Interaction of cytokines and growth factor in the regulation of verotoxin-induced apoptosis in cultured human endothelial cells.
The role of CD77, inflammatory cytokines and endothelial cell growth factor (ECGF) in verotoxin (VT)-induced apoptosis in human umbilical vein endothelial cells (HUVECs) was studied. Apoptosis was detected using annexin V and propidium iodide staining. The expression of CD77 antigen was measured on a FACStar flow cytometer using specific monoclonal antibodies. Our experiments showed that HUVECs had very low initial levels of CD77 and were resistant to VT. Treatment with tumour necrosis factor alpha (TNF-alpha) resulted in a significant upregulation of CD77 expression and sensitized endothelial cells to VT-mediated apoptosis. HUVECs incubated with a combination of cytokines [TNF-alpha and interferon gamma (IFN-gamma), both 500 U/ml] showed more pronounced upregulation of CD77 expression (> sixfold at 48 h) and underwent apoptosis, suggesting that TNF-alpha and IFN-gamma have a synergistic effect on CD77 expression in HUVECs and can induce apoptosis without VT. Cells pretreated with TNF-alpha and IFN-gamma and incubated with VT showed the most pronounced (14-fold) increase in CD77 expression. ECGF had a partial protective effect against cytokine- and VT-induced apoptosis. Taken together, our data suggest that CD77 antigen is involved in the regulation of endothelial cell apoptosis.