论文部分内容阅读
目的 研究诱导型一氧化氮合酶 (iNOS)基因转染对缺氧条件下大鼠肺动脉平滑肌细胞 (PASMCs)增殖的影响 ,探讨细胞周期蛋白依赖性激酶抑制因子 p2 1、p2 7在细胞增殖过程中的调控作用。方法 经脂质体介导将iNOS重组逆转录病毒载体 (pLNCX/iNOS)转染大鼠PASMCs,检测外源性iNOS表达及其生物学活性 ;氚标记胸腺嘧啶脱氧核苷 (3 H TdR)掺入、流式细胞技术观察iNOS转染对缺氧条件下大鼠PASMCs增殖的影响 ;逆转录 聚合酶链反应 (RT PCR)和流式细胞技术检测p2 1、p2 7的变化。 结果 转染后检测证实 ,外源性iNOS基因可有效转录、表达 ,具有生物学活性 ;iNOS转染显著抑制缺氧条件下大鼠PASMCs3 H TdR的掺入 ,缺氧组 (D组 )大鼠每分钟衰变数值为 (180 11± 2 5 2 1)次 /min ,缺氧 +DETANONOate组 (E组 ,0 5、1mmol/L)每分钟衰变数值分别为(15 36 4± 1382 )次 /min、(13712± 1782 )次 /min、低氧 +iNOS转染组 (G组 )大鼠每分钟衰变数值为(15 14 5± 15 14 )次 /min ,3组比较差异有显著性 (P <0 0 1) ;iNOS转染导致停滞于G0 /G1期的细胞比例增加 ,G组G0 /G1期细胞百分比为 6 7 8% ,与D组 (46 8% )比较差异也有显著性 (P <0 0 1) ;iNOS转染可使低氧条件下PASMCs的P2 7蛋白表达下调受到抑制 ,P2 7蛋白质表达相
Objective To investigate the effect of iNOS gene transfection on the proliferation of pulmonary artery smooth muscle cells (PASMCs) in rats under hypoxia and explore the role of iNOS gene transfection in the process of cell proliferation In the regulatory role. Methods The iNOS recombinant retroviral vector (pLNCX / iNOS) was transfected into rat PASMCs by liposome, and the expression of iNOS and its biological activity were detected. Tritiated thymidine (3 H TdR) Flow cytometry was used to observe the effect of iNOS transfection on the proliferation of rat PASMCs under hypoxic conditions. The changes of p21 and p27 were examined by reverse transcription polymerase chain reaction (RT PCR) and flow cytometry. Results After transfection, the exogenous iNOS gene could be effectively transcribed, expressed and biologically active. INOS transfection significantly inhibited the incorporation of 3 H TdR into PASMCs under hypoxia. The hypoxia group (group D) The decay value per minute was (180 11 ± 2 5 2 1) times / min. The decay values per minute for the hypoxic + DETANONOate group (group E, 0, 5 and 1 mmol / L) were (15 36 4 ± 1382) , (13712 ± 1782) times / min, and the decay value per minute in hypoxia + iNOS transfection group (G group) was (15 14 5 ± 15 14) times / min, the difference was significant among the three groups (P < The percentage of cells in G0 / G1 phase in G group was 67.8%, which was also significantly different from that in group D (46.8%) (P <0.01). The percentage of cells arrested in G0 / 0 0 1). The iNOS transfection inhibited the down-regulation of P2 7 protein expression in PASMCs under hypoxic conditions. The expression of P2 7 protein