TRANSFORMING GROWTH-FACTOR-BETA-1 INHIBITS THE PROLIFERATIVE EFFECT OF INSULIN ON HUMAN INFRAGENICULAR VASCULAR SMOOTH-MUSCLE CELLS

Purpose: The distribution of atherosclerotic arterial disease in diabe tes mellitus characteristically involves the infragenicular arterial t ree including the anterior tibial, posterior tibial, and peroneal arte ries. The proliferation of vascular smooth muscle cell (VSMC) is essen tial in the development of the atherosclerotic lesion. It has long bee n held that insulin plays a causative role in the formation of the ath erosclerotic lesion in diabetes. We studied the role played by insulin in the proliferation of these cells in culture and the interaction of insulin with transforming growth factor beta 1 (TGF beta 1), a factor known. for its possible inhibitory effects. Methods: We have grown an d characterized a line of VSMC harvested from atherosclerotic infragen icular arteries of human subjects undergoing below-knee amputation. Th e cultures were defined as being of VSMC origin by immunohistochemical staining with or-smooth muscle actin. Confluent cultures of passages 4 through 7 were seeded into six well plates at a density of 5000 cell s/well. After serum deprivation the cells were exposed to insulin (100 ng/ml) alone or in combination with TGF beta 1 (6 ng/ml). Results: Ou r findings indicate that a 48-hour incubation with insulin augments th e proliferation of human infragenicular VSMC, producing a 207% increas e in cell number when compared with control cells (11,328 +/- 686, n = 56 vs 3682 +/- 182, n = 87; p < 0.0001). The addition of TGF beta 1 i n combination with insulin abolished the accelerated growth rate seen in test groups treated with insulin alone (3614 +/- 247, n = 32 vs 11, 328 +/- 686, n = 56; p < 0.0001). Conclusion: These results strongly s uggest that insulin is a potent stimulant of human infragenicular VSMC proliferation. The mitogenic effect of insulin is inhibited by TGF be ta 1, producing proliferation rates comparable to those observed in co ntrol cells incubated with serum-free media.