Objective To investigate the effects of Visfatin on glycolipid metabolism and insulin resistance in the skeletal muscle of diabetic KKAy mice, preliminarily analyze the possible mechanism of action through the PI3K/ Akt / FoxO1 pathway. Methods Eight-week-old male C57BL/ 6J mice were randomly divided into a normal diet (ND) group, a normal diet+Visfatin (ND+V) group, a high-fat (HFD) group, and a high-fat+Visfatin (HFD+V) group. When the model was generated, male KKAy mice of the same age were randomly divided into a diabetes mellitus (DM) group and diabetes mellitus + Visfatin (DM+V) group. Among them, the ND+V, HFD+V and DM+V groups were intraperitoneally injected with recombinant Visfatin protein for 3 consecutive days. Fasting blood glucose (FBG), postprandial blood glucose(PBG), total cholesterol (TC), triglyceride (TG), free fatty acids(FFA) and fasting insulin(FIns) were detected in each group of mice. The glucose tolerance test (GTT) and insulin tolerance test (ITT) were measured; meanwhile, the area under curve (AUC) and homeostasis model assessment for insulin resistance (HOMA-IR) were calculated. At the same time, the expression levels of phosphatidylinositol 3 kinase (PI3K), protein kinase B (Akt), forkhead box transcription factor 1 (FoxO1), pyruvate dehydrogenase kinase 4 (PDK4), phosphoenolpyruvate carboxykinase (PEPCK) and other related molecules were detected by RT-qPCR and Western blot. Results Compared with the ND group and the HFD group, the body weight, food intake, FBG, PBG, TC, TG, FFA, FIns, AUC_GTT , AUC_ITT and HOMA-IR levels in the DM group were significantly increased (P< 0. 05). Compared with DM group, FBG, TC, TG, AUC_GTT , AUC_ITT , and HOMA-IR levels in DM+V group were significantly reduced (P<0. 05). Compared with the ND group and the HFD group, the mRNA expression levels of P13K and Akt in the DM group were significantly decreased (P<0. 001), but the mRNA expression levels of FoxO1 were increased (P<0. 05). Compared with the DM group, P13K and Akt mRNA expression in the DM+V group was obviously increased, but the mRNA expression of FoxO1 was significantly reduced (P<0. 01). Compared with the ND group, PI3K_110α and p-Akt expression in the DM group was reduced, and the protein expression levels of FoxO1, p-FoxO1 and PDK4 were increased. Compared with the HFD group, p-Akt and p-FoxO1 expression was lower in the DM group, and the protein expression levels of FoxO1, PDK4 and PEPCK were increased (P<0. 05). While compared with the DM group, PI3K_110α , p-Akt and p-FoxO1 expression were increased in the DM+V group, but the protein expression levels of FoxO1, PDK4 and PEPCK were reduced (P<0. 05). Conclusions Visfatin may downregulate PDK4 expression in the skeletal muscle of diabetic mice through the PI3K/ Akt / FoxO1 pathway and has a role in improving glucolipid metabolism and insulin resistance.