Objective To investigate the effects of paeonol (Pae) on the release of inflammatory factors from rat vascular endothelial cells (VECs) with lipopolysaccharide (LPS)-induced injury and the apoptosis of vascular smooth muscle cells (VSMCs), as well as whether Pae inhibits the apoptosis of VSMCs by affecting the release of inflammatory factors from VECs and regulating the p38 MAPK signaling pathway. Methods The tissue predigested adherent method was used for the primary culture of VECs and VSMCs. The Transwell chamber was used to establish a co-culture system of VSMCs and VECs. LPS was used to induce the injury of VECs. The MTT assay was used to measure the survival rate of cells; ELISA was used to measure the level of tumor necrosis factor-α (TNF-α) secreted by VECs; flow cytometry was used to measure the apoptosis rate of VSMCs; Western blot was used to measure the expression of proteins involved in the p38 MAPK signaling pathway and apoptosisrelated proteins in VSMCs. Results Compared with the normal control group, LPS significantly increased the level of TNF-α in VECs (P＜0.05),the expression of the p38 MAPK signaling pathway proteins and apoptosisrelated proteins (p-MKK3/6, p-p38, p53, and cleaved caspase-3) in VSMCs in the co-culture system (P＜0.05),and the apoptosis rate of VSMCs (P＜0.05).Compared with the LPS stimulation group, Pae significantly inhibited the secretion of TNF-α from VECs (P＜0.05)and significantly reduced the expression of the p38 MAPK signaling pathway proteins and apoptosisrelated proteins (p-MKK3/6, p-p38, p53, and cleaved caspase-3) in VSMCs in the co-culture system (P＜0.05)and the apoptosis rate of VSMCs (P＜0.05).Conclusion Pae can inhibit the release of TNF-α from VECs and the p38 MAPK signaling pathway and thus reduce the apoptosis of VSMCs.