Objective: Endothelial nitric oxide syntase(eNOS)is involved in several functions playing important role in development of type 2 diabetes mellitus(T2DM), and insulin resistance(IR). It has been demonstrated that eNOS modulates peripheral and hepatic glucose metabolism and insulin secretion, playing important role in the evolution of IR and T2DM.The aim of this study was to examine the association between eNOS intron4 VNTR polymorphism and type 2 diabetes in an Iranian population. Materials and Methods: Patients with diabetes were diagnosed according to American Diabetes Association Criteria. 5cc of peripheral blood were collected in EDTA tubes from healthy controls and patients with type2 diabetes attending diabetes clinic.Then DNA was extracted from WBCs,using salting-out method and PCR was performed to determine allele and genotype frequencies for eNOS gene VNTR polymorphism. Results: A significant difference was found in genotype frequencies of eNOS polymorphism between patients and controls (aa+ab vs bb p=0.02 OR=2.0 95%CI; 1.05-3.96). Also allele a frequency was significantly increased in patients with diabetes compared with controls (p=0.007 OR= 2.1 95%CI; 1.19-4.08). Conclusion: We found a significant difference in distribution of eNOS polymorphic variants at both allele and genotype frequencies level in diabetic patients. A allele is a risc factor of type 2 diabetes in an Iranian population.