Membrane cofactor protein (MCP), decay-accelerating factor (DAF), and CD59 are complement (C) regulatory proteins that protect human hematopoietic cells from damage induced by autologous complement. Using monoclonal antibodies in both western blotting studies and flow cytometry, we found that endothelial cells (EC) expressed on their surface MCP, DAF, and CD59 molecules. We tested whether stimulation of endothelial cells by known EC activators such as LPS, TNF-alpha, IL-1 beta, and IL-4 regulates the expression of these C regulatory proteins. EC activation was assessed in this study by an up-regulation of ICAM-1 expression. Treatment of EC with various concentrations of TNF-alpha, IL-1 beta, or IL-4, at different incubation times did not increase MCP expression on EC surface membrane. In contrast, the level of DAF was altered during EC activation: LPS or IL-1 beta treatment resulted in a slight increase of DAF expression; the most up-regulatory effect was obtained with IL-4. Up-regulation of surface DAF on EC required prolonged treatment of cells with these agents. The level of CD59 was far greater than that of DAF or MCP on EC, and was slightly up-regulated by TNF-alpha and down-regulated by IL-1 beta. These findings indicate that the levels of C regulatory proteins are regulated in an independent fashion on EC, MCP is not regulated by any cytokine tested, while DAF is an EC activation antigen, although it has a different augmentation profile from ICAM-1 since IL-4 up-regulates DAF but not ICAM-1.