Prevention of cycloheximide-induced apoptosis in hepatocytes by adenosine and by caspase inhibitors

Blom, W. Marty; Bont, H. J. G. M. De; Meijerman, Irma; Mulder, Gerard J.; Nagelkerke, J. Fred

doi:10.1016/s0006-2952(99)00268-3

Cited by 32 publications

(23 citation statements)

References 40 publications

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“…PC12 and Rat-1 cells were treated with varying doses of cycloheximide, followed by analysis of protein synthesis by [ 35 S]methionine incorporation and of apoptosis by nuclear morphology. As previously reported for other cell types (2,34,40), inhibition of protein synthesis by cycloheximide induced apoptosis of both PC12 and Rat-1 cells (Fig. 6), consistent with a requirement for translation in cell survival.…”

Section: Resultssupporting

confidence: 90%

Role of Translation Initiation Factor 2B in Control of Cell Survival by the Phosphatidylinositol 3-Kinase/Akt/Glycogen Synthase Kinase 3β Signaling Pathway

Pap

Cooper

2002

Molecular and Cellular Biology

146

104

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The phosphatidylinositol 3-kinase (PI 3-kinase)/Akt signaling pathway is an important mediator of growth factor-dependent survival of mammalian cells. A variety of targets of the Akt protein kinase have been implicated in cell survival, including the protein kinase glycogen synthase kinase 3␤ (GSK-3␤). One of the targets of GSK-3␤ is translation initiation factor 2B (eIF2B), linking global regulation of protein synthesis to PI 3-kinase/Akt signaling. Because of the central role of protein synthesis, we have investigated the involvement of eIF2B, which is inhibited as a result of GSK-3␤ phosphorylation, in programmed cell death. We demonstrate that expression of eIF2B mutants lacking the GSK-3␤ phosphorylation or priming sites is sufficient to protect both Rat-1 and PC12 cells from apoptosis induced by overexpression of GSK-3␤, inhibition of PI 3-kinase, or growth factor deprivation. Consistent with these effects on cell survival, expression of nonphosphorylatable eIF2B prevented inhibition of protein synthesis following treatment of cells with the PI 3-kinase inhibitor LY294002. Conversely, cycloheximide induced apoptosis of PC12 and Rat-1 cells, further indicating that protein synthesis was required for cell survival. Inhibition of translation resulting from treatment with cycloheximide led to the release of cytochrome c from mitochondria, similar to the effects of inhibition of PI 3-kinase. Expression of nonphosphorylatable eIF2B prevented cytochrome c release resulting from PI 3-kinase inhibition but did not affect cytochrome c release or apoptosis induced by cycloheximide. Regulation of translation resulting from phosphorylation of eIF2B by GSK-3␤ thus appears to contribute to the control of cell survival by the PI 3-kinase/Akt signaling pathway, acting upstream of mitochondrial cytochrome c release.

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Section: Resultssupporting

confidence: 90%

Role of Translation Initiation Factor 2B in Control of Cell Survival by the Phosphatidylinositol 3-Kinase/Akt/Glycogen Synthase Kinase 3β Signaling Pathway

Pap

Cooper

2002

Molecular and Cellular Biology

146

104

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“…Pre-treatment of hepatocytes with broad spectrum caspase inhibitor, z-VAD-FMK and more specific inhibition of caspase-3 (Ac-DEVD-CHO or z-DEVD-FMK), caspase-8 (z-IETD-FMK) and caspase-9 (z-LEHD-FMK) has provided evidence in the same way as had been reported in previously studied cell types and cell lines (Cain et al 1996;Jones et al 1998;Blom et al 1999;Woo et al 1999;Albright et al 2003). Similarly, we show in the present study that staurosporine-induced apoptosis can be blocked by treatment with caspase inhibitors.…”

Section: Discussionsupporting

confidence: 71%

Apoptosis and Its Suppression in Hepatocytes Culture

Mukwena

Al‐Rubeai

2004

Cytotechnology

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In order to achieve the goal of developing extracorporeal liver support devices, it is necessary to optimise bioprocess environment such that viability and function are maximised. Optimising culture medium composition and controlling the constitution of the cellular microenvironment within the bioreactor have for many years been considered vital to achieving these aims. Coupled to this is the need to understand apoptosis, the prime suspect in the demise of animal cultures, including those of hepatocytes. Results presented here show that absent nutrients including glucose and amino acids play a substantial part in the induction of apoptosis. The use of chemical apoptosis inhibitors was utilised to investigate key components of hepatic apoptosis where caspases, predominantly caspase 8, were implicated in staurosporine (STS)-induced HepZ apoptosis. Caspase 9 and 3 activation although recorded was of less significance. Interestingly, these results were not consistent with those of mitochondrial membrane depolarisation where inhibition of caspase activation appeared to drive depolarisation. Inhibition of mitochondrial permeability transition and use of anti-oxidants was unsuccessful in reducing apoptosis, caspase activation and mitochondrial membrane depolarisation. In further studies, the anti-apoptotic gene bcl-2 was over-expressed in HepZ, resulting in a cell line that was more robust and resistant to death induced by glucose and cystine deprivation and treatment with STS. Bcl-2 did not however show significant cytoprotectivity where apoptosis was stimulated by deprivation of glutamine and serum. Overall, results indicated that although apoptosis can be curbed by use of chemical inhibitors and genetic manipulation, their success is dependent on apoptotic stimuli.

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“…54,59,60 To evaluate the cell death induced under our conditions, we employed several markers to characterize apoptotic cell death, including examination of DNA fragmentation both by gel electrophoresis and in situ detection of fragmented DNA by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), measurement of nuclear fragmentation by Hoechst staining, and activation of caspase-3 and cleavage of poly (ADP ribose) polymerase (PARP).…”

Section: Resultsmentioning

confidence: 99%

p53, Apaf-1, caspase-3, and -9 are dispensable for Cdk5 activation during cell death

Lin

Zakeri

2005

Cell Death Differ

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Cyclin-dependent kinase 5 (Cdk5) is a member of the cyclindependent kinase family that is mostly seen in neurons, does not vary with cell cycle, and is activated in many neurodegenerative disorders and other non-neuronal pathologies, but its relationship to non-neuronal apoptosis is not understood, nor is the control of the activation of Cdk5 by its activators. The most widely studied activator of Cdk5, p35, is cleaved to p25 by calpain, an event that has been linked with activation of Cdk5 and neuronal death. Here we report that calpain-mediated Cdk5/p25 activation accompanies nonneuronal as well as neuronal cell death, suggesting that the p35/calpain/p25/Cdk5 activation sequence is a general feature of cell death. We further demonstrate that Cdk5 can be activated in the absence of p53, Apaf-1, caspase-9, and -3 during cell death, indicating that its activation relates more to cell death than to a specific pathway of apoptosis.

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Prevention of cycloheximide-induced apoptosis in hepatocytes by adenosine and by caspase inhibitors

Cited by 32 publications

References 40 publications

Role of Translation Initiation Factor 2B in Control of Cell Survival by the Phosphatidylinositol 3-Kinase/Akt/Glycogen Synthase Kinase 3β Signaling Pathway

Role of Translation Initiation Factor 2B in Control of Cell Survival by the Phosphatidylinositol 3-Kinase/Akt/Glycogen Synthase Kinase 3β Signaling Pathway

Apoptosis and Its Suppression in Hepatocytes Culture

p53, Apaf-1, caspase-3, and -9 are dispensable for Cdk5 activation during cell death

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