Stages of cell death,regulators of cell death,apoptosis,types,necrosis,autophagy and its types

1. Cell death,Regulators,
Apoptosis,Necrosis,Autophagy
Presented by Dr.SIBI P ITTIYAVIRAH
PROFESSOR,
DEPARTMENT OF PHARMACEUTICAL SCIENCES,CPAS,CHERUVANDOOR,
ETTUMANOOR,KERALA

2. Cell death
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.
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.

3. Programmed cell death (PCD; sometimes referred to as cellular suicide) is
the death of a cell as a result of events inside of a cell, such as apoptosis or
autophagy.
PCD is carried out in a biological process, which usually confers advantage
during an organism's lifecycle.
For example, the differentiation of fingers and toes in a developing human
embryo occurs because cells between the fingers apoptose; the result is that
the digits are separate.
PCD serves fundamental functions during both plant and animal tissue
development.
Apoptosis and autophagy are both forms of programmed cell death.
]
Necrosis is the death of a cell caused by external factors such as trauma or
infection and occurs in several different forms.

4. Necrosis was long seen as a non-physiological process that occurs as
a result of infection or injury,
but in the 2000s, a form of programmed necrosis, called
necroptosis,was recognized as an alternative form of programmed cell
death.

5. Programmed cell death is the death of a cell as a result of events inside of a cell, such as apoptosis or autophagy. PCD is carried
out in a biological process, which usually confers advantage during an organism's lifecycle.

6. Apoptosis or Type I cell-death.An etoposide-treated DU145 prostate cancer cell exploding into a cascade of apoptotic bodies. The sub images were extracted
from a 61-hour time-lapse microscopy video, created using quantitative phase-contrast microscopy. The optical thickness is color-coded. With increasing thickness,
color changes from gray to yellow, red, purple and finally black.

7. Apoptosis
Apoptosis is the process of programmed cell death (PCD) that may occur in multicellular
organisms.
Biochemical events lead to characteristic cell changes (morphology) and death.
These changes include
blebbing,
cell shrinkage,
nuclear fragmentation,
chromatin condensation,
and chromosomal DNA fragmentation.

8. In cell biology, a bleb is a bulge of the plasma membrane of a cell, human bioparticulate or
abscess with an internal environment synonymous to that of a simple cell, characterized by a
spherical, bulky morphology.
It is characterized by the decoupling of the cytoskeleton from the plasma membrane,
degrading the internal structure of the cell, allowing the flexibility required for the cell to
separate into individual bulges or pockets of the intercellular matrix.
During apoptosis, blebbing is the first phase (left) of cell disassembly.
[1

9. HeLa cells stained for nuclear DNA with the blue fluorescent Hoechst dye. The central and rightmost cell are in interphase,
thus their entire nuclei are labeled. On the left, a cell is going through mitosis and its DNA has condensed.

12. Activation Mechanisms
The initiation of apoptosis is tightly regulated by activation mechanisms, because once apoptosis has
begun, it inevitably leads to the death of the cell.
The two best-understood activation mechanisms are
the intrinsic pathway (also called the mitochondrial pathway)
and the extrinsic pathway.
The intrinsic pathway is activated by intracellular signals generated when cells are stressed and
depends on the release of proteins from the intermembrane space of mitochondria.
The extrinsic pathway is activated by extracellular ligands binding to cell-surface death receptors,
which leads to the formation of the death-inducing signaling complex (DISC).[18]

13. Signaling pathway of TNF-R1. Dashed grey lines represent multiple steps

14. A cell initiates intracellular apoptotic signaling in response to a stress,which may bring about cell suicide.
The binding of nuclear receptors by glucocorticoids,
Heat,
Radiation,
nutrient deprivation,
viral infection,
Hypoxia
increased intracellular concentration of free fatty acids
and increased intracellular calcium concentration,
for example, by damage to the membrane,
can all trigger the release of intracellular apoptotic signals by a damaged cell.
A number of cellular components, such as poly ADP ribose polymerase, may also help regulate apoptosis.[24]

