An adult human is made up of about 30 trillion (3 10 13) cells, all

of which originate from a single fertilized egg. If this first cell divides
into 2, the progeny cells into 4, and so on, it would take only about 45
rounds of division to produce the number of cells required to make an
adult human. In fact, cell division occurs constantly through our
lifetimes, such that we generate a new complete set of 3 10 13 cells
every 2 weeks. The reason that multicellular organisms do not become
infinitely large is because the proliferation of cells is balanced by cell
death.

Development of the Cell

Death Concept
*Necrosis was the earliest form of Cell Death known to man and was considered to be the only one
until Virchows landmark article titled, Cellular Pathology talked about involvement of cells in
Infecction.
* Necrosis was mentioned in Ancient Greek texts since Galens time. ( AD-129 )
*Three main avenues leading to cell death have been delineated: necrosis, apoptosis and
autophagy.
*Necrosis was defined as an accidental form of cell death caused by a hostile environment to which
a
cell could not adapt effectively. It was thus seen as a passive process in which the cell was more
sinned
against than sinning itself.
*By contrast, apoptosis is a form of cellular suicide in which the cell participates actively in its own
demise. It is a mechanism by which individual cells activate their own signaling systems to sacrifice
themselves for the preservation of the organism.
*Autophagy (see above) is also an active signaling process that is elicited when a stressful
environment
requires autodigestion of a portion of the cells macromolecular constituents.

*Necrosis is a form of pathological cell death.

*Unplanned murder of cells by external agents.
*Necrosis was defined as an accidental form of cell death caused by a hostile environment to
which a cell could not
adapt effectively.
*It was thus seen as a passive process in which the cell was more sinned against than sinning
itself.
*Typically affects geographically localized groups of cells. The response to this process is
usually acute inflammation,
which itself may generate further cell injury.

Necrosis

In cells that are injured, ATP concentrations fall so low that the Na+/K+ ATPase can no longer
operate, and therefore
ion concentrations are no longer controlled. This causes the cells to swell and then burst. The
cell contents then leak
out, causing the surrounding tissues to become inflamed. Cells that die by suicide on the
other hand shrink, and their
cell contents are packaged into small membrane-bound packets called blebs. The nuclear
DNA becomes chopped up
into small fragments, each of which becomes enclosed in a portion of the nuclear envelope.
The dying cell modifies its
plasma membrane, signaling to macrophages, which respond by engulfing the blebs and the
remaining cell fragments .
*Three morphologic changes follow:

Coagulative necrosis.
A.Normal heart. All myocytes are nucleated, and
striations are clear.
B. Myocardial infarction. The heart from a patient
Following acute myocardial infarction. The necrotic cells
are deeply eosinophilic and most have lost their nuclei.

APOPTOSIS
BARE BEGINNINGS
*Spontaneous cell death as a physiological event was discussed almost as
soon as stains became available. It was born with a bang in 1885 in a paper
by the same Walther Flemming who created the terms chromatin and
mitosis. Flemming studied ovarian follicles in mammals and noticed that
the epithelial lining of regressing follicles was littered with cells the nuclei of
which were breaking up.
*Sydney Brenner's studies on animal development began in the late1950s in what was to become the Laboratory of Molecular Biology
(LMB) in Cambridge, UK. It was at this lab that during the 1970s and
1980s, a team led by John Sulston succeeded in tracing the nematode
Caenorhabditis elegans entire embryonic cell lineage. In other words,
Sulston and his team had traced where each and every cell in the
roundworm's embryo came from during the division process, and where
it ended up.

Apoptosis as observed in 1885 by

Flemming, who called it chromatolysis.
Top left: Normal rabbit ovarian follicle
near matunity, 1 mm in diameter.
Numerous epithelial mitoses. Top right:
Early stage of involution in a
nearbyfollicle. Many epithelial cells are in
vanous stages of death by
chromatolysis; some are shed into the
lumen. Bottom: detail of the same
involuting follicle. Most epithelial cells in
contact with the ovum are normal (one is
in mitosis); those farther removed are
undergoing chromatolysis. Note the halfmoons of chromatin typical of apoptosis.

Apoptosis as seen in 1886 by a German

medical student, Franz Nissen, in the
lactating mammary gland. Nissen became
aware of Flemming's study,2' after having
completed his own, and concluded that the
name chromatolysis was very suitable also
to his own findings.

