The effect of TGFβ1 and IL-12 on calcium-independent cytotoxic pathways was investigated. We have previously demonstrated that the regulatory effect of TGFβ1 and IL-12 on human alloreative CTL activity was associated with regulation of perforin and granzyme B gene expression. To determine the effect of both cytokines on the alternative cytotoxic pathway involving FasL and mTNF, we first investigated the expression of both molecules on human primary alloactivated T cells. Our results show that human allostimulated T lymphocytes express FasL. Cell lysis experiments demonstrate that the FasL cytotoxic pathway is involved in the killing of specific target cells mediated by human alloreactive CTL. In addition, allogeneic stimulation induced significant mTNF expression on both CD4+ and CD8+ responder T cells. Using TNF-sensitive target cells, we also demonstrate that the mTNF-mediated cytotoxic pathway is involved in the cytotoxic activity of human primary allostimulated T lymphocytes. Neither TGFβ1 nor IL-12 had an effect on FasL or mTNF expression. Furthermore, addition of TGFβ1 or IL-12 at the initiation of the MLR had no significant effect on Fas- and mTNF-mediated cytotoxicity.
Taken together, our results provide a novel insight into the differences between regulation by cytokines of perforin-dependent and -independent cytotoxic mechanisms. Unlike their role in the perforin/granzyme B pathway, TGFβ1 and IL-12 do not appear to mediate any regulatory effect on FasL and mTNF cytotoxic pathways used by human alloreactive primary CTL.