Abstract: Objective: To observe the role of homocysteine（Hcy） and MMP-9 in prevention of diabetic nephropathy（DN）. Methods: Twenty-four Kkay mice were divided into three groups（n=8）:diabetes group（KA）,methionine-diet group（KB）,folic acid and vitamin B₁₂ treatment group（KC）.C57Bl/6 mice were used as controls（C57）.Four months later,the mice were sacrificed and their serum levels of Hcy,folic acid and vitamin B₁₂ were measured by fluorescence polarization immunoassay.Pathological changes in kidneys were examined with the periodic acid schiff（PAS） method with periodic acid-silver methenamine staining under electron microscope.Expression of MMP-9 protein or mRNA was detected by RT-PCR with immunohistochemical staining. Results: Different morphological changes of DN were found in Kkay mice.High Hcy levels induced by a diet enriched in methionine aggravated diabetic nephropathy and increased the expression level of MMP-9 protein and mRNA in glomerulus.The mean positive area of MMP-9 in KC group was lower than that in KB group（11.49% vs 15.90%,P＜0.05）,but had no difference between KC and KA groups（11.49% vs 11.14%,P＜0.05）. Conclusion: High levels of Hcy can worsen nephropathy,which can be effectively prevented by treatment with folic acid and vitamin B₁₂ to reduce the enhanced expression of MMP-9.