目的 检测Clorf63基因在肝癌和癌旁组织中的定位,以及Clorf63在多种人肝癌细胞系和人胃癌细胞系中的mRNA和蛋白表达水平；检测干涉Clorf63后对人胃癌细胞MKN28的细胞周期的影响.方法 免疫组化染色法检测Clorf63基因在组织中的定位；Real-time PCR和Western blot分别检测Clorf63基因在细胞中的表达；脂质体转染法将干涉片段转染至MKN28细胞中,Real-time PCR和Western blot法筛选有效干涉片段；流式细胞术检测Clorf63干涉后对MKN28细胞的细胞周期的影响.结果 Clorf63在肝癌和癌旁组织细胞中都有胞浆表达,在MKN28细胞中表达量较高,在MKN45细胞中表达量较低；成功筛选出Clorf63基因的有效干涉片段；Clorf63干涉后,胃癌细胞MKN28发生G1期阻滞,经统计学分析,Clorf63干涉后MKN28细胞增殖指数降低(P＜0.05).结论 Clorf63在人肝癌和癌旁组织细胞胞浆中都有表达,在MKN28细胞中表达量最高；干涉Clorf63基因后,可使MKN28细胞发生G1期阻滞,为该基因功能的后续研究奠定了基础.%Objective To investigate the localization of Clorf63 gene in liver cancer and para-carcinomatous tissues,and the expression of Clorf63 gene in several liver cancer cell lines and gastric cancer cell lines. To study the Clorf63 silencing effects on cell cycle of gastric cancer cell MKN28 by RNA interference. Methods Immunohistochemical staining was used to detect the localization of Clorf63 in liver cancer tissues. Interference clips were carried into MKN28 cell lines by liposomes. The mRNA and protein expression levels were detected by real-time PCR and Western blot, respectively. Flow cytometry was applied to analyze cell cycle after silencing Clorf63. Results Clorf63 was cytoplasmically expressed in liver cancer tissues and precancerous tissues. The expression of Clorf63 in MKN28 cells was the highest. On the contrary, the expression of Clorf63 gene in MKN45 cells was lower. After effective interference of Clorf63, the G1 phase was blocked and the cell proliferation index was significantly reduced in MKN28 cells (P<0.05). Conclusion Clorf63 gene was cytoplasmically expressed. The expression of Clorf63 was the highest in MKN28 cells. The effective interference of Clorf63 caused the G1 phase blocking and decreased cell proliferation index in MKN28 cells. Our study has laid foundation for the follow-up study on Clorf63 gene.
展开▼
