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Programmed Cell Death and the Control of Cell Survival
Philosophical Transactions B
Abstract
We draw the following tentative conclusions from our studies on programmed cell death (PCD): (i) the amount of normal cell death in mammalian development is still underestimated; (ii) most mammalian cells constitutively express the proteins required to undergo PCD; (iii) the death programme operates by default when a mammalian cell is deprived of signals from other cells; (iv) many normal cell deaths may occur because cells fail to obtain the extracellular signals they need to suppress the death programme; and (v) neither the nucleus nor mitochondrial respiration is required for PCD (or Bcl-2 protection from PCD), raising the possibility that the death programme, like mitosis, is orchestrated by a cytosolic regulator that acts on multiple organelles in parallel.
... Este proceso se observa de manera regular durante la embriogénesis, así como en el mantenimiento de la homeostasis tisular durante la etapa adulta. La muerte de tipo apoptótica puede presentarse tanto en condiciones patológicas como en el tejido sano, ejemplo de esto es su participación en el desarrollo de los sistemas nervioso (Raff et al., 1993) e inmune (para revisión Williams, 1994), además de presentarse en algunas demencias neurodegenerativas (Loo et al., 1993). A nivel molecular, la compleja cascada de eventos que controlan a la maquinaria apoptótica es dirigida a través de la expresión regulada de proteínas y genes asociados. ...
Durante el envejecimiento hay cambios en el rendimiento físico y sensorial que impulsan a la persona mayor a modificar sus actividades.
El modelo de atención social a la salud permite que las personas mayores conserven y desarrollen su potencial y su bienestar físico, social y emocional, así como su participación social.
... Apoptosis (or programmed cell death) is a physiologic event in response to multiple stimuli including growth factor withdrawal, radiation therapy, and chemotherapeutic agents. [19][20][21][22] Apoptotic cells undergo shrinkage, chromatin condensation, and plasma membrane blebbing with the activation of proteases and endonucleases. Their final phenotype is characterized by plasma membrane-bound ''apoptotic bodies''. ...
Flavopiridol (NSC 649890; Behringwerke L86-8275, Marburg, Germany), is a potent inhibitor of cyclin dependent kinases (CDKs) 1, 2, and 4. It has potent antiproliferative effects in vitro and is active in tumor models in vivo. While surveying the effect of flavopiridol on cell cycle progression in different cell types, we discovered that hematopoietic cell lines, including SUDHL4, SUDHL6 (B-cell lines), Jurkat, and MOLT4 (T-cell lines), and HL60 (myeloid), displayed notable sensitivity to flavopiridol-induced apoptosis. For example, after 100 nmol/L for 12 hours, SUDHL4 cells displayed a similar degree of DNA fragmentation to that shown by the apoptosis-resistant PC3 prostate carcinoma cells only after 3,000 nmol/L for 48 hours. After exposure to 1,000 nmol/L flavopiridol for 12 hours, typical apoptotic morphology was observed in SUDHL4 cells, but not in PC3 prostate carcinoma cells despite comparable potency (SUDHL4:120 nmol/L; PC3: 203 nmol/L) in causing growth inhibition by 50% (IC50). Flavopiridol did not induce topoisomerase I or II cleavable complex activity. A relation of p53, bcl2, or bax protein levels to apoptosis in SUDHL4 was not appreciated. While flavopiridol caused cell cycle arrest with decline in CDK1 activity in PC3 cells, apoptosis of SUDHL4 cells occurred without evidence of cell cycle arrest. These results suggest that antiproliferative activity of flavopiridol (manifest by cell cycle arrest) may be separated in different cell types from a capacity to induce apoptosis. Cells from hematopoietic neoplasms appear in this limited sample to be very susceptible to flavopiridol-induced apoptosis and therefore clinical trials in hematopoietic neoplasms should be of high priority.
