Characterization of Insulin Binding Sites in Cultured Smooth Muscle Cells of Rat Aorta

Summary

Insulin receptors could be demonstrated in cultured smooth muscle cells of rat aorta. The specific binding of ¹²⁵I-insulin was time-, temperature- and pH-dependent. The optimal temperature for our studies was 12°C. At this temperature maximal specific binding was 0.5% of total counts at 120 min incubation. The pH-optimum for the binding process was between 7.5 and 8. Degradation of ¹²⁵I-insulin at 12°C was 14%, no degradation of binding sites could be measured at this temperature. Dissociation of ¹²⁵I-insulin was rapid. 50% of the labeled hormone remained associated with the cells. Half-maximal inhibition of ¹²⁵I-insulin binding was produced by insulin at 4 × 10^-11 mol/l. Scatchard-analysis gave curvilinear plots, that may suggest negative cooperativity. Specificity of binding was studied in competition experiments between ¹²⁵I-insulin, insulin, proinsulin, insulin-like growth factors and human growth hormone. Half-maximal inhibition of ¹²⁵I-insulin binding was produced by proinsulin at 2 × 10^-9 mol/l and by insulin-like growth factors at 9 × 10^-9 mol/l. Human growth hormone had no significant effect on the insulin binding.

Key-Words:

Insulin Receptors - Smooth Muscle Cells - Atherosclerosis