The aim of this study was to investigate the mechanism of oridonin (ORI) in ameliorating non-alcoholic fatty liver disease (NAFLD). Oleic acid (OA) was used to induce NAFLD model in HepG2 and Huh-7 cells, and the effects of ORI on OA-induced cellular steatosis were evaluated by using the CCK-8 cell viability assay, oil red O staining, and total cholesterol (TC) and triglyceride (TG) content assays. Potential targets and action pathways for ORI in the treatment of NAFLD were explored through a network pharmacology approach. The mechanism of ORI to improve NAFLD was verified with HepG2 and Huh-7 cell steatosis models. The experimental results showed that ORI at concentrations of 5 and 10 μmol/L could improve the lipid accumulation in the steatosis models of HepG2 and Huh-7 cells, and significantly reduce the lipid content as well as the contents of total cholesterol (TC) and triglyceride (TG) in the steatosis models. Ninety-five targets of ORI and NAFLD were screened, and GO enrichment analysis showed that these genes were mainly involved in lipopolysaccharide response, fatty acid binding, and cholesterol homeostasis, and the results of KEGG pathway analysis indicated that PPAR signaling pathway might be a key pathway for the amelioration of NAFLD by ORI. The results of mechanism validation showed that the expression of PPARα, CPT-1 protein and mRNA was elevated after intervention with ORI (P<0.05, P<0.01), and the effect of ORI on PPARα and CPT-1 protein expression disappeared after treatment with the PPARα inhibitor MK886. In conclusion, ORI had an ameliorating effect on OA-induced NAFLD model in HepG2 and Huh-7 cells, and its mechanism of action was related to the PPARα/CPT-1 pathway.