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目的探讨腺病毒介导的反义血管紧张素Ⅱ(AngⅡ)1型受体(AT1R)cDNA转染对培养的人肺动脉平滑肌细胞(PASMC)迁移、增殖和凋亡的影响。方法构建重组反义人AT1R腺病毒(AdCMVahAT1)和表达β半乳糖苷酶的对照腺病毒载体AdCMVLacZ,将培养的PASMC分为DMEM组、AdCMVLacZ组和AdCMVahAT1组,分别给予不同的干预因素干预PASMC,用半定量逆转录聚合酶链反应(RT PCR)和免疫组化及彩色图像分析法检测AT1R的表达情况,并用改良的迁移小室进行细胞迁移实验。又将培养的PASMC分为DMEM组,AngⅡ组,AdCMVLacZ+AngⅡ组和AdCMVahAT1+AngⅡ组,用流式细胞仪检测各组PASMC的增殖指数和凋亡率,绘制DNA合成直方图。结果转染AdCMVahAT1后48h的PASMC,AT1RmRNA明显低于对照组[AT1R/βactin吸光度(A)比值:AdCMVahAT1组为0.48,DMEM组为0.97,AdCMVLacZ组为0.96];AdCMVahAT1组AT1R蛋白表达水平(A值为176.39±21.63)显著低于DMEM组(A值为310.21±29.82)和AdCMVLacZ组(A值为306.45±30.09),差异均有统计学意义(P均<0.01),转染AdCMVahAT1后24h的PASMC迁移距离为(40.93±9.69)μm,显著短于DMEM组的(78.23±11.79)μm和AdCMVLacZ组的(77.80±10.66)μm,差异均有统计学意义(P均<0.01)。给予AngⅡ刺激48h,AngⅡ组PASMC的增殖指数(59.69±3.46)显著高于
Objective To investigate the effect of adenovirus mediated transfection of AT1R cDNA on the migration, proliferation and apoptosis of cultured human pulmonary artery smooth muscle cells (PASMCs). Methods AdCMVahAT1 recombinant adenovirus and AdCMVLacZ adenovirus vector expressing β-galactosidase were constructed. The cultured PASMC were divided into DMEM group, AdCMVLacZ group and AdCMVahAT1 group. Different interventions were given to PASMC, The expression of AT1R was detected by semi-quantitative reverse transcriptase-polymerase chain reaction (RT PCR), immunohistochemistry and color image analysis, and the cell migration assay was performed with a modified migration chamber. The cultured PASMCs were further divided into DMEM group, AngⅡgroup, AdCMVLacZ + AngⅡgroup and AdCMVahAT1 + AngⅡgroup. Proliferation index and apoptosis rate of PASMC in each group were detected by flow cytometry, and DNA synthesis histogram was drawn. Results The expression of AT1R mRNA in AdCMVahAT1 group was significantly lower than that in control group at 48h after transfection of AdCMVahAT1 (AT1R / βactin absorbance (A) ratio: 0.48 in AdCMVahAT1 group, 0.97 in DMEM group and 0.96 in AdCMVLacZ group) (176.39 ± 21.63) was significantly lower than DMEM group (A value 310.21 ± 29.82) and AdCMVLacZ group (A value was 306.45 ± 30.09), the difference was statistically significant (P all <0.01), AdCMVahAT1 24h after transfection of PASMC The migration distance was (40.93 ± 9.69) μm, significantly shorter than that in the DMEM group (78.23 ± 11.79) μm and (77.80 ± 10.66) μm in the AdCMVLacZ group (all P <0.01). After stimulation with AngⅡ for 48h, the proliferation index of PASMC in AngⅡ group (59.69 ± 3.46) was significantly higher than that in AngⅡ group