Ceramide accumulation precedes caspase-dependent apoptosis in CHP-100 neuroepithelioma cells exposed to the protein phosphatase inhibitor okadaic acid

Abstract

The protein phosphatase inhibitor okadaic acid (OA) dosedependently induced apoptosis in CHP-100 neuroepithelioma cells when administered for 24 h at concentrations ranging from 10 ± 100 nM. Apoptosis was largely, albeit not completely, dependent on cystein protease (caspase) activation. CPP32 processing and poly(ADP-ribose) polymerase (PARP) cleavage started to be observed only at 20 nM OA; moreover, the caspase inhibitor Z-Val-Ala-DL-Asp-fluoromethylketone (Z-VAD.fmk) (100 mM) had negligible effect on apoptosis induced by 10 nM OA, but rescued from death an increasing cell fraction as OA concentration was raised from 20 ± 100 nM. Cell treatment for 24 h with OA induced ceramide accumulation; the phenomenon started to be evident at 20 nM OA and reached its maximum at 50 ± 100 nM OA. In cells exposed to 50 nM OA, ceramide was already elevated by 5 h; at this time, however, PARP cleavage and apoptosis were not yet observed. Z-VAD.fmk (100 mM) had no effect on ceramide elevation induced by 50 nM OA within 5 h, but markedly reduced ceramide accumulation as the incubation was prolonged to 24 h. The latter phenomenon was accompanied by elevation of glucosylceramide levels, thus suggesting that a caspase-dependent reduction of glucosylceramide synthesis might contribute to late ceramide accumulation. Shortchain ceramide (30 mM) induced apoptosis in CHP-100 cells and its effect was additive with that evoked by OA (10 ± 20 nM). These results suggest that ceramide generation might be an important mechanism through which sustained protein phosphatase inhibition induces caspase activation and apoptosis in CHP-100 cells.

Activation of Neutral Sphingomyelinase Participates in Ethanol-Induced Apoptosis in Hep G2 Cells

“…The interaction of ceramide and caspase-3 varies with the cell type. In PC12 cells and CHP-100 cells, ceramide formation leads to caspase-3 activation (Yoshimura et al, 1998;Spinedi et al, 1999), whereas in human T and B lymphocytes, Fas-induced generation of ceramide is activated by caspase-3 (Genestier et al, 1998). In the present study, we found that C 2 -Cer increased the activity of caspase-3, indicating that caspase-3 in Hep G2 cells can be a downstream target of ceramide.…”

Signaling events mediating activation of brain ethanolamine plasmalogen hydrolysis by ceramide

“…Consistent with this, complex modulation of Cer levels evoked by okadaic acid has been reported [34]. Although we cannot rule out the possibility that okadaic acid itself affects sphingomyelinase, it is very likely that the metabolic fate of Cer is affected.…”

Endosomes and lysosomes play distinct roles in sulfatide-induced neuroblastoma apoptosis: potential mechanisms contributing to abnormal sulfatide metabolism in related neuronal diseases

“…Accumulation of sulfatide and ceramide as well as cell apoptosis were also demonstrated with primary neuron cultures. Accordingly, these results indicate that abnormal sulfatide metabolism/trafficking through direct internalization can induce neuronal cell apoptosis through endosome-generated ceramide accumulation [26][27][28] and/or lysosome swelling and sulfatide toxicity [29,30].…”