Apoptosis illustrated in 1914 by L. Graper

as chromatolysis. Wall of the yolk sac in
the course of involution, in a 20-cm
embryo of Acanthias, a creature that we
have been unable to identify. An epithelial
cell has taken up the fragmented nucleus
of a neighboring cell that died during the
involution process.

Apoptosis actually
Whats in a name?
The term, Apoptosis was coined in 1972 by Kerr, Wyllie and Currie in
their landmark article introducing the concept of Programmed cell death
titled, APOPTOSIS: A BASIC BIOLOGICAL PHENOMENON WITH
WIDE- RANGING IMPLICATIONS IN TISSUE KINETICS.
We are most grateful to Professor James Cormack of the Department of Greek,
University of Aberdeen, for suggesting this term. The word " apoptosis " is used
in Greek to describe "the dropping off " or " falling off " of petals from flowers,
or leaves from trees. To show the derivation clearly, we propose that the stress
should be on the penultimate syllable, the second half of the word being
pronounced like " ptosis " (with the " p " silent), which comes from the same
root " to fall and is already used to describe drooping of the upper eyelid.
A general mechanism of controlled cell deletion, which is complementary
to mitosis in the regulation of animal cell populations.

The two main regulators of cell populations,

mitosis (bottom left) and apoptosis (top right). In
the apoptotic cell, the nucleus is fragmented
(karyorhexis) and the chromatin is pyknotic. One
of the nuclear fragments contains a
characteristic half-moon of condensed
chromatin. (Electron micrograph from a rat
prostate 2 days after castration. Bar = 2 Y).

Apoptosis Produces
Individual Cell Death Amidst
Viable Cells

*Apoptosis is a pattern of cell death that is triggered by a variety of

extracellular and
intracellular stimuli and is carried to its conclusion by organized
cellular signaling cascades.
*Apoptotic cells are recognized by nuclear fragmentation and pyknosis,
generally
Against background of viable cells. Importantly, apoptosis occurs
in single cells or small groups of cells, whereas necrosis
characteristically involves larger geographic areas of cell death.
.

Stages

The earliest phase is the stimulus that provokes the apoptotic response. This may be
an external signal delivered through surface receptors or may originate inside the cell
from the action of a drug, toxin, or radiation.
The next phase includes detection of this signal or metabolic state and transduction of
the signal. Signal transduction pathways send this message to the cell death effector
machinery.
The effector phase is the third part of the cell death mechanism and includes the
proteases that are activated during apoptosis, as well as their positive and negative
regulators.
The fourth phase of cell death is the postmortem phase, in which the cell's chromatin
condenses and its DNA is degraded. In vivo (but not necessarily in vitro) dying cells are
recognized and engulfed by other cells.

When, where and why?

*Programmed cell death rids the developing
animal of superfluous or unwanted cells.
*For example, large numbers of self-reactive lymphocytes or
lymphocytes that fail to produce useful antigen-specific
receptors are deleted by programmed cell death.
*In the developing nervous system, about half of the neurons
produced undergo programmed cell death soon after they
mature if they fail to establish vital connections with target.
*Cells dangerous to the organism, such as transformed
cells, damaged cells, or those infected with pathogens such
as viruses, are also eliminated by the activation
of programmed cell death.
*In case of DNA Damage, PCD may eliminate cells carrying
potentially harmful mutations, including mutations in cells
that can lead to development of Cancer.

Removal of Apoptotic Cells

Once the self-destructive process of apoptosis has propelled a cell to DNA fragmentation
and
cytoskeletal dissolution, the final phase, the apoptotic body, remains. Apoptotic
bodies are
phagocytosed by tissue macrophages.
Phosphatidylserine (PS), a phospholipid that is normally on the interior aspect of the cell
membrane, is
externalized in cells undergoing apoptosis. PS is recognized by macrophages and
activates ingestion of an
apoptotic cells mortal remains without release of intracellular constituents, thus
avoiding an inflammatory
reaction.
Mononuclear phagocytes ingest the debris from apoptotic cells, but recruitment of
neutrophils or lymphocytes
is rare. This situation is unlike that of cells that undergo necrotic cell death, which tends
to elicit acute
inflammatory responses.

Molecular mechanism
*Insight into the molecular basis of apoptosis was first revealed in studies on the
nematode
worm C. elegans, whose cells can be followed with absolute precision during
embryonic
development.
*Of the 1090 cells produced during the development of this worm,131 cells are
normally
destined to die by apoptosis yielding 959 somatic cells seen in adult worm .
(Sulston and Horvitz, 1977; Sulston et al., 1983).
* Nearly all dying cells undergo the same sequence
(Sulston and Horvitz, 1977; Sulston et al., 1983).

of morphological changes .

