Abstract: Objective:To perform a primary culture of mouse distal pulmonary artery smooth muscle cells (PASMCs) , and to investigate the effect of hypoxia on PASMCs proliferation and expression of classical protein kinase C (cPKC) isoforms.Methods:Primary culture of mouse distal PASMCs was performd. PASMCs of 4 to 8 passage were used in the experiment. Western blotting and immunofluorescence staining were used to examine the expression of cPKC isoforms in PASMCs. The PASMCs were then divided into normoxic group and hypoxic group, which were cultured in usual incubator (5% CO 2) and hypoxic incubator (3% O 2, 5% CO 2) , respectively. After 24 h, 48 h, and 72 h culture in normoxia and hypoxia, Brdu method was used to examine the changes in cell proliferation in the two groups; Western blotting and flow cytometry were used to examine the changes in expression of cPKC isoform proteins in PASMCs. Results:Western blotting and immunofluorescence staining showed that mouse PASMCs expressed cPKCα, cPKCβⅠ and cPKCβⅡ, but not cPKCγ. Immunofluorescence staining showed that, compared with the normoxia group, the ratio of Brdu-positive cells to DAPI-positive cells in PASMCs in the hypoxic group increased significantly at 24 h, 48 h, and 72 h of hypoxia (all P<0.05) , and was most significant at 72 h. Western blotting and flow cytometry showed that the expression of cPKCα, cPKCβⅠ, and cPKCβⅡ proteins in PASMCs of hypoxic group increased at 24 h, 48 h, and 72 h of hypoxia compared with the normoxic group (all P<0.05) . Conclusion:Hypoxia may induce abnormal proliferation of mouse distal PASMCs by regulating cPKCα, βⅠ and βⅡ signaling pathways.