Despite several studies examining the contribution of allorecognition pathways to acute and chronic rejection of vascularized murine allografts, little data describing activation of alloreactive T cells by mouse vascular endothelium exist. We have used primary cultures of resting or IFN-γ-activated C57BL/6 (H-2b) vascular endothelial cells as stimulators and CD8+ T lymphocytes isolated from CBA/J (H-2k) mice as responders. Resting endothelium expressed low levels of MHC class I, which was markedly up-regulated after activation with IFN-γ. It also expressed moderate levels of CD80 at a resting state and after activation. Both resting and activated endothelium were able to induce proliferation of unprimed CD8+ T lymphocytes, with proliferation noted at earlier time points after coculture with activated endothelium. Activated endothelium was also able to induce proliferation of CD44low naive CD8+ T lymphocytes. Activated CD8+ T lymphocytes had the ability to produce IFN-γ and IL-2, acquired an effector phenotype, and showed up-regulation of the antiapoptotic protein Bcl-xL. Treatment with CTLA4-Ig led to marked reduction of T cell proliferation and a decrease in expression of Bcl-xL. Moreover, we demonstrate that nonhemopoietic cells such as vascular endothelium induce proliferation of CD8+ T lymphocytes in a B7-dependent fashion in vivo. These results suggest that vascular endothelium can act as an APC for CD8+ direct allorecognition and may, therefore, play an important role in regulating immune processes of allograft rejection.