... These results strongly suggest that progesterone inhibits the proliferation of cervical cancer cells and switches acidotoxic cell death from necrosis to apoptosis by suppressing TRPM7 expression and activity. Apoptosis is the process by which cells are programmed to die in an orderly and timely manner and also the prerequisite process to promptly remove dying and damaged cells (Wyllie et al., 1980;Raff et al., 1994). Cancer cells, in contrast, escape from the apoptotic cell death by acquiring a dysfunctional apoptotic program (Hacker and Vaux, 1995;Thompson, 1995;Renehan et al., 2001). ...
Because intravaginal pH is strongly acidic, it is important to investigate the effects of acidosis on cervical cancer cells. Recently, in response to strong acidosis, human cervical cancer HeLa cells were shown to exhibit necrosis after showing persistent cell swelling induced by Cl- influx. Since cation influx should be accompanied with Cl- influx to drive water inflow causing cell swelling, we here studied on the nature of acidotoxic cation conductance. The mRNA/protein expression was assessed by RT-PCR and Western blotting. Ionic currents were measured by patch-clamping techniques. Cell counting/viability and colorimetric assays were applied to assess proliferation rate and caspase 3/7 activity, respectively. Cell volume and size were measured by electronic sizing and video-microscopic measurements, respectively. Acid exposure enhanced TRPM7 activity endogenously expressed in HeLa cells and exogenously overexpressed in HEK293T cells. Gene silencing of TRPM7 abolished acid-induced cell swelling and necrosis but rather induced activation of apoptotic caspase 3/7 in HeLa cells. Overexpression with the pore charge-neutralizing D1054A mutant suppressed acid-enhanced cation currents, acid-induced cell swelling, and acidotoxic necrosis in HEK293T cells. Progesterone treatment was surprisingly found to suppress molecular and functional expression of TRPM7 and cell proliferation in HeLa cells. Furthermore, in the progesterone-treated cells, acid exposure did not induce persistent cell swelling followed by necrosis but induced persistent cell shrinkage and apoptotic cell death. These results indicate that in the human cervical cancer cells, TRPM7 is essentially involved in acidotoxic necrotic cell death, and progesterone inhibits TRPM7 expression thereby inhibiting acidotoxic necrosis by switching to apoptosis.
... The balance between cell proliferation and cell death is critical for normal development and homeostasis in multicellular organisms [1][2][3][4], and disruption of this balance leads to serious human diseases, such as cancers and neurodegenerative diseases [1,[5][6][7][8]. Apoptosis, a type of programmed cell death, is mediated by the sequential activation of caspases, a family of cysteine proteases that cleave specifically after aspartic acid residues [9,10]. ...
Abnormal regulation of caspase-2-mediated neuronal cell death causes neurodegenerative diseases and defective brain development. PIDDosome is caspase-2 activating complex composed of PIDD, RAIDD, and caspase-2. Recent whole-exome sequencing study showed that the RAIDD mutations in the death domain (DD), including G128R, F164C, R170C, and R170H mutations, cause thin lissencephaly (TLIS) by reducing caspase-2-mediated neuronal apoptosis. Given that the molecular structure of the RAIDD DD:PIDD DD complex is available, in this study, we analyzed the molecular mechanisms underlying TLIS caused by the RAIDD TLIS variants by performing mutagenesis and biochemical assays.
... Programmed cell death is necessary for normal development and growth in multicellular organisms, which produce billions of apoptotic cells (ACs) daily (1,2). Exogenous pathogenic microbes also threaten organisms' lives and development (3). ...
