Aim: Effect of vitamin D supplementation on peripheral conversion of T4 into T3 in diabetic rats has not been previously investigated. The present study aimed to assess the effect of vitamin D3 administration on deiodinase 2 (D2) expression in diabetic rats. Methods: Thirty male Wistar rats (200-250 g) were included into; control, diabetic and diabetic supplemented with vitamin D3 groups. Diabetes was induced by single intraperitoneal injection of streptozotocin 45 mg/kg in citrate buffer. Vitamin D3 was administered orally in a dose of 500 IU/kg/day in corn oil for 10 weeks. Serum levels of free T4, free T3 and TSH were measured. Tissue homogenates from liver, kidney, muscle, femur bone, heart and brain were obtained and assessed for expression of D2 mRNA. Results: Diabetic rats demonstrated significant increase in free T4 and significant decrease in free T3 together with non significant increase in TSH level compared to control rats. However the changes in thyroid profile were ameliorated by vitamin D3 administration to diabetic rats. D2 mRNA expression was significantly reduced in all tissue homogenates obtained from diabetic rats compared to control rats. Vitamin D3 treated diabetic rats showed significantly enhanced D2 mRNA expression in liver and brain homogenates, while in remaining tissues; the increase in D2 mRNA did not reach significance. Conclusion: Diabetes mellitus inhibited peripheral conversion of T4 into T3 secondary to reduction in the expression of D2 in almost all body tissues. Vitamin D3 administration to diabetic rats greatly corrected the alterations in thyroid profile and D2 expression. The present results point to the possible beneficial role of vitamin D3 against the subclinical hypothyroid state that could associate diabetes mellitus.