15. Before the actual process of cell death is precipitated by enzymes, apoptotic signals
must cause regulatory proteins to initiate the apoptosis pathway.
This step allows those signals to cause cell death, or the process to be stopped,
should the cell no longer need to die.
Several proteins are involved, but two main methods of regulation have been
identified:
the targeting of mitochondria functionality,[27]
or directly transducing the signal via adaptor proteins to the apoptotic mechanisms.
An extrinsic pathway for initiation identified in several toxin studies is an increase in
calcium concentration within a cell caused by drug activity, which also can cause
apoptosis via a calcium binding protease calpain.

18. Intrinsic pathway
The intrinsic pathway is also known as the mitochondrial pathway.
Mitochondria are essential to multicellular life.
Without them, a cell ceases to respire aerobically and quickly dies.
This fact forms the basis for some apoptotic pathways.
Apoptotic proteins that target mitochondria affect them in different ways. They may cause
mitochondrial swelling through the formation of membrane pores, or they may increase the
permeability of the mitochondrial membrane and cause apoptotic effectors to leak out.
Nitric oxide has been implicated in initiating and inhibiting apoptosis through its possible
action as a signal molecule of subsequent pathways that activate apoptosis.[31

19. During apoptosis, cytochrome c is released from mitochondria through the actions of
the proteins Bax and Bak.
Once cytochrome c is released it binds with Apoptotic protease activating factor – 1
(Apaf-1) and ATP, which then bind to pro-caspase-9 to create a protein complex
known as an apoptosome.
The apoptosome cleaves the pro-caspase to its active form of caspase-9, which in
turn cleaves and activates pro-caspase into the effector caspase-3.

20. Mitochondria also release proteins known as SMACs (second mitochondria-derived
activator of caspases) into the cell's cytosol following the increase in permeability of the
mitochondria membranes.
SMAC binds to proteins that inhibit apoptosis (IAPs) thereby deactivating them, and
preventing the IAPs from arresting the process and therefore allowing apoptosis to
proceed.
IAP also normally suppresses the activity of a group of cysteine proteases called
caspases,which carry out the degradation of the cell.
Therefore, the actual degradation enzymes can be seen to be indirectly regulated by
mitochondrial permeability.

21. Extrinsic Pathway
Two theories of the direct initiation of apoptotic mechanisms in mammals have been suggested: the
TNF-induced (tumor necrosis factor) model and
the Fas-Fas ligand-mediated model,
both involving receptors of the TNF receptor (TNFR) familycoupled to extrinsic signals.
TNF path
TNF-alpha is a cytokine produced mainly by activated macrophages, and is the major extrinsic mediator of
apoptosis. Most cells in the human body have two receptors for TNF-alpha: TNFR1 and TNFR2.
The binding of TNF-alpha to TNFR1 has been shown to initiate the pathway that leads to caspase activation
via the intermediate membrane proteins TNF receptor-associated death domain (TRADD) and Fas-associated
death domain protein (FADD).

22. Binding of this receptor can also indirectly lead to the activation of transcription factors involved
in cell survival and inflammatory responses
The link between TNF-alpha and apoptosis shows why an abnormal production of TNF-alpha
plays a fundamental role in several human diseases, especially in autoimmune diseases.
The TNF-alpha receptor superfamily also includes death receptors (DRs), such as DR4 and
DR5. These receptors bind to the proteinTRAIL and mediate apoptosis.
Apoptosis is known to be one of the primary mechanisms of targeted cancer therapy.

23. Fas path
The fas receptor (First apoptosis signal) – (also known as Apo-1 or CD95) is a
transmembrane protein of the TNF family which binds the Fas ligand (FasL).
The interaction between Fas and FasL results in the formation of the death-inducing
signaling complex (DISC), which contains the FADD, caspase-8 and caspase-10.
In other types of cells (type II), the Fas-DISC starts a feedback loop that spirals into
increasing release of proapoptotic factors from mitochondria and the amplified activation
of caspase-8.