The swift clearance of apoptotic cells (ACs) (efferocytosis) by phagocytes is a critical event during development of all multicellular organisms. It is achieved through phagocytosis by professional or amateur phagocytes. Failure in this process can lead to the development of inflammatory autoimmune or neurodegenerative diseases. AC clearance has been conserved throughout evolution, although many details in its mechanisms remain to be explored. It has been studied in the context of mammalian macrophages, and in the nematode Caenorhabditis elegans, which lacks “professional” phagocytes such as macrophages, but in which other cell types can engulf apoptotic corpses. In Drosophila melanogaster, ACs are engulfed by macrophages, glial, and epithelial cells. Drosophila macrophages perform similar functions to those of mammalian macrophages. They are professional phagocytes that participate in phagocytosis of ACs and pathogens. Study of AC clearance in Drosophila has identified some key elements, like the receptors Croquemort and Draper, promoting Drosophila as a suitable model to genetically dissect this process. In this review, we survey recent works of AC clearance pathways in Drosophila, and discuss the physiological outcomes and consequences of this process.
... It has been reported that SCT signaling is required for the survival of neuronal progenitors under normal and toxic conditions (Hwang et al., 2009;Jukkola et al., 2011). As a consequence of the early arrival to the IGL, the inability of immature GCs to establish proper synaptic connections with target neurons results in cell death (Raff et al., 1989). In consistent with these studies, excess apoptosis of GCs was found in both EGL and IGL of Sct−/− mice. ...
Postnatal development of the cerebellum is critical for its intact function such as motor coordination and has been implicated in the pathogenesis of psychiatric disorders. We previously reported that deprivation of secretin (SCT) from cerebellar Purkinje neurons impaired motor coordination and motor learning function, while leaving the potential role of SCT in cerebellar development to be determined. SCT and its receptor (SCTR) were constitutively expressed in the postnatal cerebellum in a temporal and cell-specific manner. Using a SCT knockout mouse model, we provided direct evidence showing altered developmental patterns of Purkinje cells (PCs) and granular cells (GCs). SCT deprivation reduced the PC density, impaired the PC dendritic formation, induced accelerated GC migration and potentiated cerebellar apoptosis. Furthermore, our results indicated the involvement of protein kinase A (PKA) and extracellular signal regulated kinase (ERK) signaling pathways in SCT-mediated protective effects against neuronal apoptosis. Results of this study illustrated a novel function of SCT in the postnatal development of cerebellum, emphasizing the necessary role of SCT in cerebellar-related functions.
PIDDosome formation followed by caspase-2 activation is critical for genotoxic stress-induced apoptotic cell death. Failure of proper caspase-2 activation causes a neurodevelopmental disorder and intellectual disability. R815W, R862W, and Q863stop mutations in p53-induced protein with a death domain (PIDD), a component of the PIDDosome, also lead to this disorder. However, the molecular mechanisms underlying this pathogenesis remain elusive. In this study, we analyzed the molecular mechanisms underlying the pathogenesis of the PIDD DD pathogenic variants R815W, R862W, and Q863stop. We determined that these mutations prevented the interaction between PIDD and RIP-associated Ich-1/Ced-3 homologous protein with a death domain (RAIDD), a molecule that mediates PIDDosome formation. The disruption of this interaction affects PIDDosome formation and caspase-2 activation.
All cells are constantly exposed to conflicting environment cues that signal cell survival or cell death. Survival signals are delivered by autocrine or paracrine factors that actively suppress a default death pathway. In addition to survival factor withdrawal, cell death can be triggered by environmental stresses such as heat, UV light, and hyperosmolarity or by dedicated death receptors (e.g., FAS/APO-1 and tumor necrosis factor [TNF] receptors) that are counterparts of growth factor or survival receptors at the cell surface. One of the ways that cells integrate conflicting exogenous stimuli is by phosphorylation (or dephosphorylation) of cellular constituents by interacting cascades of serine/threonine and tyrosine protein kinases (and phosphatases). Survival factors (e.g., growth factors and mitogens) activate receptor tyrosine kinases and selected mitogen-activated, cyclin-dependent, lipid-activated, nucleic acid-dependent, and cyclic AMP-dependent kinases to promote cell survival and proliferation, whereas environmental stress (or death factors such as FAS/APO-1 ligand and TNF-alpha) activates different members of these kinase families to inhibit cell growth and, under some circumstances, promote apoptotic cell death. Because individual kinase cascades can interact with one another, they are able to integrate conflicting exogenous stimuli and provide a link between cell surface receptors and the biochemical pathways leading to cell proliferation or cell death.
