Effects of an aminosteroid inhibitor of phospholipase C-dependent processes on the TCR-mediated signal transduction pathway in human T cells.

Phospholipase C (PLC) is a key enzyme in the T cell antigen receptor (TCR)-mediated signal transduction pathway in human T cells. Agonist-induced PLC activation leads to a cascade of intracellular events that ultimately regulate gene transcription and T cell activation. We studied the effects of U-73122, a putative inhibitor of PLC-dependent events, on TCR/CD3 complex-mediated early and late events in human T cells. Both anti-CD3 monoclonal antibody-induced 1,4,5-inositol trisphosphate (IP3) and free intracytoplasmic calcium [Ca2+]i increases were inhibited by U-73122 (0.05-0.1 microM), but not by the related inactive analog, U-73343. U-73122 did not affect thapsigargin-evoked [Ca2+]i increase in T cells, indicating a specific mode of inhibition of CD3 signaling. Late events in T cell activation like CD3-mediated T cell proliferation and mitogen-induced interleukin 2 receptor (IL2-R) expression were also inhibited by this agent. T cell proliferation induced by a combination of a phorbol ester and ionomycin was not affected by U-73122. Although an agonist effect on basal IP3 and [Ca2+]i levels was observed with high concentrations of U-73122, the inhibitor alone did not induce any proliferative effect or IL2-R expression in T cells. Our results demonstrate for the first time that U-73122 is a specific inhibitor of PLC-dependent processes in human T cells and could serve as a valuable tool for studying T cell signal transduction pathways.