Thy-1 is a glycosylphosphatidylinositol-anchored member of the Ig superfamily whose function, particularly in the human, remains unknown. We have demonstrated that human Thy-1 is expressed on endothelial cells (EC) both in situ and on the surface of cultured human umbilical vein EC and dermal microvascular EC (DMEC). The expression of the molecule decreased with serial passage but was restored by treatment of EC with PMA and phorbol-12,13-dibutyrate (PBu), which increased Thy-1 by up to 100-fold in a dose-dependent manner. This increase was first detectable at 12 h post-stimulation, peaked at 48 h, and was maintained at 72 h. In PBu-stimulated DMEC, Western blotting revealed Thy-1 to be a 29-kDa molecule, while Northern analysis demonstrated an increase in steady-state Thy-1 mRNA. Thy-1 expression was also induced on DMEC by treatment with TNF. Inhibition studies showed that the induction of Thy-1 by PBu and TNF was protein synthesis dependent. The up-regulation of Thy-1 by PBu, but not TNF, was inhibited by the protein kinase C inhibitor RO31-8220, suggesting the presence of protein kinase C-dependent and -independent pathways for Thy-1 expression. To investigate the function of Thy-1 on human EC, we studied changes in intracellular calcium concentration ([Ca2+]i) following cross-linking of Thy-1 on human umbilical vein EC. This resulted in a rapid transient rise of [Ca2+]i in EC. Our results demonstrate for the first time the presence of Thy-1 on cultured human EC. The expression on EC, the inducibility by TNF, and the ability to transmit signals suggest that Thy-1 may have an important role in inflammatory responses.