Aim To explore the effect of miR-301a on the expression of inflammatory factors in macrophages, from the perspective of microRNA to clarify the pathogenesis of atherosclerosis, then provide a new way of prevention and treatment of atherosclerosis. Methods ApoE－/－ mice was fed with high fat diet to establish atherosclerosis model, the expression of inflammatory factors in the atery were analyzed by real-time PCR. MiR-301a mimic and miR-301a inhibitor were transfected into RAW264.7 cells, the expression of inflammatory factors was analyzed by real-time PCR, the protein level of NF-κB repressing factor (NKRF) was measured by Western blot, the cellular location of p65 was determined by immunofluorescence. Results The level of miR-301a was increased in ateries of ApoE－/－ mice. Overexpression of miR-301a decreased the protein levels of NKRF and increased the mRNA levels of inflammatory factors, p65 was located in nuleus in the RAW264.7 cells transfected with miR-301a mimic. Inhibition of miR-301a increased the protein levels of NKRF and decreased the mRNA levels of inflammatory factors, p65 was located in cytoplasm in the RAW264.7 cells transfected with miR-301a inhibitor. Conclusion MiR-301a regulates the mRNA expression of TNF-α, IL-6 and MCP-1 in RAW264.7 cells via targeting NKRF to regulate the cellular location of p65.