Osteopontin is produced by rat cardiac fibroblasts and mediates A(II)-induced DNA synthesis and collagen gel contraction

Angiotensin II (A(II)) is a critical factor in cardiac remodeling which involves hypertrophy, fibroblast proliferation, and extracellular matrix production. However, little is known about the mechanism by which A(II) accelerates these responses. Osteopontin is an acidic phosphoprotein with RGD (arginine-glycine-aspartate) sequences that are involved in the vascular smooth muscle cell remodeling process, We identified the presence of osteopontin mRNA and protein in cultured rat cardiac fibroblasts and its prominent regulation by A(II) (10(-11) M). Osteopontin message levels were increased fourfold (P < 0.01) and protein fivefold (P < 0.05) at 24 h after addition of A(II) (10(-7) M), This response was inhibited by the AT(1) receptor blocker, losartan, Osteopontin mRNA levels were increased in hypertrophied ventricles from animals with renovascular hypertension (1.6-fold, P < 0.05) and aortic banding (2.9-fold, P < 0.05). To examine the function of osteopontin, we determined its effects on (a) the ability of cardiac fibroblasts to contract three-dimensional collagen gels and (b) cardiac fibroblast growth. A monoclonal antibody against osteopontin partially blocked A(II)-induced three-dimensional collagen gel contraction by cardiac fibroblasts (64 +/- 4 vs, 86 +/- 5% in the presence of antibody, P < 0.05), while osteopontin itself promoted contraction of the gels by fibroblasts (71 +/- 5%, P < 0.05 compared with control), Either a monoclonal antibody against beta(3) integrin which is a ligand for osteopontin or the RGD peptide blocked both A(II) and osteopontin-induced collagen gel contraction, Thus, the osteopontin RGD sequence binds to beta(3) integrins on the fibroblast to promote fibroblast binding to collagen, A(II) induced a threefold increase in DNA synthesis of cardiac fibroblasts, which was completely blocked by antibodies against osteopontin and beta(3) integrin, or by RGD peptide, but not by controls. Thus, A(II)-induced growth of cardiac fibroblasts also requires osteopontin engagement of the beta(3) integrin, Taken together, these results provide the first evidence that osteopontin is a potentially important mediator of A(II) regulation of cardiac fibroblast behavior in the cardiac remodeling process.