*A number of mutations affecting the process of cell death have been isolated.
These mutations,
which have defined the genes ced-7 (for cell death abnormal), ced-2, and nut-7
(nuclease
deficient) (Sulston, 1976; Hedgecock et al., 1983), affect all programmed cell deaths.
Thus, all cell deaths
appear to involve the same genetic (and, hence, molecular) processes.

Crucial Findings
*In 1986,Robert Horvitz and his colleagues at the Massachusetts
Institute of
Technology discovered that worms carrying a mutation in the CED-3
gene proceed
through development without losing any of their cells to apoptosis.
This finding
suggested that the product of the CED-3 gene played a crucial role in
the process of
apoptosis in this organism.
*Two genes, CED-3 and CED-4, were identified as genes required for
cell death:
Inactivating mutations in either of these two genes prevents all
developmentally
occurring cell deaths in C. elegans. This observation also provided
the first direct
evidence that cells die by an intrinsic suicide program.
*CED-3 is a protease, whereas CED-4 is a protein that activates CED3 by interacting
with it in dying cells. Another gene, CED-9,acts to protect cells from

*Complicating the issue, CED-9 is also expressed in many of the 131

cells
that are fated to die during development. How then do these cells die?
Death of these cells requires the expression of another gene, EGL-1.
EGL-1 antagonizes
CED-9 by physically interacting with it, thus freeing CED-4 to activate
CED-3.
Genetic studies have shown that the over expression of EGL-1 or
mutations that
hyper activate EGL-1 (gain of function mutation) can induce ectopic cell
death in C.
elegans, which can be suppressed by gain-of-function mutation of CED9 or loss of
function mutations of CED-3 or CED-4.
*In contrast, a loss of function mutation of EGL-1 cannot suppress cell
deaths caused by
the loss of CED-9 activity (loss of function CED-9). These results are
consistent with
EGL-1 acting upstream of the CED-3, CED-4, and CED-9 genes in the

*The identification of key components of the genetic program in the

nematode led to
the search for their homologs in mammals. These studies have
resulted in the
identification of a family of proteins, collectively called caspases, as
the mammalian
homologs of CED-3.
*The homolog of CED-4 in mammals is a protein called Apaf-1,
whereas proteins
related in sequence to CED-9 comprise the Bcl-2 family. In contrast
with the situation
in C. elegans, however, where CED-9 is antiapoptotic, the Bcl-2 family
of proteins is composed
of both antiapoptotic and proapoptotic members. Some of these
proapoptotic Bcl-2
proteins function as the mammalian counterparts of EGL-1.
*Consistent with the evolutionary conservation of the mechanisms
underlying
apoptosis, the overexpression of the nematode proteins CED-3 or

Isolation of Cell Death Mutants by Direct Screening

Programmed cell deaths can easily be identified in living nematode

using Nomarski
differential interference contrast microscopy (Sulston and Horvitz,
1977). The first sign
of the impending death of a cell is a slight increase in its refractility.
*The nucleus of the dying cell becomes increasingly refractile until it
resembles a flat
button; this stage persists for lo-30 min.
*Subsequently, the nucleus of the dying cell decreases in refractility,
begins to appear
crumpled, and then gradually disappears. This process is completed in
less than 1 hr
.(Sulston and Horvitz, 1977;Sulston et al., 1983).

Ced-9 mutant (below)

Absence of Cell Deaths in ced-3 Animals

Ghgm;;

References
*Cell and Molecular Biology- Gerard Karp.
*Apoptosis, Oncosis, and NecrosisGuido Majno and Isabelle Joris From the Department of Pathology, University of
Massachusetts Medical School, Worcester,
Massachusetts. American Journal of Pathology, Vol. 146, No. 1, January 1995 Copyright )
American Society for Investigative
Pathology.

*Apoptosis: A basic biological phenomenon with wide-ranging

implications in tissue
Kinetics.
J. F. R. KERR*, A. H. WYLLIE AND A. R. CURRIE. Br. J. Cancer (1972) 26, 239

*Genetic Control of Programmed Cell Death in the Nematode C.

elegans.
Hilary M. E.llis, and H. Robert Horvitz.Cell. Vol 44, 817-829, March 28, 1986, Copyright 0 1986 by
Cell Press.

Apoptosis - .mkv