The short-chain fatty acid n-butyrate has recently been shown in vitro to specifically downregulate T cell reactivity to nominal antigen or to alloantigen, which possibly results from inhibition of cell cycle progression in early G1 phase during antigen contact. In the present study, we investigated the effect of cyclosporin A (CyA) on the modulation of alloreactivity in human mixed lymphocyte culture (MLC) by n-butyrate. Whereas in primary culture, CyA additively enhanced inhibition of DNA synthesis by n-butyrate, the effect of this agent on secondary T cell reactivity was clearly antagonized by CyA. Thus, specific downregulation of proliferative responsiveness to restimulation with antigen from the original donor, observed in cultures pretreated with n-butyrate alone, was at least partially prevented by the addition of CyA to the primary culture. Our in vitro finding indicates that specific downregulation of T cell alloreactivity by n-butyrate might depend on a calcium-dependent T cell receptor (TCR)-mediated signal sensitive to the immunosuppressive action of CyA.