The additive effects of glucose and insulin on the proliferation of infragenicular vascular smooth muscle cells

Purpose: Peripheral vascular disease involving the infragenicular arterial tree is common in patients with diabetes mellitus (DM). Accelerated prolife ration of vascular smooth muscle cells (VSMCs) plays an important role in t he development of atherosclerosis. Insulin and glucose stimulate VSMC proli feration and are elevated in patients with non-insulin-dependent DM. We hav e previously described the mitogenic effect of insulin on VSMCs in vitro; t he effects of insulin and glucose separately and in combination on the prol iferation of VSMCs grown in serum-free media were studied. Methods: Human infragenicular VSMCs isolated from diabetic patients with en d-stage peripheral vascular disease undergoing below-knee amputation were u sed. Cells from passages 3 to 5 were grown in serum-free media with varying glucose (0.05%, 0.1%, 0.2%, 0.4%, 0.6%, and 0.8%) and insulin (no added in sulin, 100 ng/mL, and 1000 ng/mL) concentrations for 6 days. Results: Insulin stimulated VSMC growth at glucose concentrations more than 0.2% (0.4% glucose with no added insulin resulted in 13,073 +/- 336 cells/ mL, 0.4% glucose with 100 ng/mL insulin resulted in 16,536 +/- 1175 cells/m L, 0.4% glucose with 1000 ng/mL insulin resulted in 17,500 +/- 808 cells/mL , 0.6% glucose with no added insulin resulted in 14,167 +/- 1062 cells/mL, 0.6% glucose with 100 ng/mL insulin resulted in 18,984 +/- 1265 cells/mL, 0 .6% glucose with 1000 ng/mL insulin resulted in 20,450 +/- 1523 cells/mL, 0 .8% glucose with no added insulin resulted in 15,853 +/- 1650 cells/mL, 0.8 % glucose with 1000 ng/mL insulin resulted in 26,302 +/- 1919 cells/mL; P < .05 compared with glucose with no added insulin). Glucose stimulated VSMC proliferation up to a concentration of 0.2% (42% and 117% higher growth at 0.1% and 0.2% glucose, respectively, compared with the baseline, P < .05), regardless of the insulin concentration in the media. The greatest growth ( 26,302 +/- 1919 cells/mL) occurred in the group with the highest concentrat ion of both insulin (1000 ng/mL) and glucose (0.8% glucose; P < .05). Conclusion: Both insulin and glucose stimulate the growth of diabetic infra genicular VSMCs. The mitogenic effects of insulin and glucose are additive and may contribute to the development of atherosclerosis in patients with D M.