Objective To investigate the role and mechanism of miR-214 targeting PTEN-mediated PI3K/AKT signaling pathway in reducing oxidative stress and renal interstitial fibrosis in diabetic nephropathy in rats. Methods SD rats were divided into control group (control group), model group (DN group), miR-214 negative control group (DN+miR-NC group) and miR-214 mimic group (DN+miR-214 mimics group). qRT-PCR was used to detect the expression of miR-214 in kidney tissue of each group of rats. The electronic balance was used to weigh the body weight and kidney weight of each group of rats, and calculates the renal index. Blood glucose meter was used to measure fasting blood glucose (FPG) of rats in each group. Biuret method was used to determine the 24-hour urine protein (UP) level of rats in each group. Automatic biochemical analyzer to detect serum creatinine (Scr) and serum urea nitrogen (BUN) levels in each group of rats. HE staining was used to observe the pathological changes of kidney tissue in each group of rats. Masson staining was used to observe the renal interstitial fibrosis in each group of rats. ELISA was used to detect the expression of malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), and glutathione peroxidase (GSH-PX) in the serum of each group of rats; Immunohistochemical method was used to detect the expression of TGF-β1 in kidney tissue of rats in each group. Bioinformatics tools was used to predict target genes. Dual luciferase reporters was used to detect interactions between genes or proteins; Western blot was used to detect the expression of PTEN, p-PI3K, p-AKT and p-mTOR proteins in the kidney tissues of rats in each group. Results miR-214 in renal tissue of DN rats was low expressed (P＜0.001), and significant up-regulated in renal tissue of DN+miR-214 mimics group rats (P＜0.001). miR-214 mimics increased body weight in rats, decreased renal index, FPG, UP levels, Scr levels, and the BUN level (all P＜0.05). In the DN+miR-214 mimics group, the infiltration of inflammatory cells in the renal interstitial tissue and the deposition of collagen fibers in the renal interstitial tissue were reduced. miR-214 mimics reduced MDA levels in rat serum, elevated SOD, GSH, GSH-PX levels (all P＜0.05), and TGF-β1 expression in rat kidney tissues was decreased (P＜0.05). There is a targeting relationship between miR-214 and PTEN. miR-214 mimics down-regulated PTEN protein expression in rat kidney tissues, and up-regulated p-PI3K, p-AKT, and p-mTOR protein expressions (all P＜0.05). Conclusion miR-214 is under-expressed in the kidney tissues of DN rats, but over-expression of miR-214 has a protective effect on the kidney tissues of DN rats, which may be related to reducing the level of oxidative stress in kidney tissues, the expression of TGF-β1 protein, and inhibiting PI3K/AKT pathway activation is involved.