INDUCTION OF APOPTOSIS IN MDA-231 CELLS BY PROTEIN-SYNTHESIS INHIBITORS IS SUPPRESSED BY MULTIPLE AGENTS

In the present study we investigated the ability of several diverse ag ents to inhibit MDA-231 cell death induced by two different protein sy nthesis inhibitors, cycloheximide (CHX) and ricin. Cell death was eval uated by several techniques: trypan blue staining, determination of th e released lactic dehydrogenase, transmission electron microscopy, and DNA fragmentation. Results from DNA gel electrophoresis and electron microscopy suggest a mechanism of death by apoptosis which terminates in necrosis. Approximately 60% of cell death was induced either by a c ontinuous exposure to 30 mu g/ml CHX for 48 hr or by a 1-hr exposure t o 250 pg/ml ricin followed by a subsequent incubation of 48 hr in the absence of the drug. Cell survival, in the protein synthesis-inhibited cells, was enhanced by the following diverse agents: the growth facto rs EGF (20 ng/ml) and IGF-1 (20 ng/ml), the protein kinase C activator 12-0-tetradecanoyl-phorbol-13-acetate (5 ng/ml), the protein kinase A activator 8-bromoadenosine 3':5'-cyclic monophosphate (650 mu g/ml), the nuclease inhibitor aurintricarboxylic acid (100 mu g/ml), and feta l bovine serum (5%). The survival agents that stimulated protein synth esis in the control untreated cells had no effect on the CHX-inhibited protein synthesis, which indicates that new protein synthesis is not required for cell survival. The same survival agents attenuated the co ntinuous decrease in protein synthesis in the ricin-exposed cells; the refore, the involvement of new protein synthesis in the survival mecha nism could not be excluded. The protein kinase C inhibitor staurospori ne blocked, in a dose-dependent manner, the survival effect of 12-0-te tradecanoyl-phorbol-13-acetate and EGF, but not that of aurintricarbox yclic acid or fetal bovine serum, in the protein synthesis-inhibited c ells. These results provide evidence for several distinctive pathways, the activation of which inhibits MDA-231 cell death induced by protei n synthesis inhibitors. Some of these pathways involved activation of protein kinases, probably protein kinase C.