RNA干扰沉默STIM1表达对低氧下大鼠肺动脉平滑肌细胞增殖的影响  

Effect of RNA interference targeting STIM1 gene on proliferation of hypoxic pulmonary artery smooth muscle cells in rat

作　　者：后显华[1,2] 陈建[1,2] 罗勇军[3] 刘福玉[2] 杨晓红[2] 袁志兵[1,2] 徐刚[2] 陈德伟[1,2] 高钰琪[1,2] 

机构地区：[1]第三军医大学高原军事医学系病理生理与高原生理学教研室,重庆400038 [2]第三军医大学教育部高原医学重点实验室,全军高原生理与高原病研究重点实验室,重庆400038 [3]第三军医大学高原疾病学教研室,重庆400038

出　　处：《第三军医大学学报》2011年第5期437-440,共4页Journal of Third Military Medical University

基　　金：“十一五”国家科技支撑计划(2009BAI85B01);国家重点基础研究发展计划(973计划,2006CB504101)~~

摘　　要：目的应用RNA干扰抑制肺动脉平滑肌细胞(pulmonary aretery smooth muscle cells,PASMCs)基质交感分子1(stromal interaction molecule 1,STIM1)基因表达,探讨STIM1表达抑制后对PASMCs缺氧增殖反应的影响。方法采用离体培养大鼠肺动脉平滑肌细胞,设对照组、阴性siRNA组(SiNT)及阳性siRNA组(SiSTIM1),低氧(3%O2)培养24 h,Western blot法检测PASMCs中STIM1基因表达变化,MTT比色法及[3H]-TdR掺入法检测PASMCs的增殖情况,流式细胞仪检测PASMCs的细胞周期变化。结果 Western blot检测SiSTIM1组STIM1蛋白相对灰度值为(0.355±0.008),显著低于对照组(0.624±0.017)和SiNT组(0.618±0.021)(P<0.05)。MTT比色实验及[3H]-TdR掺入实验显示低氧培养24 h后,对照组和SiNT组PASMCs较常氧组增殖显著(P<0.05),而SiSTIM1组PASMCs增殖活性较对照组和SiNT组显著降低(P<0.05)。流式细胞仪检测结果表明低氧培养24 h后,对照组和SiNT组PASMCs G0/G1期细胞比例较常氧组显著降低(P<0.05),而SiSTIM1组G0/G1期细胞比例则显著高于对照组和SiNT组(P<0.05)。结论 RNA干扰介导STIM1基因沉默可明显抑制低氧下大鼠肺动脉平滑肌细胞增殖。Objective To inhibit the expression of stromal interaction molecule 1（STIM1） gene of pulmonary artery smooth muscle cells（PASMCs） through RNA interference,and to explore the proliferation change of PASMCs under hypoxia condition after gene silencing.Methods In vitro cultured PASMCs were divided into a blank control group,a negative control siRNA group（SiNT） and a positive siRNA group（SiSTIM1） that were cultured in hypoxia condition（3% O2） for 24 h,as well as a normal group that was cultured in normal O2 condition（21% O2） for 24 h.Western blotting,MTT assay and -TdR incorporation assay,and flow cytometry were employed to test STIM1 gene expression,cell proliferation,and cell cycle of PASMCs,respectively.Results Western blotting results showed that the relative grey value of STIM1 protein in the SiSTIM1 group（0.355±0.008） was remarkably lower than that in the control group（0.624±0.017） and the SiNT group（0.618±0.021）（P0.05）.MTT assay and -TdR incorporation assay exhibited that,after 24 h hypoxic culture,the PASMCs in the blank control group and the SiNT group proliferated significantly as compared with those in the normal group（P0.05） and the SiSTIM1 group（P0.05）.Flow cytometry results discovered that,after hypoxic culture for 24 h,the percentages of PASMCs in G0/G1 phase in the blank control group and the SiNT group were remarkably lower than those in the normal group（P0.05） and the SiSTIM1 group（P0.05）.Conclusion RNA interference-mediated STIM1 gene silencing can significantly inhibit the proliferation of PASMCs under hypoxia.

关 键 词：基质交感分子1 RNA干扰 肺动脉 平滑肌细胞 缺氧 细胞增殖 

分 类 号：R322.121[医药卫生—人体解剖和组织胚胎学] R364.4[医药卫生—基础医学]