Mechanisms Of Diabetic Vascular Complications

Objestives To observe the changes of platelet nitric oxide synthase (NOS) activity in diabetic patients and effects of advanced glycation end products (AGEs) on platelet NOS activity of normal subjects and its mechanisms.Methods (1) Peripheral venous blood was collected from eight diabetic patients, platelets were isolated with gel-filtration chromatography, NOS activity was measured as formation of 3H-L-citrulline from 3H-L-arginine. (2) Platelets from six normal subjects were prepared and incubated with different concentrations of AGEs (100-400 ug/ml) for 30 minutes, then platelets were divided into two aliquots, one was used for NOS activity measurements, and the other for purifying eNOS with immuno -precipitation method. Both eNOS expression and eNOS protein phosphorylation were detected by western blotting.Results (l)Platelet NOS activity was significantly decreased in diabetic patients as compared with normal subjects(p<0.01), it was still142 %lower after histamine stimulation(p<0.05). (2) AGEs significantly inhibited platelet NOS activity in a concentration-dependent manner. (3) eNOS was highly expressed in platelets. The expression of eNOS and serine phosphorylation of eNOS protein in platelet had no significant reduction after platelets were incubated with AGEs for 30 minutes.Conclusion: (l)Platelet NOS activity of diabetic patients was reduced; (2)AGEs inhibited platelet eNOS activity; (3)The expression and serine phosphorylation of eNOS protein in platelet had no significant reduction after platelets were incubated with AGEs for 30 minutes.