Background: Periostin mediates inflammation and fibrosis by regulating ECM adhesion, migration, and differentiation in multiple organ diseases. Studies have shown Periostin mainly located in the dilated mesangium, tubulointerstitial and fibrotic regions of the diabetic nephropathy, which is negatively correlated with renal function. However, the underlying mechanisms remain poorly explored. Methods: The expression of periostin in HK-2 cells was investigated under high glucose and high concentration of TGF-β1. The signaling pathway of periostin involved in epithelial-mesenchymal transdifferentiation of HK-2 cells was also validated. The expression of periostin were investigated by RT-PCR with different concentration of glucose and TGF-β1. Periostin expression in HK-2 cells stimulated by high glucose and high TGF-β1 was detected by western blotting and immunofluorescence. The expression of E-Cad, α-SMA and p38 proteins were also detected. The effects of p38 inhibitors on periostin, E-Cad and α-SMA in high glucose environment were also investigated and verified. Results: High concentration of glucose and TGF-β1 increased the fibrosis of HK-2 cells, prompting high expression of periostin. The combined stimulation of high glucose (22 mmol/L) and high TGF-β1(10 ng/mL) upregulated the expression of periostin obviously, stimulating the expression of α-SMA and p38 while inhibiting the secretion of E-Cad. More importantly, p38 inhibitors reduced the expression of periostin and α-SMA protein while promoted E-Cad protein expression in HK-2 cells under high glucose conditions. Furthermore, p38-MAPK signal pathway was involved in the epithelial-mesenchymal transition of human renal tubules in high glucose environment. Conclusion: Periostin, a downstream factor of TGF-β1, is positively regulated by TGF-β1 under high glucose condition, affecting the epithelial-interstitial differentiation of HK-2 cells via the p38-MAPK signaling pathway. Therefore, periostin may serve as a biomarker of renal fibrosis in diabetic kidney disease.