IN THE ABSENCE OF EPSTEIN-BARR-VIRUS INFECTION, PHORBOL ESTER MODULATES APOPTOSIS IN CYCLOHEXIMIDE-TREATED BURKITTS-LYMPHOMA (BJA-B) CELLS

We have shown previously that cycloheximide (CHX), a potent protein sy nthesis inhibitor, induces high levels of apoptosis in Epstein-Barr vi rus free (EBV(-)) Burkitt's lymphoma (BJA-B) cells! with comparably re duced levels of apoptosis in the EBV positive (EBV(+)) cells, Modulati on of CHX-induced apoptosis in EBV(-) and (+) B cells is reported here using concurrent treatment with phorbol ester (phorbol 12,13-dibutyra te, PdBu). Cells were collected at 0, 3, 6, 12, 24 and 48 hours after treatment with (i) 1 mu g/ml CHX, (ii) 0.1 mu g/ml PdBu (1 hour pretre atment before Oh), or (iii) CHX + PdBu (CHX added at Oh, 1 hour after PdBu). Control cultures were untreated. Apoptotic, necrotic or viable cells were quantified using histological, ultrastructural and biochemi cal parameters. Protein synthesis was assessed using S-35-methionine i ncorporation. Intracellular calcium concentrations were measured using flow cytometry. PdBu alone had little effect on cell death. High leve ls of CHX-induced apoptosis in EBV(-) cells were significantly reduced by concurrent addition of PdBu (P < 0.005). In contrast, low levels o f CHX-induced apoptosis in EBV(+) cells were not significantly altered by PdBu treatment. In EBV(-) cells, a negative relationship was obser ved between levels of apoptosis and calcium concentrations, whereas in EBV(+) cells, there was negligible correlation between these paramete rs. Thus high levels of CHX-induced apoptosis in EBV(-) cells occur vi a a PKC-dependent pathway, whereas CHX treatment of EBV(+) cells induc es comparatively low levels of apoptosis that occur via a PKC-independ ent mechanism. The results application in the therapeutic intervention for cancers developing in association with EBV infection.