解剖学报

解剖学报 ›› 2017, Vol. 48 ›› Issue (5): 545-549.doi: 10.16098/j.issn.0529-1356.2017.05.008

• 肿瘤生物学 • 上一篇    下一篇

支架蛋白活化蛋白激酶C受体1在胃癌细胞中的表达及其与胃癌细胞增殖的关系

刘超1 任丽莉2 王一曌1 刘乙蒙1 肖建英3*   

  1. 1. 锦州医科大学基础医学院发育生物学教研室，辽宁 锦州 121001; 2.锦州医科大学基础医学院神经生物学教研室，辽宁 锦州 121001; 3.锦州医科大学教务处，辽宁 锦州 121001
  • 收稿日期:2016-11-07 修回日期:2017-05-27 出版日期:2017-10-06 发布日期:2017-10-06
  • 通讯作者: 肖建英 E-mail:xiaojianying@lnmu.edu.cn
  • 基金资助:
    国家自然科学基金资助课题

Expression of receptor for activated C kinase 1 in gastric cancer cells and its relationship with the proliferation of gastric cancer cells

LIU Chao1 REN Li-li2 WANG Yi-zhao1 LIU Yi-meng1 XIAO Jian-ying3*   

  1. 1. Department of Developmental Biology; 2. Neurobiology; 3. Teaching Affairs Department of Jinzhou Medical University, Liaoning Jinzhou 121001, China
  • Received:2016-11-07 Revised:2017-05-27 Online:2017-10-06 Published:2017-10-06
  • Contact: XIAO Jian-ying E-mail:xiaojianying@lnmu.edu.cn

PDF

摘要：

目的 探讨活化蛋白激酶C受体1C(RACK1, GNB2L1)在胃癌细胞HGC27中的表达及过表达RACK1对HGC27生长增殖的影响。方法 体外培养胃癌未分化细胞HGC27和正常胃黏膜上皮细胞系GES-1，收集细胞48 h后，提取mRNA和蛋白，利用RT-PCR检测RACK1 mRNA在HGC27和GES-1细胞中的表达；利用Western blotting法检测RACK1蛋白在两种细胞中的表达；以人胚肾HEK293细胞cDNA为模板，构建pcDNA3.1A-flag-RACK1重组质粒，利用Lipo2000转染入HGC27细胞中，Western blotting法检测质粒的转染效率，MTT法检测过表达RACK1对HGC27胃癌细胞系生长增殖的影响。 结果 HGC27胃癌细胞中RACK1的mRNA和蛋白水平表达低于GES-1细胞（P<0.01）。双酶切鉴定和测序分析表明，pcDNA3.1A-flag-RACK1重组质粒构建成功。将该质粒转入HGC27细胞后，与未转染组相比，空载转染组RACK1蛋白表达无明显区别（P>0.05），pcDNA3.1-RACK1转染组RACK1蛋白表达明显升高（P<0.01）；转染pcDNA3.1A-flag-RACK1转染组细胞存活率在72 h和96 h明显少于pcDNA3.1空载对照组（P<0.01）。结论 支架蛋白RACK1的mRNA和蛋白水平在HGC27细胞中低表达，上调RACK1的表达可明显抑制HGC27细胞增殖。

关键词: 蛋白激酶1活化受体, 胃癌细胞, 增殖, HGC27细胞, 免疫印迹法

Abstract:

Objective To study the expression of the receptor for activated protein C kinase 1(RACK1, GNB2L1) in gastric cancer cells HGC27 and the effect of overexpressed RACK1 on the growth of HGC27 cells. Methods The gastric carcinoma cells HGC27 and normal gastric mucosa GES-1 cells were cultured in vitro and collected 48hours later. RNA and protein were extracted from the cells. The expression of RACK1 mRNA and protein was detected by RT-PCR and Western blotting in HGC27 and GES-1 cells, respectively. The recombinant plasmid pcDNA3.1A-flag-RACK1 was constructed using human embryo kidney cell HEK293 cDNA as template and transfected into HGC27 cells with Lipo2000, and the transfection efficiency was evaluated by Western blotting and the survival rate of HGC27 cells was detected by MTT method . Results The expressions of RACK1 mRNA and protein in HGC27 cells were lower than that in GES-1 cells (P<0.01). Double enzyme digestion analysis and sequencing analysis showed that the recombinant plasmid of pcDNA3.1A-flag-RACK1 was successfully constructed. There was no significant difference between untransfected group and pcDNA3.1 vector group（P>0.05）, while the RACK1 protein expression was significantly increased compared with untransfected group and vector transfected group(P<0.01). The survival rate of cells transfected with pcDNA3.1A-flag-RACK1 at 72 hours and 96 hours was significantly less than that of cells with pcDNA3.1 transfection (P<0.01). Conclusion The expression level of mRNA and protein of scaffold protein RACK1 in HGC27 gastric carcinoma cells is downregulated. Overexpressed RACK1 can significantly inhibit the growth of HGC27 cells. This study provides a new theoretical and experimental basis for RACK1 as a new molecular marker and star molecule in signaling pathways.

Key words: Receptor for activated C kinase 1, Gastric cancer cell, Proliferation, HGC27 cell, Western blotting