引用本文:曹建佳1,何 江2,李兴森3,陈柯宏1,盛 夏1,李文宾1,李美材1,黄 飚3,王德林1.shRNA 沉默YAP基因对人肾癌786-O细胞增殖和凋亡的影响[J].重庆医科大学学报,2014,38(6):753~758
shRNA 沉默YAP基因对人肾癌786-O细胞增殖和凋亡的影响
Influence of YAP gene silencing by shRNA on the proliferation and apoptosis of human renal cancer cell 786-O
DOI:
中文关键词:  肾细胞癌  Yes-相关蛋白  RNA干扰  慢病毒
英文关键词:renal cell cancer  Yes-associated protein(YAP)  RNA interference  lentivirus
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作者单位
曹建佳1,何 江2,李兴森3,陈柯宏1,盛 夏1,李文宾1,李美材1,黄 飚3,王德林1 1. 重庆医科大学附属第一医院泌尿外科重庆 4000162. 重庆医科大学附属大学城医院消化神经中心重庆 4013313. 重庆市綦江区人民医院泌尿外科重庆 401420 
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中文摘要:
      目的:探讨慢病毒干扰载体沉默Yes-相关蛋白(Yes-associated protein,YAP)对人肾癌786-O细胞增殖和凋亡的影响。方法:用细胞免疫荧光法检测肾癌786-O细胞中YAP蛋白表达情况;构建针对YAP基因的shRNA慢病毒干扰载体转染786-O细胞。RT-PCR和Western blot分别检测干扰786-O细胞前后 YAP mRNA及蛋白的表达情况;CCK-8(cell counting kit-8)法检测沉默YAP后细胞增殖的改变;流式细胞仪(flow cytometry,FCM)检测细胞凋亡和周期的变化。结果:786-O细胞中YAP蛋白表达于细胞浆和细胞核;shRNA-YAP慢病毒干扰载体转染4 d后,可明显下调786-O细胞YAP mRNA及蛋白表达水平(P=0.000),并且明显抑制细胞增殖和促进细胞凋亡(P=0.000);细胞周期紊乱,G1 期细胞明显增加,S期细胞明显降低(P=0.000)。结论:YAP-shRNA慢病毒干扰载体能有效抑制YAP基因在786-O细胞中表达,进而抑制细胞增殖并促进细胞凋亡。
英文摘要:
      Objective:To investigate the effect of Yes-associated protein(YAP) gene silencing by shRNA on the proliferation and apoptosis of human renal cancer cells 786-O. Methods:Expression of YAP protein in 786-O cells was detected by immunofluores-cence assay. Lentiviral interference vectors of shRNA-YAP were constructed successfully and were transfected into 786-O cells. Meanwhile blank control group and negative control group were divided. Expression of YAP mRNA and protein in 786-O cells was detected by RT-PCR and Western blot,respectively. CCK-8(cell counting kit-8) assay was applied to examine the effect of YAP si-lencing on the proliferation in 786-O cells. Apoptosis and cell cycle of 786-O cells were examined by flow cytometry. Results:YAP protein was expressed in cytoplasm and nucleus of 786-O cells. On 4 d after shRNA-YAP lentiviral interference vectors being in-fected,expression of YAP mRNA and protein in 786-O cells was significantly reduced(P=0.000),cell proliferation was inhibited,and apoptosis was increased significantly(P=0.000). YAP silencing resulted in cell cycle distribution disorder in 786-O cells,which the number of G1 phase cells was significantly increased while the number of S phase cells was significantly decreased compared with that of blank control group and negative control group(P=0.000). Conclusion:shRNA-YAP lentiviral interference vectors can effectively decrease YAP gene expression in 786-O cells,inhibit cell proliferation and promote cell apoptosis.
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