Expression of Thrombomodulin in Atherosclerotic Lesions and Mitogenic Activity of Recombinant Thrombomodulin in Vascular Smooth Muscle Cells

Abstract

Abstract —Thrombomodulin (TM), a thrombin receptor protein found on the endothelial cell surface, contains 6 tandem epidermal growth factor (EGF)–like structures. Recombinant human TM peptide containing these 6 EGF-like domains (rTM e 1–6) exhibits mitogenic activity in Swiss 3T3 cells. We examined the localization of TM in atherosclerotic lesions and the effects of rTM e 1–6 on the growth of cultured rat vascular smooth muscle cells (SMCs). Immunohistochemical analysis demonstrated that TM antigen was localized on monocytes, macrophages, and vascular SMCs. In cultured vascular SMCs, rTM e 1–6 accelerated [ 3 H]thymidine uptake into DNA in a dose-dependent manner up to 3.4 times the control level. This mitogenic activity was abolished by addition of polyclonal anti-human TM antibody. The rTM e 1–6–induced mitogenesis was enhanced by EGF. However, a neutralizing monoclonal antibody against the EGF receptor (monoclonal antibody 225) did not inhibit the mitogenic activity of rTM e 1–6. Calphostin C, a specific protein kinase C inhibitor, and lavendustin-A, an inhibitor of EGF receptor–specific protein tyrosine kinase, inhibited the mitogenic activities of both rTM e 1–6 and EGF. Finally, rTM e 1–6 treatment increased the level of phosphorylated mitogen-activated protein kinase in SMCs. Together, these results suggest that TM expression in atherosclerotic lesions may be associated with promotion of atherosclerosis through its mitogenic activity in vascular SMCs.