α-Synuclein and neuronal cell death

Mutations in the DJ-1 gene have been linked to autosomal recessive familial Parkinson's disease. To understand the function of DJ-1, we determined the DJ-1 expression in both zebrafish and post mortem human brains. We found that DJ-1 was expressed early during zebrafish development and throughout adulthood. Knock down (KD) of DJ-1 by injection of morpholino did not cause dramatic morphologic alterations during development, and no loss of dopaminergic neurons was observed in embryos lacking DJ-1. However, DJ-1 KD embryos were more susceptible to programmed cell death. While a slight reduction in staining for islet-1 positive neurons was observed in both DJ-1 KD and H₂O₂ treated embryos, the number of apoptotic cells was significantly increased in both KD and H₂O₂ treated embryos. Interestingly, DJ-1 expression was increased in brains of zebrafish under conditions of oxidative stress, indicating that DJ-1 is a part of stress-responsive machinery. Since oxidative stress is one of the major contributors to the development of Alzheimer's disease (AD), we also examined DJ-1 expression in AD brains. Using DJ-1 specific antibodies, we failed to detect a robust staining of DJ-1 in brain tissues from control subjects. However, DJ-1 immunoreactivity was detected in hippocampal pyramidal neurons and astrocytes of AD brains. Therefore, our results strongly suggest that DJ-1 expression is not necessary during zebrafish development but can be induced in zebrafish exposed to oxidative stress and is present in human AD brains.     

Apoptosis or programmed cell death?

Although there are different ways in which cells may die, it is now thought that in a developmental context cells are induced to positively commit suicide whilst in a homeostatic context the absence of certain survival factors may provide the impetus for suicide. There appears to be some variation in the morphology and indeed the biochemistry of these suicide pathways; some treading the path of "apoptosis", others following a more generalized pathway to deletion, but both usually being genetically and synthetically motivated. There is some evidence that certain symptoms of "apoptosis" such as endonuclease activation can be spuriously induced without engaging a genetic cascade, however, presumably true apoptosis and programmed cell death must be genetically mediated. It is also becoming clear that mitosis and apoptosis are toggled or linked in some way and that the balance achieved depends on signals received from appropriate growth or survival factors.     

Giant cell formation: the way to cell death or cell survival?

The study of giant cells in populations of different tumor cells and evaluation of their role in cancer development is an expanding field. The formation of giant cells has been shown to be followed by mitotic catastrophe, apoptosis, necrosis, and other types of cell elimination. Reports also demonstrate that giant cells can escape cell death and give rise to new cancer cells. However, it is not known if the programmed cell death is involved in this type of cell cycle disorders. Here we describe principal events that are observed during giant cell formation. We also consider the role of giant cells in cancer development, taking into account both published work and our own recent data in this field.     

Does autophagy contribute to cell death?

Autophagy (specifically macroautophagy) is an evolutionarily conserved catabolic process where the cytoplasmic contents of a cell are sequestered within double membrane vacuoles, called autophagosomes, and subsequently delivered to the lysosome for degradation. Autophagy can function as a survival mechanism in starving cells. At the same time, extensive autophagy is commonly observed in dying cells, leading to its classification as an alternative form of programmed cell death. The functional contribution of autophagy to cell death has been a subject of great controversy. However, several recent loss-of-function studies of autophagy (atg) genes have begun to address the roles of autophagy in both cell death and survival. Here, we review the emerging evidence in favor of and against autophagic cell death, discuss the possible roles that autophagic degradation might play in dying cells, and identify salient issues for future investigation.     

The end of autophagic cell death?

In the mammalian system, cell death is often preceded or accompanied by autophagic vacuolization, a finding that initially led to the widespread belief that so-called "autophagic cell death" would be mediated by autophagy. Thanks to the availability of genetic tools to disable the autophagic machinery, it has become clear over recent years that autophagy usually constitutes a futile attempt of dying cells to adapt to lethal stress rather than a mechanism to execute a cell death program. Recently, we systematically addressed the question as to whether established or prospective anticancer agents may induce "autophagic cell death". Although a considerable portion among the 1,400 compounds that we evaluated induced autophagic puncta and actually increased autophagic flux, not a single one turned out to kill tumor cells through the induction of autophagy. Thus, knockdown of essential autophagy genes (such as ATG5 and ATG7) failed to prevent and rather accelerated chemotherapy-induced cell death, in spite of the fact that this manipulation efficiently inhibits autophagosome formation. Herein, we review these finding and--polemically--raise doubts as to the very existence of "autophagic cell death".     

