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Review
Department of Medicine, Centre of Excellence for Arthritis and Rheumatology, Lousiana State Uni6ersity Health Sciences Center, Shre6eport,
LA 71130 -3932, USA
b
Department of Pediatrics, Lousiana State Uni6ersity Health Sciences Center, 1501 Kings Highway, Shre6eport, LA71130 -3932, USA
Received 9 March 2000; accepted 15 May 2000
Abstract
Apoptosis or programmed cell death, is essential for the normal functioning and survival of most multi-cellular organisms. The
morphological and biochemical characteristics of apoptosis, however, are highly conserved during the evolution. It is currently
believed that apoptosis can be divided into at least three functionally distinct phases, i.e. induction, effector and execution phase.
Recent studies have demonstrated that reactive oxygen species (ROS) and the resulting oxidative stress play a pivotal role in
apoptosis. Antioxidants and thiol reductants, such as N-acetylcysteine, and overexpression of manganese superoxide (MnSOD)
can block or delay apoptosis. Bcl-2, an endogenously produced protein, has been shown to prevent cells from dying of apoptosis
apparently by an antioxidative mechanism. Taken together ROS, and the resulting cellular redox change, can be part of signal
transduction pathway during apoptosis. It is now established that mitochondria play a prominent role in apoptosis. During
mitochondrial dysfunction, several essential players of apoptosis, including pro-caspases, cytochrome C, apoptosis-inducing factor
(AIF), and apoptotic protease-activating factor-1 (APAF-1) are released into the cytosol. The multimeric complex formation of
cytochrome C, APAF-1 and caspase 9 activates downstream caspases leading to apoptotic cell death. All the three functional
phases of apoptosis are under the influence of regulatory controls. Thus, increasing evidences provide support that oxidative stress
and apoptosis are closely linked physiological phenomena and are implicated in pathophysiology of some of the chronic diseases
including AIDS, autoimmunity, cancer, diabetes mellitus, Alzheimers and Parkinsons and ischemia of heart and brain. 2000
Elsevier Science Ireland Ltd. All rights reserved.
Keywords: Reactive oxygen species; Apoptosis-inducing factor; Apoptotic protease-activating factor-1
1. Introduction
Apoptosis, or programmed cell death (PCD), is a
naturally occurring cell death process, essential for the
normal development and homeostasis of all multicellular organisms [1]. This process is also important for
removing damaged, infected, or potentially neoplastic
cells. However, both too little and too much apoptotic
cell death can lead to adverse biological consequences
[24]. Rheumatoid arthritis and cancer are the best examples for too little apoptosis. Ischemic heart disease,
AIDS and neurodegenerative diseases such as
* Corresponding author. Tel.: +1-318-6756086; fax: + 1-3186756059.
E-mail address: sjain@lsuhsc.edu (S.K. Jain).
0928-4680/00/$ - see front matter 2000 Elsevier Science Ireland Ltd. All rights reserved.
PII: S 0 9 2 8 - 4 6 8 0 ( 0 0 ) 0 0 0 5 3 - 5
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2. Mediators of apoptosis
Most of the mediators of apoptosis can be broadly
placed in one of five categories based on the primary
perturbation within the cell, i.e. (1) the cell surface, (2)
the cytosol, (3) the cytoskeleton, (4) the mitochondrion,
(5) and the nucleus. At the cell surface, some of the
well-characterized death receptors are Fas (CD95), tumor necrosis factor receptor-1 (TNF-Rl), CAR1, DR3,
DR4, and DR5 [2429]. The ligands that activate these
death receptors are structurally related molecules that
belong to the tumor necrosis factor (TNF) gene superfamily [1,30]. Interaction of death receptors with ligands may lead to the initiation of a death signal,
depending upon the presence of intracellular death
domains and its physical association with the adapter
protein called Fas-associated death domain (FADD)
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Table 2
Diseases that are influenced by oxidative stress or apoptosis
Neurodegenerative diseases (Alzheimers, Parkinsons)
Autoimmune disease (e.g. rheumatoid arthritis)
Human immunodeficiency virus (AIDS)
Diabetes mellitus
Cancers of lung, colon, breast and others
Alcohol induced liver disease
Hepatitis-C induced liver disease
Ischemic reperfusion damage-heart, liver
tegrity and function [62 72]. The lipid radicals generated during the early encounter with an oxidant add
molecular oxygen to produce lipid dioxyl radical. This
pro-oxidant abstracts, an allylic hydrogen from another
unsaturated side chain, producing a lipid hydroperoxide
(LOOH) and thus propagating the chain [62 64]. Iron,
a transition metal, is also well known for its crucial role
in the initiation of new lipid-radical chain reactions
[65].
Lipid peroxidation is an important biological consequence of oxidative cellular damage and aging [6673].
A partial list of conditions or diseases that are likely to
involve oxidative stress is given in Table 2. This includes several neurodegenerative diseases and AIDS.
Other sources of ROS include radiation (e.g. UV), toxic
chemicals (e.g. paraquat and endosulfan) and
chemotherapeutic agents (e.g. adriamycin and
bleomycin) [13,74,75]. The classical view of ROS as
villains that indiscriminately destroy biological macromolecules has undergone a shift, in which, positive
physiological roles are considered as well. In summary,
oxidative stress can have positive responses, such as,
proliferation or activation, as well as negative responses, such as, lipid peroxidation, DNA damage, cell
growth inhibition or cell death.
