by apoptosis

maintains organ size and function

cells lost by apoptosis are replaced by cell division

remember limited replicative potential of normal cells restricts how many times this can occur before tissue renewal declines

apoptosis in pathologic conditions

DNA damage due to radiation, chemotherapy

accumulation of misfolded proteins leads to ER stress which ends with apoptosis

cell death in viral infections that induce apoptosis such as HIV and Adenovirus or by the host immune response such as hepatitis

organ atrophy after duct obstruction

too much apoptosis

tissue atrophy (neurodegeneration, thin skin)

too little apoptosis

hyperplasia (cancer, atherosclerosis)

apoptosis in neurodegeneration (parkinsons, alzheimers, huntington disease)

neurons are post-mitotic (cannot replace themselves; neuronal stem cell replacement is inefficient)

neuronal death is caused by loss of proper connections, loss of proper growth factors, and/or damage (especially oxidative damage)

neuronal dysfunction or damage results in loss of synapses or loss of cell bodies (synaptosis, can be reversible; apoptosis, irreversible)

apoptosis in aging

both too much and too little apoptosis

too much → tissue degeneration

too little → dysfunctional cells accumulate hyperplasia (precancerous lesions)

apoptosis in cancer

apoptosis eliminates damaged cells

tumor suppressor p53 controls senescence and apoptosis responses to damage

most cancer cells are defective in apoptotic response (damaged, mutant cells survive)

high levels of anti-apoptotic proteins or low levels of pro-apoptotic proteins → cancer

t or f apoptosis can be triggered by a range of internal and external circumstances

true

stages of classic apoptosis

healthy cell → death signal (extrinsic or intrinsic) → commitment to die (reversible) → execution (irreversible) → dead cell (condensed, cross-linked) → engulfment (macrophages, neighboring cells) → degradation

cell death by injury

mechanical damage

exposure to toxic chemicals

cell death by suicide

internal signals and external signals

what makes a cell decide to commit suicide?

withdrawal of positive signals

receipt of negative signals

withdrawal of positive signals

growth factors of neurons

interleukin-2 (IL-2)

receipt of negative signals

increased levels of oxidants within the cell

damage to DNA by oxidants

death activators:

tumor necrosis factor alpha (TNF-a)

lymphotoxin (TNF-b)

fas ligand (FasL)

the extracellular signal molecules

mitogens, growth factors, survival factors

mitogens

which stimulate cell division, primarily by relieving intracellular negative controls that otherwise block progress through the cell cycle

growth factors

which stimulate cell growth (an increase in cell mass) by promoting the synthesis of proteins and other macromolecules and by inhibiting their degradation

survival factors

which promote cell survival by suppressing apoptosis

execution phase

the intrinsic and extrinsic pathways converge to a caspase activation cascade

caspases are conserved across species

synthesized as inactive precursors activated by proteolytic cleavage

family of at least 12 proteases a few of which are involved in inflammation and many of which are involved in apoptosis

removal of dead cells

dying cells secrete factors the recruit phagocytes

this facilitates prompt clearance before they undergo secondary necrosis

dead cells disappear without a trace and do not produce inflammation

mechanisms of apoptosis

triggered by cell-death signals

death receptor (extrinsic) pathway

mitochondrial (intrinsic) pathway

execution phase

removal of dead cells

this is carefully controlled by a set of enzymes called caspases

apoptosis: control

physiological receptor pathway & intrinsic damage pathway → mitochondrial signals → caspase cleavage cascade → orderly cleavage of proteins and DNA → crosslinking of cell corpses; engulfment

major players in apoptosis

caspases

adaptor proteins

TNF & TNFR family

Bcl-2 family

mitochondrial (intrinsic) pathway

cell injury

growth factor withdrawal

DNA damage (by radiation, toxins, free radicals)

protein misfolding (ER stress)

death receptor (extrinsic) pathway

receptor-ligandinteractions - Fas & TNF receptor

extrinsic pathway

death ligands → death receptors → initiator caspase 8 → effector caspase 3 → apoptosis

intrinsitc pathway

DNA damage & p53 → mitochondria/cytochrome C → initiator caspase 9 → effector caspase 3 → apoptosis

extrinsic pathway of apoptosis

signals which originate from outside of the cell

can be triggered by an NK cell that has detected abnormal behavior

these ligands bind to a surface-receptor, causing a conformational change inside of the cell. this acts as a signal transduction and activates a protein cascade that produces caspases

what is the extrinsic pathway triggered by

external stimuli or ligand molecules such as TRAIL

death receptors are members of the tumor necrosis factor receptor family and related protein called Fas(CD95)

these molecules contain a death domain

if activated, apoptosis is initiated

extrinsic pathway is mediated by the death receptors such as

TNFR1, CD95, FasR, APO-1, DR4 and Dr5

in cancer cells, TRAIL resistance may be caused by?