Activation—induced cell death in B lymphocytes

Upon encountering the antigen(Ag),the immune system can either develop a specific immune response of enter a specific state of unresponsiveness,tolerance.The response of B cells to their specific Ag can be activation and proliferation,leading to the immune response,or anergy and activation-induced cell death(AICD),leading to tolerance.AICD in B lymphocytes is a highly regulated event initiated by crosslinking of the B cell receptor (BCR).BCR engagement initiates several signaling events such as activation of PLCγ,Ras,and PI3K,which generally speaking,lead to survival.However,in the absence of survival signals(CD40 or IL-4R engagement),BCR crosslinking can also promote apoptotic signal transduction pathways such as activation of effector caspases,expression of pro-apoptotic genes,and inhibition of pro-survival genes.The complex interplay between survival and death signals determines the B cell fate and, consequently,the immune response.     

A second signal for autophagic cell death?

Dictyostelium cells in monolayers in vitro lend themselves well to a study of autophagic cell death (ACD). There is no apoptosis machinery in the protist Dictyostelium, no caspase nor Bcl-2 family members (except a paracaspase whose inactivation does not alter cell death), thus there is no apoptosis that could interfere with, or substitute for, non-apoptotic cell death. Also, Dictyostelium, a eukaryote, has a haploid genome, which facilitates random insertional mutagenesis.     

Do inducers of apoptosis trigger caspase-independent cell death?

Apoptotic cell death is mediated by molecular pathways that culminate in the activation of a family of cysteine proteases, known as the caspases, which orchestrate the dismantling and clearance of the dying cell. However, mounting evidence indicates that a cell that has been treated with an apoptotic inducer can also initiate a suicide programme that does not rely on caspase activation. Here, we present recent findings and discuss the physiological relevance of caspase-independent cell death.     

Ammonium ion transport—a cause of cell death

Ammonium can be transported into the cell by ion pumps in the cytoplasmic membrane. Ammonia then diffuse out through the cell membrane. A futile cycle is created that results in cytoplasmic acidification and extracellular alkalinisation. Ammonium transport can be quantified by measuring the extracellular pH changes occurring in a cell suspension (in PBS) after addition of ammonium. By using this technique, in combination with specific inhibitors of various ion pumps, it was shown that ammonium ions are transported across the cytoplasmic membrane by the Na⁺K⁺2Cl^--cotransporter in both hybridoma and myeloma cells. Further, the Na⁺/H⁺ exchanger, which regulates intracellular pH by pumping out protons, was shown to be active during ammonium exposure. The viability of hybridoma cells suspended in PBS and exposed to NH _inf4 ^sup+ for only 90 min, was reduced by 11% (50% necrosis and 50% apoptosis). A control cell suspension did not loose viability during this time. Turning off the activity of the Na⁺/H⁺ exchanger (by amiloride) during ammonium exposure decreased viability further, while inhibiting transport itself (by bumetanide) restored viability to the same level as for the control experiment with bumetanide alone. These results show that one effect of ammonia/ammonium on cell physiology is specifically related to the inward transport of ammonium ions by membrane bound ion pumps.Abbreviations q _pH specific rate of pH increase (pH units per min and 10⁶ cells per ml)     

Cell death in Leishmania induced by stress and differentiation: programmed cell death or necrosis?

Unicellular organisms, such as the protozoan parasite Leishmania, can be stimulated to show some morphological and biochemical features characteristic of mammalian apoptosis. This study demonstrates that under a variety of stress conditions such as serum deprivation, heat shock and nitric oxide, cell death can be induced leading to genomic DNA fragmentation into oligonucleosomes. DNA fragmentation was observed, without induction, in the infectious stages of the parasite, and correlated with the presence of internucleosomal nuclease activity, visualisation of 45 to 59 kDa nucleases and detection of TUNEL-positive nuclei. DNA fragmentation was not dependent on active effector downstream caspases nor on the lysosomal cathepsin L-like enzymes CPA and CPB. These data are consistent with the presence of a caspase-independent cell death mechanism in Leishmania, induced by stress and differentiation that differs significantly from metazoa.     