Retinal degeneration is one of the leading causes of blindness in the developed world. There is currently no effective therapy. In order to improve our understanding of these conditions, we characterised three animal models of retinal degeneration, namely pigmented retinal degeneration slow (rds) mouse, retinal dystrophy (Rdy) cats and the miniature longhaired dachshund dog (MLHD). We then carried out a series of treatment trials using both inhibitors of caspases and neurotrophic factors in these animals. Using immunohistochemical techniques, there was mis-localisation of opsin, reduction of synaptophysin in the outer plexiform layer and increased expression of glial fibrillary acidic protein in Muller cells in all of these animal models. Apoptosis was found to be the main mode of photoreceptor cell death. Photoreceptors in the pigmented rds mouse were found to degenerate more slowly than the albino rds mouse. The Rdy cats had opsin labelled neurites similar to those described in human retinal degeneration. The MLHD dog was demonstrated to have a cone-rod dystrophy (CORD) based on electrophysiological studies. It represents the first animal model of CORD. As apoptosis is the main mode of cell death, we attempted to either block the apoptotic pathway by caspases inhibitors or prevent photoreceptors entering the apoptotic pathway using neurotrophic factors. It appears that DEVD, a caspase-3 inhibitor, might slow photoreceptor cell loss in the pigmented rds mice, but z-VAD and YVAD did not. We have also found that Axokine, a ciliary neurotrophic factor analogue, might prolong photoreceptor cell survival in the Rdy cats, but brain derived neurotrophic factor had no effect. Furthermore, Axokine appears to preserve retinal function in the MLHD dogs as measured by electroretinography for a short period of time after an initial suppression.
Background
Oligodendrocytes, which form myelin, enable rapid and efficient nerve conduction. Destruction of myelin causes demyelinating diseases such as multiple sclerosis. Primary oligodendrocyte progenitor cells (OPCs) from postnatal rodents have been utilized to elucidate the developmental mechanism of oligodendrocytes in vitro. However, this process is complicated and takes up to several weeks.
New method
We established a method to culture OPCs from neonatal rat brain in DMEM/F-12 with Stem-Pro, bFGF (10 ng/mL), and rhPDGF (30 ng/mL). The culture, without shaking or immunopanning, became OPC-enriched rather than a mixed glial culture.
Results
Immunofluorescent analysis using cell lineage markers suggested that these cells were initially glial progenitors, which gradually changed to OPCs with a few cells further differentiating into oligodendrocytes. Using compounds that promote OPC differentiation, we confirmed that these cells were compatible for high-throughput screening in a 96-well plate format. In co-culture with dorsal root ganglion neuron, OPCs showed myelin sheath-like morphologies. This method was also applicable to mouse OPCs.
Comparison with existing methods
Although the purity of the OPCs was not comparable to that after immunopanning, most cells were of the oligodendrocyte lineage at 8 DIV, while less than 10% were astrocytes. This method requires mediums with only two growth factors without any specific equipment like antibodies or magnet and takes simple procedures.
Conclusions
The simplicity and high yield of our method make it a good choice when working with oligodendrocytes/OPCs. We believe that this method is an affordable protocol for various biological applications without any special techniques or equipment.
Interactions between different tissue compartments are crucial for embryonic development. These interactions include long-range effects by serum factors (hormonese and growth factors), adhesive interactions between cells, and inductive interactions between two dissimilar cell populations located in close proximity to each other. In vitro cultures of embryonic tissues are well suited for analysis of these phenomena. In such cultures, the normal histoarchitecture is retained, but it is nevertheless possible to manipulate morphogenesis at least to some extent. Organ culture of embryonic tissues has been used to study the development of most parenchymal organs, such as the lung, heart, liver, many different glands, and the kidney. These studies have convincingly demonstrated the role of inductive interactions in development (see Wessells, 1977). We are using the developing kidney to study these phenomena.
