Diabetic nephropathy (DN) is one of the most serious complications of diabetes mellitus (DM). Recent researches show that DNA methylation plays a role in DN. However, the exact mechanism is not fully understood. MicroRNAs (miRNAs) are a group of endogenous non-coding small RNAs that are involved in the regulation of the development of DN. We have previously demonstrated that let-7a was down-expressed in DN by microarray, but the mechanism is unclear. In this study, let-7a-3 was found to be the only gene with the CpG island in the promoter region among the three let-7a members (let-7a-1, let-7a-2 and let-7a-3) by bioinformatic methods. Also, the expression levels of three homologues of let-7a were tested by real-time PCR, and DNA methylation of the let-7a-3 gene in the promoter region was analyzed by quantitative methylation-specific PCR (qMSP) in 60 individuals, with 20 cases in the control (CON), DM and DN groups respectively. Additionally, the target gene of let-7a-UHRF1 was proved by bioinformatic analysis and dual-luciferase reporter assay. Results showed that let-7a-3 was down-regulated in DN patients. Moreover, qMSP data showed that the average methylation ratio of the let-7a-3 promoter in the DN group was significantly higher than that in the CON and DM groups (P<0.05). Data also showed that let-7a negatively regulated the mRNA and protein expressions of methylation-related gene-UHRF1 through UHRF1 3'UTR. And the expressions of UHRF1 and DNMT1 were increased in DN patients. Therefore, we concluded that promoter hypermethylation and down-expression of let-7a-3 may play a role in DN by targeting UHRF1.