24. Caspases
Caspases play the central role in the transduction of ER apoptotic signals. Caspases are proteins that are
highly conserved, cysteine-dependent aspartate-specific proteases.
There are two types of caspases: initiator caspases, caspase 2,8,9,10,11,12, and effector caspases, caspase
3,6,7. The activation of initiator caspases requires binding to specific oligomeric activator protein.
Effector caspases are then activated by these active initiator caspases through proteolytic cleavage. The
active effector caspases then proteolytically degrade a host of intracellular proteins to carry out the cell death
program.
Caspase-independent apoptotic pathway
There also exists a caspase-independent apoptotic pathway that is mediated by AIF (apoptosis-inducing
factor).[42]

25. Negative regulators of apoptosis
Negative regulation of apoptosis inhibits cell death signaling pathways, helping tumors to evade
cell death and developing drug resistance.
The ratio between anti-apoptotic (Bcl-2) and pro-apoptotic (Bax) proteins determines whether a
cell lives or dies.[47][48]
Many families of proteins act as negative regulators categorized into either antiapoptotic factors,
such as IAPs and Bcl-2 proteins or prosurvival factors like cFLIP, BNIP3, FADD, Akt, and NF-
κB.[49]

26. Autophagy is generally activated by conditions of nutrient deprivation
but has also been associated with physiological as well as pathological
processes such as development, differentiation, neurodegenerative
diseases, stress, infection and cancer.
● Autophagic or Type II cell-death.
(Cytoplasmic: characterized by the formation
of large vacuoles that eat away organelles in a
specific sequence prior to the destruction of the
nucleus.)

27. Diagram of the process of autophagy, which produces the structures autophagosomes, AP, and autolysosomes, AL; (B) Electron micrograph of
autophagic structures AP and AL in the fat body of a fruit fly larva; (C) Fluorescently labeled autophagosomes AP in liver cells of starved mice.

28. Macroautophagy, often referred to as autophagy, is a catabolic process that
results in the autophagosomic-lysosomal degradation of bulk cytoplasmic
contents, abnormal protein aggregates, and excess or damaged organelles.
Microautophagy, on the other hand, involves the direct engulfment of cytoplasmic
material into the lysosome.This occurs by invagination, meaning the inward folding of
the lysosomal membrane, or cellular protrusion.

29. Chaperone-mediated autophagy, or CMA, is a very complex and specific pathway, which
involves the recognition by the hsc70-containing complex.
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.

31. Mitophagy is the selective degradation of mitochondria by autophagy. It often
occurs to defective mitochondria following damage or stress.
Mitophagy promotes the turnover of mitochondria and prevents the accumulation
of dysfunctional mitochondria which can lead to cellular degeneration.
It is mediated by Atg32 (in yeast) and NIX and its regulator BNIP3 in mammals.
Mitophagy is regulated by PINK1 and parkin proteins.
The occurrence of mitophagy is not limited to the damaged mitochondria but also
involves undamaged ones.[34]

32. Lipophagy is the degradation of lipids by autophagy,[36]
a function which has been
shown to exist in both animal and fungal cells.
The role of lipophagy in plant cells, however, remains elusive.
In lipophagy the target are lipid structures called lipid droplets (LDs), spheric
"organelles" with a core of mainly triacylglycerols (TAGs) and a unilayer of
phospholipids and membrane proteins. In animal cells the main lipophagic pathway is
via the engulfment of LDs by the phagophore, macroautophagy.
In fungal cells on the other hand microplipophagy constitutes the main pathway and is
especially well studied in the budding yeast Saccharomyces cerevisiae
Lipophagy was first discovered in mice and published 2009.[