The development of most regions of the vertebrate nervous system includes a distinct phase of neuronal degeneration during which a substantial proportion of the neurons initially generated die. This degeneration primarily adjusts the magnitude of each neuronal population to the size or functional needs of its projection field, but in the process it seems also to eliminate many neurons whose axons have grown to either the wrong target or an inappropriate region within the target area. In addition, many connections that are initially formed are later eliminated without the death of the parent cell. In most cases such process elimination results in the removal of terminal axonal branches and hence serves as a mechanism to "fine-tune" neuronal wiring. However, there are now also several examples of the large-scale elimination of early-formed pathways as a result of the selective degeneration of long axon collaterals. Thus, far from being relatively minor aspects of neural development, these regressive phenomena are now recognized as playing a major role in determining the form of the mature nervous system.
We have cloned the C. elegans cell death gene ced-3 . A ced-3 transcript is most abundant during embryogenesis, the stage during which most programmed cell deaths occur. The predicted CED-3 protein shows similarity to human and murine interleukin-1β-converting enzyme and to the product of the mouse nedd-2 gene, which is expressed in the embryonic brain. The sequences of 12 ced-3 mutations as well as the sequences of ced-3 genes from two related nematode species identify sites of potential functional importance. We propose that the CED-3 protein acts as a cysteine protease in the initiation of programmed cell death in C. elegans and that cysteine proteases also function in programmed cell death in mammals.
Mutations in the gene ced-4 block almost all of the programmed cell deaths that normally occur during Caenorhabditis elegans development. We have cloned the ced-4 gene using a ced-4 mutation caused by the insertion of the transposon Tc4. When microinjected into a ced-4 animal, a 4.4 kb DNA fragment derived from the wild-type strain and corresponding to the region of the Tc4 insertion in the mutant ced-4(n1416) rescues the Ced-4 mutant phenotype. The ced-4 gene encodes a 2.2 kb RNA transcript. This mRNA is expressed primarily during embryogenesis, when most programmed cell deaths occur. The Ced-4 protein, as deduced from cDNA and genomic DNA clones, is 549 amino acids in length. Two regions of the putative Ced-4 protein product show some similarity to known calcium-binding domains.
Programmed cell death is a physiological process that eliminates unwanted cells. The bcl-2 gene regulates programmed cell death in mammalian cells, but the way it functions is not known. Expression of the human bcl-2 gene in the nematode Caenorhabditis elegans reduced the number of programmed cell deaths, suggesting that the mechanism of programmed cell death controlled by bcl-2 in humans is the same as that in nematodes.
Kidney development starts with an epithelial bud and a domain of committed mesenchyme inducing one another so that the former gives a bifurcating duct system and the latter nephrons. As these events take place in vitro, the mechanisms underpinning nephrogenesis can be investigated experimentally. Recent work has shown the diversity of regulatory molecules expressed during kidney development and begun to clarify the molecular basis of mesenchyme induction, but there is more to come.
Programmed cell death occurs in most animal tissues at some stage of their development, but the molecular mechanism by which it is executed is unknown. For some mammalian cells, programmed death seems to occur by default unless suppressed by signals from other cells. Such dependence on specific survival signals provides a simple way to eliminate misplaced cells, for regulating cell numbers and, perhaps, for selecting the fittest cells. But how general is this dependence on survival signals?
The gene ced-9 of the nematode Caenorhabditis elegans acts to protect cells from programmed cell death. A mutation that abnormally activates ced-9 prevents the cell deaths that occur during normal C. elegans development. Conversely, mutations that inactivate ced-9 cause cells that normally live to undergo programmed cell death; these mutations result in embryonic lethality, indicating that ced-9 function is essential for development. The ced-9 gene functions by negatively regulating the activities of other genes that are required for the process of programmed cell death.