解整合素样金属蛋白酶8基因调控磷脂酰肌醇3激酶／哺乳动物雷帕霉素靶蛋白通路对结肠癌细胞HCT8的影响  

Effect of a disintegrin and metalloprotease domain 8 gene on colon cancer cells HCT8 by regulating protein kinase B/the mammalian target of rapamycin pathway

作　　者：郭书伟[1] 刘洪洲[1] 李永胜[1] 吕彦东[1] 郭志强[1] 陈俊生[1] 赵晓中[1] 张利中[1] 

机构地区：[1]山西省长治医学院附属和平医院普通外科,长治046000

出　　处：《中华实验外科杂志》2017年第1期31-34,共4页Chinese Journal of Experimental Surgery

摘　　要：目的观察解整合素样金属蛋白酶8（ADAM8）基因对结肠癌HCT8细胞增殖及增殖信号转导途径磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白（PI3K/Akt/mTOR）的影响。方法体外培养结肠癌HCT8细胞，构建含ADAM8基因过表达质粒载体和ADAM8基因RNA干扰质粒载体；转染至HCT8细胞，分为过表达组、干扰组和对照组。采用5-乙炔基-2’脱氧尿嘧啶核苷（EdU）和3-（4，5-二甲基吡啶-2-基）-5-（3-羧基甲氧基苯基）-2-（4-磺苯基）-2H-四唑（MTS）方法检测细胞增殖，PI3K酶活检测试剂盒检测细胞内PI3K酶活，Western blot方法检测细胞内Akt、磷酸化蛋白激酶B（p-Akt）相对表达量和mTOR的表达量。结果（1）过表达组ADAM8表达量显著高于对照组（t＝1.143，P＝0.035），而干扰组ADAM8的表达量显著低于对照组（t＝1.203，P＝0.027），说明过表达载体和干扰表达载体构建成功。（2）过表达组细胞增殖能力[（1.31±0.15） copies/ml]显著高于对照组[（1.00±0.10） copies/ml，t＝1.157，P＝0.033]，干扰组细胞增殖能力[（0.71±0.09） copies/ml]显著低于对照组[（1.00±0.10） copies/ml，t＝1.172，P＝0.031]。（3）与对照组比较，过表达组和干扰组细胞内PI3K的酶活变化差异无统计学意义。ADAM8过表达后细胞内p-Akt表达量增加，被干扰表达后p-Akt表达量降低，差异有统计学意义（t＝1.319，P＝0.018）。ADAM8过表达后细胞内mTOR表达量增加，被干扰表达后mTOR表达量降低，差异有统计学意义（t＝1.390，P＝0.014）。结论ADAM8可通过增强Akt的磷酸化和mTOR的表达促进HCT8细胞增殖。Objective To investigate the effects of a disintegrin and metalloprotease domain 8 （ADAM8） gene on the proliferation of colon cancer cells HCT8, and the phosphatidylinositol-3-kinase/protein kinase B/the mammalian target of Rapamycin （PI3K/Akt/mTOR） signal transduction pathway.Methods The HCT8 cells were cultured in vitro, and transfected with ADAM8 overexpression plasmid vector （overexpression group） and ADAM8 gene RNA interference plasmid vector （interference group） respectively, A control group was set up. Cell proliferation was detected by 5-Ethynyl -2’-deoxyuridine （EdU） and 3-（4, 5-Dimethylthiazol-2-yl）-5-（3-carboxymethoxyphenyl）-2-（4-sulfophenyl）-2H-tetrazolium （MTS） methods. The PI3K enzyme activity was detected by PI3K activity assay kit. The relative expression of Akt, phosphorylated Akt （p-Akt） and the expression of mTOR were detected by Western blotting.Results （1） The expression of ADAM8 in overexpression group was significantly higher than that in control group （t=1.143, P=0.035）, while the expression of ADAM8 in interference group was significantly lower than that in control group （t=1.203, P=0.027）, which indicated that overexpression vector and interference expression vector were successfully constructed. （2） The cell proliferation capacity of overexpression group [（1.31±0.15） copies/ml] was significantly higher than that of control group [（1.00±0.10） copies/ml, t=1.157, P=0.033], and the cell proliferation ability [（0.71±0.09） copies/ml] in overexpression group was significantly lower than that of control group [（1.00±0.10） copies/ml, t=1.172, P=0.031]. （3） As compared with the control group, there was no significant difference in the changes of enzyme activities of PI3K in the overexpression and interference groups. The expression of p-Akt was increased, and the expression of p-Akt was decreased after the overexpression of ADAM8. The difference was statistically significant （t=1.319, P=0.018）. The expression o

关 键 词：解整合素样金属蛋白酶8 结肠癌 干扰表达 

分 类 号：R735.35[医药卫生—肿瘤]