Role of protein kinase C and cytokines on the function and production of cytolytic granules in alpha CD3-activated killer-cell-mediated killing of tumor cells.
The effects of PMA and staurosporine (PKC depletor/antagonist) and IL-2/IL4 were used to determine the role of PKC and cytokine on alpha CD3-induced activated killer cells (CD3-AK). The present study examines their effects on the production of BLT-esterase and on the effector function of CD3-AK cells as well as the cytolytic granules. The production of BLT-esterase generally correlated with the cytolytic activity of CD3-AK cells and was reduced by PKC depletor/inhibitor but increased by IL-4. In studying the effector function of CD3-AK cells, we found that adding PMA or SSP at the effector phase inhibited the PKC-dependent slow lysis. PMA, but not SSP, also reduced fast lysis, which was shown to be a PKC-independent event. Additional experiments were performed to determine the effect of PKC on the lytic granules and to ascertain whether PMA has other effects on the effector-to-target relationship unrelated to PKC. It was found that neither PMA nor SSP affects the function of cytolytic granules, as measured by hemolytic assay against anucleated target (SRBC). These findings indicate that PKC has no direct effect on the granules. During testing against the nucleated tumor target through a novel approach using non-cytolytic surrogate killers, the lytic activity of the granules was inhibited by PMA, suggesting that exocytosis or delivery of granules to nucleated target cells may require mobilization of intracellular Ca2+ in the killer cells, and this process is inhibited by PMA. Our findings indicate that PKC and cytokines regulate the production but not the lytic activity of cytolytic granules. Nonetheless, delivery of cytolytic granules from killer cells to the nucleated tumor target appears to be a Ca(2+)-dependent event unrelated to PKC.
Wu
J
J
Office of the Director, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.
Shiver
J
J
Hargrove
M E
ME
Ting
C C
CC
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000906:Antibodies; D000920:Antibody-Dependent Cell Cytotoxicity / Q000502:physiology*; D017252:CD3 Complex / Q000494:pharmacology*; D016207:Cytokines / Q000502:physiology*; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000502:physiology*; D003602:Cytotoxicity, Immunologic; D005260:Female; D053804:Granzymes; D016219:Immunotherapy, Adoptive; D007694:Killer Cells, Natural / Q000187:drug effects / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009374:Neoplasms, Experimental / Q000628:therapy*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D012697:Serine Endopeptidases / Q000378:metabolism; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000470:Alkaloids; D000906:Antibodies; D017252:CD3 Complex; D016207:Cytokines; D011493:Protein Kinase C; D053804:Granzymes; D012697:Serine Endopeptidases; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate