转染Kv1.5基因抑制人气道平滑肌细胞的增殖

Transfection of human Kv1.5 gene inhibits the proliferation of human airway smooth muscle cells

目的:转染Kv1.5基因对人气道平滑肌细胞(HASMCs)增殖及凋亡的影响。方法:通过脂质体介导瞬时转染Kv1.5基因于培养的HASMCs中,以转染空载体pRc/CMV的细胞及未转染质粒的细胞为对照;用Westernblotting检测平滑肌细胞Kv1.5蛋白表达;用荧光光度法检测HASMCs胞内钙浓度;用流式细胞术观察细胞周期;用MTT法检测HASMCs增殖及DNA末端转移酶介导的原位缺口末端标记技术(TUNEL)检测细胞的凋亡。结果:(1)转染质粒组Kv1.5蛋白质的表达明显高于未转染组及空载体转染组(P<0.01);(2)转染质粒组细胞胞内钙浓度明显低于未转染组及空载体转染组(P<0.05),且其细胞周期中的G0/G1期细胞比例明显高于、细胞增殖率显著低于未转染组及空载体转染组(P<0.01);同时,转染质粒组细胞的凋亡率明显高于未转染组及空载体转染组(P<0.01)。结论:转染Kv1.5基因能抑制HASMCs的增殖、促进其凋亡,为进一步探讨哮喘气道重塑的机制及其治疗提供实验依据。

AIM： To investigate the effect of human Kv1. 5 gene transfection on the proliferation and apoptosis in human airway smooth muscle cells （HASMCs）. METHODS： Kv1. 5 gene was transiently transfected into HASMCs with Lipofectamine 2000 and the level of Kv1. 5 protein was observed by Western blotting. Ca2 ＋ concentration in HASMCs was investigated by fluorescent quantitation using fluorospectrophotometer. Flow - cytometry, MTr method and TUNEL were used to detect the effects of Kv1. 5 gene transfection on proliferation and apoptosis in HASMCs. RESULTS ： （ 1 ） Western blotting showed that transfection of human Kv1. 5 plasmid significantly increased the Kv1. 5 protein expression compared to control group （P 〈0. 01 ）. （2） There was a decrease in Ca^2＋ concentration and the numbers of G0/G1 HASMCs in Kv1. 5 gene transfection group compared to control group （P 〈0. 05 ）. The proliferation rate in Kv1. 5 gene transfection group was significantly decreased, and the apoptosis rate in Kv1. 5 gene transfection group was significantly increased compared to control group （P 〈0. 01 ）. CONCLUSION： Transfection of human Kv1. 5 plasmid inhibits proliferation and increases apoptosis in HASMCs, which provides the experimental evidence for investigating the mechanism and treatment of airway remodeling in asthma.