A 20-day treatment with LF15–0195, a deoxyspergualine analog, induced long-term heart allograft survival in the rat without signs of chronic rejection. LF15–0195-treated recipients did not develop an anti-donor alloantibody response. Analysis of graft-infiltrating cells, IL10, TNFα, IFNγ mRNA and iNOS protein expression in allografts, 5 days after transplantation, showed that they were markedly decreased in allografts from LF15–0195-treated recipients compared with allografts from untreated recipients. Surprisingly, spleen T cells from LF15–0195 recipients, 5 days after grafting, were able to proliferate strongly in vitro, when stimulated with donor cells, but had reduced mRNA expression for IFNγ compared with spleen T cells from untreated graft recipients. Furthermore, when T cells from naive animals were stimulated in vitro, using anti-CD3 and anti-CD28, LF15–0195 also increased T-cell proliferation in a dose-dependent fashion; however, these cells expressed less of the Th1-related cytokines, IFNγ and IL2, compared with untreated cells, suggesting that LF15–0195 could act on T-cell differentiation. In conclusion, we show here that a short-term treatment with LF15–0195 induced long-term allograft tolerance, decreasing the in situ anti-donor response, and we illustrate evidence for the development of regulatory mechanisms.