Background and Aim Vascular smooth muscle cell (VSMC) apoptosis plays a critical role in the pathogenesis of many angiocalcified diseases. Recent studies have shown that lysine methyltransferase SET8 was involved in regulating cell proliferation and apoptosis. The purpose of this study was to explore whether SET8 can influence calcification by regulating proliferation and apoptosis of VSMC. Methods VSMC were obtained from rat thoracic aorta, and then randomly divided into control group, the empty plasmid group and SET8-shRNA group. Transfection was performed with cationic lipid vectors (LipofectamineTM2000) on VSMC. Calcium deposition was measured by alizarin red staining and calcium content measurement; The proliferation of VSMC in vitro was measured by MTT assay. The expressions of proliferation-related genes survivin and apoptosis-related genes caspase-3 were determined by real-time PCR and Western blot assay. Results The expression of SET8 protein in VSMC was effectively inhibited by SET8-shRNA. The calcium content was decreased in cells after SET8-shRNA transfection. Proliferation of VSMC was inhibited after transfected with SET8-shRNA 12 h, 24 h, 36 h and 48 h (P<0.05). The expressions of survivin was decreased in cells, but expressions of caspase-3 was increased after SET8-shRNA transfection (P<0.05 for all). Conclusion Interfering with SET8 can increase the expression of apoptosis gene and inhibit the expression of proliferative genes, and SET8 may be involved in regulating the calcification of rat blood vessels.