33. Functions[edit]
Nutrient starvation[edit]
Autophagy has roles in various cellular functions. One particular example is in yeasts, where the nutrient
starvation induces a high level of autophagy. This allows unneeded proteins to be degraded and the amino
acids recycled for the synthesis of proteins that are essential for survival.
In higher eukaryotes, autophagy is induced in response to the nutrient depletion that occurs in animals at birth
after severing off the trans-placental food supply, as well as that of nutrient starved cultured cells and tissues.[
In microbiology, xenophagy is the autophagic degradation of infectious particles. Cellular autophagic machinery
also play an important role in innate immunity. Intracellular pathogens, such as Mycobacterium tuberculosis (the
bacterium which is responsible for tuberculosis) are targeted for degradation by the same cellular machinery
and regulatory mechanisms that target host mitochondria for degradation.

34. Infection
Vesicular stomatitis virus is believed to be taken up by the autophagosome from the cytosol and
translocated to the endosomes where detection takes place by a pattern recognition receptor called toll-like
receptor 7, detecting single stranded RNA.
Following activation of the toll-like receptor, intracellular signaling cascades are initiated, leading to
induction of interferon and other antiviral cytokines.
]
Galectin-8 has recently been identified as an intracellular "danger receptor", able to initiate autophagy
against intracellular pathogens. When galectin-8 binds to a damaged vacuole, it recruits an autophagy
adaptor such as NDP52 leading to the formation of an autophagosome and bacterial degradation.[84]
Repair mechanisms
Autophagy degrades damaged organelles, cell membranes and proteins, and insufficient autophagy is
thought to be one of the main reasons for the accumulation of damaged cells and aging.[85]
Autophagy and
autophagy regulators are involved in response to lysosomal damage, often directed by galectins such as
galectin-3 and galectin-8. These in turn recruit receptors such as TRIM16[86]
and NDP52[84]
and directly
affect mTOR and AMPK activity, whereas mTOR and AMPK inhibit and activate autophagy, respectively.[87]

35. Necrosis (from Ancient Greek νέκρωσις, nékrōsis, "death") is a form of cell
injury which results in the premature death of cells in living tissue by autolysis.[
1]
Necrosis is caused by factors external to the cell or tissue, such as infection,
or trauma which result in the unregulated digestion of cell components.
In contrast, apoptosis is a naturally occurring programmed and targeted cause
of cellular death.
While apoptosis often provides beneficial effects to the organism, necrosis is
almost always detrimental and can be fatal.[2]

36. Cellular death due to necrosis does not follow the apoptotic signal transduction pathway, but
rather various receptors are activated and result in the loss of cell membrane integrity[3]
and an
uncontrolled release of products of cell death into the extracellular space.
This initiates in the surrounding tissue an inflammatory response, which attracts leukocytes and
nearby phagocytes which eliminate the dead cells by phagocytosis.
However, microbial damaging substances released by leukocytes would create collateral
damage to surrounding tissues.
This excess collateral damage inhibits the healing process.
Thus, untreated necrosis results in a build-up of decomposing dead tissue and cell debris at or
near the site of the cell death.
A classic example is gangrene. For this reason, it is often necessary to remove necrotic tissue
surgically, a procedure known as debridement.

37. Morphological patterns[edit]
There are six distinctive morphological patterns of necrosis:[6]
Coagulative necrosis is characterized by the formation of a gelatinous (gel-like) substance
in dead tissues in which the architecture of the tissue is maintained,[6]
and can be
observed by light microscopy.
Coagulation occurs as a result of protein denaturation, causing albumin to transform into a
firm and opaque state.
This pattern of necrosis is typically seen in hypoxic (low-oxygen) environments, such as
infarction.
Coagulative necrosis occurs primarily in tissues such as the kidney, heart and adrenal
glands.
Severe ischemia most commonly causes necrosis of this form.

