Objective To observe the effects of 1,25-dihydroxyvitamin D3 on the expressions of transforming growth factor-β1(TGF-β1), fibronectin(FN),and vascular endothelial growth factor(VEGF) in rats with diabetic nephropathy(DN), and to elucidate the protective mechanism played by 1,25-dihydroxyvitamin D3. Methods DN models were estabolished by injecting streptozotoein ( STZ ) into male SD rats, which were divided into TGF-β1 overexpression group, TGF-β1 overexpression plus vitamin D3 group, TGF-β1 low-expression group, TGF-β1 low-expression plus vitamin D3 group, TGF-β1 normal-expression group, and TGF-β1 normal-expression plus vitamin D3 group. After 1,25-dihydroxyvitamin D3 treatment for 37 days, renal function and blood biochemical parameters were evaluated. The morphology and fibrosis of kidney tissues were observed. The expressions of TGF-β1, FN, and VEGF in kidney cortex were measured by immunohistochemistry, realtime PCR, and Western blotting. Results The levels of cholesterol, triglyceride, creatinine,plasma glucose, HbA1C , and 24 h urinary protein were lower in vitamin D3treated groups than those in corresponding control groups(P<0. 05). The degree of renal fibrosis was raised with the increased level of TGF-β1. Vitamin D3 treatment decreased the fibrosis in diabetic kidney. There were significant differences in the mRNA and protein expressions of TGF-β1 in three control groups(P<0. 05). With the increased levels of TGF-β1, the expressions of FN and VEGF were increased. The expressions of TGF-β1, FN, and VEGF were lowered by vitamin D3compared with the corresponding control groups(P<0. 05). Conclusion 1,25-dihydroxy-vitamin D3 may protect the renal tissure in diabetic rats via inhibiting the expressions of TGF-β1, FN, and VEGF in the kidney.