various genetic mutations (resulting in altered apoptotic signaling proteins) or over expression of anti-apoptotic Bcl-2 proteins

ligand-induced cell death

FasL - Fas (CD95), TNF - TNF-R, TRAIL-DR$ (Trail-R)

explain the signaling and control pathway of apoptosis

1. apoptotic signals can be externally or internally driven

   1. internal signals involve p53 and the mitochondria

   2. mitochondira release cytochrome c

   3. external signals activate initiator enzyme activators

2. initiator caspases: 6, 8, 9, 12

3. execution caspases: 2, 3, 7

4. apoptosis events: execution caspases trigger the events that cause programmed cell death

How is apoptosis regulated

bcl-2: inhibits apoptosis by blocking cytochrome C-mediated signaling

inhibitors: block apoptotic signals, activators of initiator enzymes, initiator caspases and apoptosis events

survival factors: external and internal signals that promote inhibitors and help prevent apoptosis

overall: these mechanisms regulate or prevent the caspase cascade, stopping programmed cell death

intrinsic pathway of apoptosis

signals which originate from inside of the cell

can be triggered by DNA damage or the presence of an abnormal protein like p53

this activates a caspase cascade

cells may also initiate apoptosis if certain factors required for growth are not present

explain the intrinsic pathway of apoptosis

activation: usually activated by cytochrome C

location: cytochrome c is found between the inner and outer mitochondrial membranes

cell stress: makes mitochondrial membranes more permeable, allowing cytochrome c to leak into the cytosol

cause of stress, radiation, heat, nutrient deprivation, viral infection, increased calcium ion concentration in cytoplasm, glucocorticoids binding to receptors on the nuclear envelope

final step: once in cytosol, cytochrome c activates procaspases into caspases initiating apoptosis

essence of intrinsic pathway

pro-apoptotic and protective molecules that regulate mitochondrial permeability and the release of death molecules sequestered in the mitochondria are maintained in balance normally

imbalance initiates the death pathway

intrinsic pathway (damage):

mitochondria → cytochrome c release → pro-caspase 9 cleavage →pro-execution caspase (3) cleavage → caspase (3) cleavage of cellular proteins, nuclease activation → death

intrinsic pathway is initiated by

apoptogenic triggers or cellular stress

caspases

are a family of proteases

procaspases

synthesised in a cells cytoplasm as inactive

once activated caspase molecules have several effects for example

activate more procaspase to caspase

hydrolyse structural proteins in the cytoplasm (actin)

hydrolyse structural proteins in the nucleus (lamins)

activate dna-hydrolysing enzymes

structural proteins are hydrolysed

the nucleus breaks up

the cell shrinks and forms extensions called blebs

small membrane bound fragments bud off from these blebs and are engulfed by

phagocytic white blood cells

what activates caspase initally - intrinsic pathway

an intercellular signal in response to stress

mitochondria release substances into the cytosol of the cell

one of these is cytochrome c one component of the electron transport chain of aerobic respiration

in the cytosol, cytochrome c causes hydrolysis of inactive procaspase to active caspase

what activates caspase initially - extrinsic pathway

a signal from a different cell often a killer t cell

each cell has so called death receptors on its cell surface membrane

when bound to a complementary protein on the cell-surface membrane of another cell, the death receptor recruits adaptor proteins inside the cytoplasm that attract procaspase molecules and hydrolyse them to active caspase

the caspase cascade involved in apoptosis

proteins preent in the cytoplasm are invovled in the caspase cascade

they start off inactive and through post-translational modification become active

this allows a rapid response even if the cells nucleus is not functioning properly

cleavage of the proteins occurs - subunits are removed and reassembled to form active caspases

an initiator caspase starts apoptosis and activates executioner caspases which degrade DNA keratin and actin. each executioner can degrade over 600 components

how do cancer cells survive and grow by avoiding apoptosis

cancer cells develop defects in apoptosis allowing them to:

survive despite DNA damage and genetic instability

accumulate more aggressive cells

avoid cell death from low oxygen and oxidative stress

survive detached from the extracellular matrix allowing metastasis

how do anti-apoptotic Bcl-2 proteins prevent apoptosis

bcl-2 proteins block the intrinsic apoptosis pathway

pro-apoptotic proteins (BIM, PUMA, tBID) activate Bax/Bak

cancer cells increase anti-apoptotic Bcl-2 proteins to block these signals

this prevents mitochondrial cytochrome c release and apoptosis

how do cancer cells avoid apoptosis by suppressing p53

p53 is a tumor suppressor that promotes apoptosis

p53 activates pro-apoptotic proteins PUMA and NOXA

cancer cells mutate/delete p53 or increase MDM2 (inhibits p53)

without p53 cancer cells survive and continue growing

how do cancer cells change their metabolism to avoid apoptosis

p53 inhibits glycolysis (breakdown of glucose)

low glucose can activate p53 and PUMA promoting apoptosis

cancer genes (RAS, AKT, MYC) increase aerobic glycolysis

this helps cancer cells avoid metabolism-related apoptosis