IGF-I but not insulin inhibits apoptosis at a low concentration in vascular smooth muscle cells

Abstract

Apoptosis of vascular smooth muscle cells (VSMCs) is of importance in the development of diabetic angiopathy. Our aim was to evaluate the effect of insulin and IGF-I on apoptosis in VSMCs. Rat aortic VSMCs were used and apoptosis was induced by serum starvation. As apoptotic markers we measured caspase-3 activity, histone-associated DNA fragments by ELISA and nuclear morphology by DAPI (4',6-diamidino-2-phenylindole) staining. Phosphorylation of IGF-I receptors was evaluated by Western blot. Serum starvation had increased caspase-3 activity even after 3 h. The highest activity was found after 3-12 h. IGF-I 10(-9 )M inhibited serum starvation-induced caspase-3 activity with a maximal effect after 12 h. When studied after starvation for 12 h, significant inhibitory effects on caspase-3 were found at IGF-I concentrations of 10(-8)-10(-7) M (P<0.01) and at an insulin concentration of 10(-6 )M (P<0.01). DNA fragmentation was detected by ELISA after 24 h and chromatin condensation and nuclear fragmentation by DAPI staining after 24 and 48 h respectively. IGF-I dose-dependently reduced apoptosis evaluated by ELISA, reaching a maximal effect at 10(-9) M. Insulin reduced apoptosis but the effect was weaker and a higher concentration was needed. IGF-I (10(-8 )M) and insulin at a very high concentration (10(-6) M) phosphorylated IGF-I receptors. Taken together, IGF-I and insulin have antiapoptotic effects on VSMCs but the effect of insulin is only found at high unphysiological concentration.