38. Liquefactive necrosis (or colliquative necrosis), in contrast to coagulative necrosis, is
characterized by the digestion of dead cells to form a viscous liquid mass.
This is typical of bacterial, or sometimes fungal, infections because of their ability to
stimulate an inflammatory response.
The necrotic liquid mass is frequently creamy yellow due to the presence of dead
leukocytes and is commonly known as pus.
Hypoxic infarcts in the brain presents as this type of necrosis, because the brain
contains little connective tissue but high amounts of digestive enzymes and lipids,
and cells therefore can be readily digested by their own enzymes.

39. Gangrenous necrosis can be considered a type of coagulative necrosis that resembles
mummified tissue. It is characteristic of ischemia of lower limb and the gastrointestinal
tracts. If superimposed infection of dead tissues occurs, then liquefactive necrosis
ensues (wet gangrene).[8]
Caseous necrosis can be considered a combination of coagulative and liquefactive
necrosis,[5]
typically caused by mycobacteria (e.g. tuberculosis), fungi and some
foreign substances. The necrotic tissue appears as white and friable, like clumped
cheese. Dead cells disintegrate but are not completely digested, leaving granular
particles.[5]
Microscopic examination shows amorphous granular debris enclosed within
a distinctive inflammatory border.[6]
Some granulomas contain this pattern of
necrosis.[9]

40. Fat necrosis is specialized necrosis of fat tissue,[9]
resulting from the action of activated lipases
on fatty tissues such as the pancreas. In the pancreas it leads to acute pancreatitis, a condition
where the pancreatic enzymes leak out into the peritoneal cavity, and liquefy the membrane by
splitting the triglyceride esters into fatty acids through fat saponification.[6]
Calcium, magnesium
or sodium may bind to these lesions to produce a chalky-white substance.
Fibrinoid necrosis is a special form of necrosis usually caused by immune-mediated vascular
damage. It is marked by complexes of antigen and antibodies, referred to as immune complexes
deposited within arterial walls[6]
together with fibrin.[6]

41. Causes
Necrosis may occur due to external or internal factors.
External factors[edit]
External factors may involve mechanical trauma (physical damage to the body which causes
cellular breakdown), damage to blood vessels (which may disrupt blood supply to associated
tissue), and ischemia.[12]
Thermal effects (extremely high or low temperature) can result in
necrosis due to the disruption of cells.
In frostbite, crystals form, increasing the pressure of remaining tissue and fluid causing the
cells to burst.[12]
Under extreme conditions tissues and cells die through an unregulated
process of destruction of membranes and cytosol.[13]

42. Internal factors[edit]
Internal factors causing necrosis include: trophoneurotic disorders (diseases that occur due to defective nerve
action in a part of an organ which results in failure of nutrition); injury and paralysis of nerve cells. Pancreatic
enzymes (lipases) are the major cause of fat necrosis.[12]
Necrosis can be activated by components of the immune system, such as the complement system; bacterial
toxins; activated natural killer cells; and peritoneal macrophages.[1]
Pathogen-induced necrosis programs in cells
with immunological barriers (intestinal mucosa) may alleviate invasion of pathogens through surfaces affected by
inflammation.[1]
Toxins and pathogens may cause necrosis; toxins such as snake venoms may inhibit enzymes
and cause cell death.[12]
Necrotic wounds have also resulted from the stings of Vespa mandarinia.[14]
Pathological conditions are characterized by inadequate secretion of cytokines. Nitric oxide (NO) and reactive
oxygen species (ROS) are also accompanied by intense necrotic death of cells.[12]
A classic example of a
necrotic condition is ischemia which leads to a drastic depletion of oxygen, glucose, and other trophic factors
and induces massive necrotic death of endothelial cells and non-proliferating cells of surrounding tissues
(neurons, cardiomyocytes, renal cells, etc.).[1]
Recent cytological data indicates that necrotic death occurs not
only during pathological events but it is also a component of some physiological process.[12]
Activation-induced death of primary T lymphocytes and other important constituents of the immune response are
caspase-independent and necrotic by morphology; hence, current researchers have demonstrated that necrotic
cell death can occur not only during pathological processes, but also during normal processes such as tissue
renewal, embryogenesis, and immune response.[12]