The extrinsic cell death pathway and the élan mortel

Early in the exploration of the chemical nature of life, it was widely believed that the molecules of living organisms, by their very nature, differ from those of inorganic material molecules and possess a vital force ('élan vital'). Similarly, early scientific thinking on the subject of cell death and its induction by cytotoxic cells of the immune system was pervaded by a sense that the molecules mediating these functions possess intrinsic deadly activity and are dedicated exclusively to death-related tasks. This impression was also reflected in the initial notions of the mode of action of intracellular proteins that signal for death. It is now gradually becoming clear, however, that proteins participating in death induction also have functions unrelated to death. Nevertheless, as exemplified by studies of the function of caspase-8 (an enzyme that signals both for activation of the extrinsic cell-death pathway and for non-death-related effects), analysis of the mechanistic basis for such heterogeneity might allow identification of distinct structural determinants in the proteins participating in death induction that do bear death specificity.Cell Death and Differentiation (2008) 15, 1533-1541; doi:10.1038/cdd.2008.41.     

The apoptosis endonucleases: cleaning up after cell death?

The term apoptosis describes the predictable structural changes associated with many forms of programmed cell death. One of the first visible events of apoptosis is the collapse of the nucleus. Nuclear degradation is manifested by digestion of chromatin into nucleosome-sized fragments or multiples of these. This digestion of DNA is enzymatic, and several attempts have been made to characterize apoptosis-specific endodeoxyribonucleases. Although there are strong candidates for such enzymes, direct evidence for their role in apoptosis is yet to be provided.     

Necrosis: a specific form of programmed cell death?

For a long time necrosis was considered as an alternative to programmed cell death, apoptosis. Indeed, necrosis has distinct morphological features and it is accompanied by rapid permeabilization of plasma membrane. However, recent data indicate that, in contrast to necrosis caused by very extreme conditions, there are many examples when this form of cell death may be a normal physiological and regulated (programmed) event. Various stimuli (e.g., cytokines, ischemia, heat, irradiation, pathogens) can cause both apoptosis and necrosis in the same cell population. Furthermore, signaling pathways, such as death receptors, kinase cascades, and mitochondria, participate in both processes, and by modulating these pathways, it is possible to switch between apoptosis and necrosis. Moreover, antiapoptotic mechanisms (e.g., Bcl-2/Bcl-x proteins, heat shock proteins) are equally effective in protection against apoptosis and necrosis. Therefore, necrosis, along with apoptosis, appears to be a specific form of execution phase of programmed cell death, and there are several examples of necrosis during embryogenesis, a normal tissue renewal, and immune response. However, the consequences of necrotic and apoptotic cell death for a whole organism are quite different. In the case of necrosis, cytosolic constituents that spill into extracellular space through damaged plasma membrane may provoke inflammatory response; during apoptosis these products are safely isolated by membranes and then are consumed by macrophages. The inflammatory response caused by necrosis, however, may have obvious adaptive significance (i.e., emergence of a strong immune response) under some pathological conditions (such as cancer and infection). On the other hand, disturbance of a fine balance between necrosis and apoptosis may be a key element in development of some diseases.     

Senescence and programmed cell death: substance or semantics?

The terms senescence and programmed cell death (PCD) have led to some confusion. Senescence as visibly observed in, for example, leaf yellowing and petal wilting, has often been taken to be synonymous with the programmed death of the constituent cells. PCD also obviously refers to cells, which show a programme leading to their death. Some scientists noted that leaf yellowing, if it has not gone too far, can be reversed. They suggested calling leaf yellowing, before the point of no return, 'senescence' and the process after it 'PCD'. However, this runs into several problems. It is counter to the historical definitions of senescence, both in animal and plant science, which stipulate that senescence is programmed and directly ends in death. It would also mean that only leaves and shoots show senescence, whereas several other plant parts, where reversal has not (yet) been shown, have no senescence, but only PCD. This conflicts with ordinary usage (as in root and flower senescence). Moreover, a programme can be reversible and therefore it is not counter to logic to regard the cell death programme as potentially reversible. In green leaf cells a decision to die, in a programmed way, has been taken, in principle, before the cells start to remobilize their contents (that is, before visible yellowing) and only rarely is this decision reversed. According to the arguments developed here there are no good reasons to separate a senescence phase and a subsequent PCD phase. Rather, it is asserted, senescence in cells is the same as PCD and the two are fully synchronous.