ENHANCEMENT OF TUMOR-NECROSIS-FACTOR CYTOTOXICITY BY LITHIUM-CHLORIDEIS ASSOCIATED WITH INCREASED INOSITOL PHOSPHATE ACCUMULATION

We have previously reported that LiCl increases considerably the cytot oxic activity of TNF towards some transformed cell lines such as L929. Here we show that treatment of these cell lines with the combination of TNF and LiCl leads to the prolonged accumulation of inositol monoph osphate, inositol bisphosphate, and inositol trisphosphate, whereas tr eatment with TNF or LiCl alone did not. In contrast, both a LiCl-unres ponsive TNF-sensitive cell line and TNF-resistant cell lines did not r espond with increased accumulation of inositol phosphates (IP(n)) upon treatment with the combination of TNF and LiCl. Furthermore, the comb ination of TNF and LiCl induced a transient increase in cytidine dipho sphate-diacylglycerol in L929 cells. Increased IP(n) and cytidine diph osphate-diacylglycerol accumulation preceded the onset of cell killing by approximately 1 h. TNF-mediated cytotoxicity and TNF-induced IP(n) accumulation were equally sensitive to inhibition by the phospholipas e inhibitor neomycin and to stimulation by the protein kinase inhibito r staurosporine. Characterization of the inositol bisphosphate isomers by HPLC analysis revealed that the TNF + LiCl-induced increase in IP( n) levels was due to activation of a phospholipase C and not of a phos pholipase D. In contrast to TNF, several other cytotoxic agents did no t increase IP(n) production upon application in the presence of LiCl. The TNF + LiCl-induced increase in inositol triphosphate suggests a ro le for intracellular Ca2+ mobilization in TNF action. Moreover, severa l agents that lower the intracellular Ca2+ concentration inhibited TNF cytotoxicity. In conclusion, our data provide evidence that TNF cytot oxicity and its enhancement by LiCl are mediated by increased IP(n) ac cumulation resulting in Ca2+ mobilization.