【摘要】 目的 探讨缺氧诱导因子-1α(HIF-1α)调控经典型瞬时受体电位通道6(TRPC6)在新生大鼠缺氧性肺动脉高压(HPH)肺血管重塑中的作用机制。方法 将32只Wistar新生大鼠随机分为常氧组、HPH组、过表达HIF-1α+HPH组(HIF-1α组)、HIF-1α抑制剂+HPH组(2ME组)。常氧组在常氧环境下饲养，其余3组建立HPH模型，其中HIF-1α组大鼠通过尾静脉注射携带HIF-1α基因的腺病毒载体，2ME组每日皮下注射2-甲氧基雌二醇(2ME),各组以干预处理后14 d作为实验终点。直接测压法测量右心室收缩压(RVSP);称重计算右心室肥厚指数(RVHI);苏木精-伊红染色观察肺远端小动脉形态，计算血管重塑指标(MT%、MA%)进行定量分析；免疫组化法检测肺组织增殖细胞核抗原(PCNA)表达以评估肺血管平滑肌细胞增殖水平，同时检测HIF-1α、TRPC6蛋白在肺组织中的表达水平；实时荧光定量-聚合酶链反应法检测HIF-1α、TRPC6、PCNA的转录水平。结果 与常氧组比较，HPH组新生大鼠RVSP、RVHI、MT%、MA%升高(P<0.05),肺组织HIF-1α、TRPC6、PCNA的蛋白表达和mRNA水平升高(P<0.05);与HPH组比较，HIF-1α组新生大鼠RVSP、RVHI、MT%、MA%升高(P<0.05),肺组织HIF-1α、TRPC6、PCNA的蛋白表达和mRNA水平升高(P<0.05);与HPH组比较，2ME组RVSP、RVHI、MT%、MA%降低(P<0.05),肺组织HIF-1α、TRPC6、PCNA蛋白表达水平降低(P<0.05),TRPC6、PCNA mRNA水平降低(P<0.05);与HIF-1α组比较，2ME组RVSP、RVHI、MT%、MA%降低(P<0.05),肺组织HIF-1α、TRPC6、PCNA蛋白表达和mRNA水平降低(P<0.05)。结论 慢性缺氧刺激下HIF-1α可能通过上调TRPC6,促进新生大鼠HPH肺血管重塑，该研究可能为新生儿HPH提供新的治疗靶点。更多还原

【Abstract】 Objective To investigate the mechanism by which hypoxia-inducible factor-1α(HIF-1α) regulates the classical transient receptor potential channel 6(TRPC6) to mediate pulmonary vascular remodeling in a neonatal rat model of hypoxic pulmonary hypertension(HPH). Methods A total of 32 neonatal Wistar rats were randomly divided into normoxia group, HPH group, overexpression of HIF-1α+HPH group(HIF-1α group) and HIF-1α inhibitor+HPH group(2ME group). The rats in the normoxia group were fed in normoxia environment, and the other 3 groups were established HPH model, among which the rats in the HIF-1α group were injected with adenovirus vector carrying HIF-1α gene through tail vein, and the rats in the 2ME group were injected 2ME every day. The experimental end point was 14 days after intervention. Right ventricular systolic pressure(RVSP) was measured using direct catheterization. Right ventricular hypertrophy index(RVHI) was calculated based on ventricular weight measurements. Morphology of distal pulmonary arterioles was examined by hematoxylin-eosin(HE) staining. Vascular remodeling indices(medial wall thickness percentage(MT%) and medial wall area percentage(MA%) were calculated for quantitative assessment.Immunohistochemistry was used to detect the expression of PCNA in lung tissues to evaluate the proliferation level of pulmonary vascular smooth muscle cells, and the expression levels of HIF-1α and TRPC6 proteins in lung tissue were also detected. Transcriptional levels of HIF-1α, TRPC6 and PCNA were quantified using quantitative real-time polymerase chain reaction(qRT-PCR). Results Compared with normoxia group, the RVSP, RVHI, MT% and MA% of neonatal rats in the HPH group increased(P<0.05), and the protein expression and mRNA levels of HIF-1α, TRPC6 and PCNA in lung tissue increased(P<0.05). Compared with the HPH group, the RVSP, RVHI, MT% and MA% of neonatal rats in the HIF-1α group increased(P<0.05), and the protein expression and mRNA levels of HIF-1α, TRPC6 and PCNA in lung tissue increased(P<0.05). Compared with the HPH group, the RVSP, RVHI, MT% and MA% of neonatal rats in the 2ME group decreased(P<0.05), and the protein expression levels of HIF-1α, TRPC6 and PCNA in lung tissue decreased(P<0.05), and the mRNA levels of TRPC6 and PCNA decreased(P<0.05). Compared with the HIF-1α group, the RVSP, RVHI, MT% and MA% of neonatal rats in the 2ME group decreased(P<0.05), and the protein expression and mRNA levels of HIF-1α, TRPC6 and PCNA in lung tissue decreased(P<0.05). Conclusion HIF-1α may promote pulmonary vascular remodeling in neonatal rats with HPH by up-regulating TRPC6 under chronic hypoxia stimulation, which may provide a new therapeutic target for neonatal HPH.更多还原