ABSTRACT
Objective_To investigate the effects of hypoxia-inducible factor-1 alpha ( HIF-1α) inhibitor YC-1 on hy-poxia induced human pulmonary artery smooth muscle cells ( HPASMCs) proliferation, apoptosis and the expression of P53, and to explore the molecular mechanism in the processes.Methods_HPASMCs were cultured in DMEM me-dium supplemented with 10%FBS in vitro.Then divided them into four groups:normoxia, hypoxia and hypoxia+YC-1(0.01 and 0.05 mmol/L).Cell proliferation was measured by CCK-8 and apoptosis was detected by flow cytom-etry.The expressions of HIF-1αand P53 were tested by Western blot, and the mRNA expression of P53 was tested by RT-PCR.Results_Hypoxia can promote the proliferation of HPASMCs.Treatment of HPASMCs with different concentrations of YC-1 intervention for 24h obviously dropped proliferation rate (P<0.05), and the apoptosis rate increased significantly (P<0.05).YC-1 can also down-regulate the expression of HIF-1αand up-regulate the ex-pression of P53 significantly ( P<0.05 ) .Conclusions_YC-1 can inhibit hypoxia-induced HPASMCs proliferation and promote apoptosis, the mechanism is potentially related to the up-regulation of P53 expression.
ABSTRACT
AIM: To find out the mechanism of focal adhesion kinase(FAK) facilitating human pulmonary artery smooth muscle cells(HPASMCs) proliferation.METHODS: HPASMCs were isolated from normal part of lungs of two carcinoma patients who undergone lung partial resection.Cultured HPASMCs stimulated by fibronection(40 mg/L) were passively transfected with ODNs,sense focal adhesion kinase(FAK),mismatch sense and antisense-FAK,respectively.Expression of FAK,Jun NH2-terminal kinase(JNK) and cyclin-dependent kinase2(CDK2) proteins were detected by immunoprecipitation and Western blotting.Cell cycle and cell apoptosis were analyzed by flow cytometry.In addition,cytoplasma FAK expression was detected by immunohistochemistry staining.RESULTS: The protein expressions of FAK,JNK and CDK2 in HPASMCs decreased in FAK ASODNs group and increased in FAK SODNs group.Meanwhile,the proportion of cells at G1 phase decreased significantly in FAK SODNs group,while the cells at S phase increased significantly.In contrast,the proportion of cells at G1 phase was increased significantly in FAK ASODNs group.The level of cell apoptosis in FAK ASODNs group was higher.FAK expression in FAK SODNs group was strongly stained by immunocytochemistry,whereas that in FAK ASODNs group was weakly stained.CONCLUSION: The results suggest that FAK via JNK,CDK2 signaling pathway enhances HPASMCs proliferation.