MiRNAs play important roles in initiation and progress of many pathologic processes. MiR-21 was closely associated with diabetic nephropathy (DN). However, whether serum miR-21 was as a potential diagnostic biomarker for DN and the relationship between serum miR-21 and tissue miR-21 remained unclear. In this study, real-time RT-PCR, cell transfection, luciferase reporter gene assays, western blot and confocal microscope were used, respectively. Here, we found that serum and renal tissue miR-21 was significantly elevated with the progress of DN. Moreover, luciferase reporter gene assays showed that smad7 was a validated miR-21 target, cell transfection showed that miR-21 overexpression downregulated target smad7 expression. Interestingly, serum miR-21 was significantly consistent with the alterations of tissue miR-21 with the development of DN. In addition, serum miR-21 was also positively correlated with GBM, GA, ACR and CCF, while negatively correlated with Ccr. Importantly, antagomiR-21 not only alleviated GBM, GA, ACR and CCF, but also ameliorated Ccr by increasing target smad7. In conclusion, our data demonstrated that serum miR-21 was closely associated with renal structure and function, and serum miR-21 may be regarded as a potential diagnostic biomarker of DN.