ROLE OF INTRACELLULAR CALCIUM ([CA2-MUSCLE CELLS TO FIBRINOGEN(](I)) AND TYROSINE PHOSPHORYLATION IN ADHESION OF CULTURED VASCULAR SMOOTH)

Objective: Fibrinogen is an independent risk factor for cardiovascular disease. This study has investigated the role of intracellular Ca2+ ( [Ca2+](i)) and tyrosine phosphorylation in the attachment of human and rat-derived cultured vascular smooth muscle cells to fibrinogen. Meth ods: Cells were cultured from human saphenous vein segments (HVSMC) an d from an established rat aortic cell line (A7r5). [Ca2+](i) was measu red using fura-2 and adhesion was studied using pre-coated 96 well pol ystyrene plates. Results: Fibrinogen increased [Ca2+](i) in both cell types. In A7r5 cells fibrinogen-induced increases in [Ca2+](i) were pa rtially inhibited by a peptide containing the amino acid sequence Arg- Gly-Asp (RGD) which interferes with binding to integrins. In contrast RGD increased [Ca2+](i) in HVSMC, but did not inhibit responses to fib rinogen. Ni2+, an inorganic calcium channel blocker largely abolished the rise in [Ca2+](i), but blockers of voltage-operated calcium channe ls failed to affect [Ca2+](i) responses to fibrinogen in either cell t ype. Genistein, an inhibitor of tyrosine kinase inhibited fibrinogen-i nduced rises in [Ca2+](i), while daidzein, an inactive analogue, was w ithout effect. Adhesion of cells to fibrinogen was concentration- and time-dependent. Cell adhesion to fibrinogen was partially inhibited by RGD peptide in both cell types. Adhesion of cell to fibrinogen was in hibited by chelation of [Ca2+](i) with BAPTA-AM, inhibition of Ca2+ en try by Ni2+, and inhibition of tyrosine kinases by genistein, but hepa rin had no effect on adhesion. Conclusions: Vascular smooth muscle cel ls attach to fibrinogen in part through RGD-type interactions. Activat ion of tyrosine kinase(s) and a subsequent rise in [Ca2+](i) appear to be important signals mediating the response to fibrinogen. (C) 1998 E lsevier Science B.V. All rights reserved.