胰岛素样生长因子结合蛋白7基因对胃癌细胞周期和体内外增殖的影响  

Effects of insulin-like growth factor binding protein 7 on cell cycle and proliferation of gastric cancer in vitro and in vivo

作　　者：贺德志[1] 韩艳妙 李建生[1] 

机构地区：[1]郑州大学第一附属医院消化一科,450052 [2]郑州大学第一附属医院内分泌科,450052

出　　处：《中华消化杂志》2016年第11期746-751,共6页Chinese Journal of Digestion

基　　金：河南省医学科技攻关计划（201403071）;河南省教育厅科学技术研究重点项目（14A320040）

摘　　要：目的利用过表达和干扰技术，观察胰岛素样生长因子结合蛋白7（IGFBP7）对胃癌细胞增殖和细胞周期，以及对细胞周期素D1和细胞周期素依赖性激酶4（CDK4）表达的影响，构建裸鼠移植瘤模型，观察IGFBP7对胃癌瘤体生长的影响。 方法采用小干扰核糖核酸（siRNA）干扰胃癌细胞株MKN-28中IGFBP7表达，并建立空白对照组和阴性对照组。采用pcDNA3.1-IGFBP7质粒转染SGC-7901细胞，使之过表达IGFBP7并建立相应的空白对照组和空载体组。培养48 h，采用MTT实验和克隆形成实验检测IGFBP7干扰和过表达后胃癌细胞体外增殖情况；流式细胞术检测IGFBP7对细胞周期的影响，蛋白质印迹法检测各组细胞中细胞周期素D1和CDK4。分别用过表达IGFBP7的SGC-7901稳定转染细胞及未转染SGC7901细胞建立裸鼠移植瘤模型，观察20 d内IGFBP7对胃癌移植瘤体生长的影响。采用单因素方差分析、LSD法和独立样本t检验比较组间差异。结果MTT实验和克隆形成实验发现，与空白和阴性对照组比较，干扰IGFBP7的表达能促进MKN-28细胞的增殖（3.013±0.322、2.903±0.210比4.502±0.356，F＝18.31，P＝0.002 8），细胞克隆形成增多，G1期细胞减少[（57.29±1.30）%、（52.27±0.90）%比（36.81±0.83）%，F＝321.57，P〈0.01]；与空白对照组和空载体组比较，增加IGFBP7的表达能抑制SGC-7901细胞的增殖（3.142±0.320、3.214±0.226比1.813±0.165，F＝22.35，P＝0.001 7），细胞克隆形成减少，使细胞周期处于G1期细胞增多[（49.34±1.20）%、（47.42±0.71）%比（57.73±0.73）%，F＝109.230，P 〈0.01）]。蛋白质印迹法发现干扰IGFBP7能够增加细胞周期素D1和CDK4的表达，过表达IGFBP7的作用相反（P均〈0.01）。裸鼠移植瘤模型实验发现过表达IGFBP7组的裸鼠种植瘤的体积与质量均小于对照组[（773.50±113.45） mm3比（1 038.75±101.31） mm3，（786±50） mg比（1 145±85） mg，t＝7ObjectiveTo investigate the effects of insulin-like growth factor binding protein 7 （IGFBP7） on the proliferation, cell cycle of gastric cancer cell and the expression of cynlin D1, cyclin-dependent kinase（CDK）4, and to observe the effects of IGFBP7 on the growth of gastric tumor xenografts in nude mice.MethodsThe MKN-28 cell line was interfered by small interfere ribonucleic acid （siRNA） （interfered group）, and blank control group, negative control group were also set. The overexpression of IGFBP7 in SGC-7901 induced by pcDNA3.1-IGFBP7 plasmid infection （overexpression group）, and blank control group, empty vector group were also set. Western blotting were used to observe the interference and over expression of IGFBP7 in the cell lines after 48 h. After IGFBP7 was knockdown or overexpressed, the cell proliferation of gastric cancer cells was detected by 3-（4, 5-dimethyl-2-thiazolyl）-2, 5-diphenyl-2-H-tetrazolium bromide （MTT） assay and colony formation assay. The cell cycle was determined by flow cytometry. Cynlin D1 and CDK4 were examined by Western blotting. Tumor xenografts in nude mice model was established with SGC-7901 cells from overexpressed group and untransfected SGC7901 cells. The effects of IGFBP7 on the growth of gastric cancer was observed within 20 days. Single factor analysis of variance was used, LSD test and independent sample t test were performed to compare the differences between groups.ResultsCompared with blank control group and negative control group, the proliferation of MKN-28 cells of interfered group increased （3.013±0.322, 2.903±0.210 vs 4.502±0.356, F=18.31, P=0.002 8） with more colony formation, and less cells at G1 stage （（57.29±1.30）%, （52.27±0.90）% vs （36.81±0.83）%, F=321.57）, and the differences were statistically significant （all P〈0.01）. Compared with blank control group and empty vector group, the proliferation of SGC-7901 cells of overexpressed group decreased （3.142±0.320, 3.214±0.226 vs 1.813±0.165, F=22.35,

关 键 词：胃肿瘤 人胰岛素样生长因子结合蛋白7 肿瘤细胞株 细胞增殖 细胞周期 动物实验 

分 类 号：R735.2[医药卫生—肿瘤]