157
not. This resulted in a dramatic shutdown of mitochondrial protein biosynthesis. Transcription factors such as
NF-kB, p53 and AP-1 are also sensitive to redox
changes in mammalian cells, primarily through redox
regulation of their DNA binding regions [13,84,88,97].
Cell death in the nematode Caenorhabditis elegans is
undoubtedly the best system for the study the genetics
of programmed cell death. During development, a process resembling apoptosis deletes 131 cells out of 1090
cells of this nematode. A total of 14 suicide genes that
are important for development have been identified so
far, of which three genes Ced3, Ced4 and Ced9
have received most attention. The Ced9 gene negatively
regulates the Ced3 and Ced4 genes [1,35,9799]. The
homologues of Ced9 in humans have been identified as
Bcl-2, which protect mammalian cells. Caspase 3
(YAMA/CPP32), a proteolytic enzyme in humans is
believed to be an equivalent of Ced3 in nematode. The
homologue to Ced4 in human has been identified to be
APAF-1 (apoptotic protease-activating factor-1) [35].
Other genes of importance are c-fos, c-jun as well as
c-myc, p53, clusterin, RP-2 and RP-8 [100,101]. However, the complex mechanisms controlling gene expression that determines cell survival and cell death are still
unresolved.
158
159
Fig. 1. Schematic model of mammalian cell death pathway. Death signal may be delivered at the cell surface by direct ligand-receptor interaction.
This followed by clustering of death receptors and activation of caspase-8. Alternatively, cytotoxic drugs, ionizing radiation may directly activate
caspase-9, otherwise a later event. Mitochondria plays a central role in apoptosis by releasing apoptogenic factors and proteases into the cytosol.
A major checkpoint in this pathway is the ratio between pro-apoptotic (Bax) to anti-apoptotic (Bcl-2) members. Miitochondrial dysfunction
includes PT, change in mitochondrial membrane potential, production of ROS, release of cytochrome C, AIPF-1 and caspases-2, -3, and -9 into
the cytosol, where they form multimeric complexes. This activates downstream caspases and degradation of death substrates in the nucleus, which
ultimately leads to cell death.
160
efficient means of amplifying the apoptotic signal transduced by caspase 8 even under very low concentration.
In the absence of mitochondria, high concentrations of
caspase 8 were required to activate downstream caspase
cascade. Interestingly, caspase 8 can activate the mitochondria-dependent pathway even when the Bcl-2
protein is present. This helps explain the failure of Bcl-2
to inhibit CD95-dependent apoptosis consistently [117].
Scaffidi et al., in 1998 [118] have identified two types of
CD95-mediated cell death. Recently, it has been
demonstrated that Apaf-1 / and caspase 9 / T
cells remain sensitive to Fas-induced killing [119,120].
However, Fas-induced apoptosis was markedly reduced
in embryonic fibroblasts with Apaf-1 / phenotype
[121], suggesting Fas (CD95) can activate different
pathways in different cell types. These observations in
T cells indicate that the apoptotic function of mitochondria (at least the cytochrome C release part) can be
bypassed in these cells.
Other less defined pathways that lead to apoptosis
are the ones initiated by potent cytotoxic chemicals or
gamma irradiation. These pathways seem to rely on the
apoptotic function of mitochondria, since cells with
Apaf-1 / and caspase-9 / phenotypes are principally resistant to these death-inducing agents [119
121]. Further research in this area may help us to better
understand some of the missing links.
9. Conclusions
The role of oxidative stress as a mediator of apoptotic cell death in diverse cell systems is now better
understood. In recent years, mitochondria have gained
considerable importance both as a site for ROS production and as a major player in apoptosis. The involvement of mitochondria or ROS in apoptosis is not
without controversy. In some experimental systems,
apoptosis can occur at very low oxygen tension, when
ROS are unlikely to occur. The protooncogene
product, Bcl-2, can inhibit apoptosis both in the presence and in the absence of reactive oxygen products.
Further research is needed to clarify the underlying
mechanisms.
Recent developments suggest that mtDNA may be a
good indicator of oxidative damage in apoptosis. It is
becoming apparent that the redox status of a cell can
have a complex and multilayered effect on cell survival
and cell death. Future investigations will unravel
whether oxidative stress is a pre-requisite for all apoptotic events. This would be an attractive mechanism
whereby a panoply of seemingly diverse injuries could
rapidly converge on common apoptotic pathways.
More in-depth studies are needed to elucidate the role
of BH3-domain-only subset of molecules and their interplay with ROS. This could open new sites of drug
targeting. Future studies may also improve our understanding whether the oxidation states of mitochondrial
thiols alone function as a prominent apoptotic sensor.
A better understanding of the molecular machinery of
apoptosis in the pathophysiology will undoubtedly
provide novel therapeutic interventions and care for a
large number of patients with chronic diseases.
Acknowledgements
We thank Georgia Morgn First for the critical reading of the manuscript. This work was supported in part
by the funds by Feist Weiller Cancer Center and the
Center of Excellence for Arthritis and Rheumatology of
the Louisiana State University Health Sciences Center
at Shreveport, and a grant-in aid from the American
Heart Association (Southeast).
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