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

Angelo Spinedi, Sabrina Di Bartolomeo, Federica Di Sano, Carlo Rodolfo, Arianna Ambrosino, Mauro Piacentini

Research output: Contribution to journalArticlepeer-review

Abstract

The protein phosphatase inhibitor okadaic acid (OA) dose-dependently 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 μM) 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 CA 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 μM) 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. Short chain ceramide (30 μM) 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.

Original languageEnglish
Pages (from-to)618-623
Number of pages6
JournalCell Death and Differentiation
Volume6
Issue number7
Publication statusPublished - 1999

Keywords

  • Apoptosis
  • Caspases
  • Ceramide
  • Neuroepithelioma
  • Okadaic acid

ASJC Scopus subject areas

  • Cell Biology

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