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目的探讨异钩藤碱对血小板源性生长因子BB(PDGF-BB)诱导的肺动脉平滑肌细胞(PASMCs)增殖的影响及其机制。方法建立PDGF-BB诱导的PASMCs增殖模型,通过MTT比色法和台盼蓝拒染法检测异钩藤碱对PASMCs增殖的影响及细胞毒性,流式细胞术检测细胞周期变化,Western blotting检测细胞周期相关蛋白的表达及血小板源性生长因子受体β(PDGF-Rβ)信号通路蛋白表达及磷酸化的影响。结果与正常对照相比,加入PDGF-BB(20 ng/m L)培养24 h,可促进PASM Cs增殖约2.5倍;异钩藤碱剂量依赖性地抑制PDGF-BB诱导的PASM Cs增殖;异钩藤碱(25μmol/L)明显抑制PDGF-BB诱导的S期细胞比例升高;Western blotting结果显示,异钩藤碱可抑制PDGF-BB诱导的cyclin D1、CDK6的表达和p27kip1的降解,下调PDGF-Rβ及其下游信号分子AKT、糖原合成激酶(GSK)3β的磷酸化。结论异钩藤碱通过将细胞周期阻滞于G0/G1期来抑制PDGF-BB诱导的PASMCs增殖,其机制可能为通过抑制PDGF-Rβ及其下游信号通路的磷酸化、进而调节细胞周期相关蛋白的合成和降解来实现。
Objective To investigate the effects and mechanisms of isorhynchophylline on the proliferation of pulmonary artery smooth muscle cells (PASMCs) induced by platelet-derived growth factor BB (PDGF-BB). Methods The proliferation model of PASMCs induced by PDGF-BB was established. The proliferation and cytotoxicity of isorhynchophylline on PASMCs were detected by MTT colorimetric assay and trypan blue exclusion assay. The cell cycle was detected by flow cytometry. Expression of Cyclin D1, Cyclin D1, Cyclin D1, Cyclin D1, Cyclin D1 and Cyclooxygenase - 2 in Peripheral Blood and the Expression of PDGF - Rβ Signaling Pathway and Phosphorylation of. Results Compared with the normal control group, PDGF-BB (20 ng / mL) cultured for 24 h promoted the proliferation of PASM Cs by about 2.5-fold. Isoproterenol inhibited the proliferation of PASM-Cs induced by PDGF-BB in a dose- Rhynchophylline (25μmol / L) significantly inhibited the proportion of S phase cells induced by PDGF-BB. Western blotting showed that isorhline inhibited the expression of cyclin D1, CDK6 and p27kip1, and down-regulated PDGF-Rβ and its downstream signaling molecule AKT, glycogen synthesis kinase (GSK) 3β phosphorylation. Conclusion The inhibition of PDGF-BB-induced proliferation of PASMCs by isorhynchophylline may be through the inhibition of PDGF-Rβ and its downstream signaling pathway phosphorylation by blocking the cell cycle in G0 / G1 phase, and then regulating the expression of cell cycle related proteins The synthesis and degradation to achieve.