抑制S1PR2蛋白表达对卵巢上皮性癌SKOV3细胞增殖能力的影响  

Effect of SIPR2 inhibition on epithelial ovarian cancer SKOV3 cell proliferation in vitro and in vivo

作　　者：戴岚[1] 刘艺璇[1] 谢蕾[1] 狄文[1] 

机构地区：[1]上海交通大学医学院附属仁济医院妇产科暨上海市妇科肿瘤重点实验室,200127

出　　处：《中华妇产科杂志》2018年第2期106-110,共5页Chinese Journal of Obstetrics and Gynecology

基　　金：国家自然科学基金（81402128）

摘　　要：目的探讨抑制1-磷酸鞘氨醇受体2（S1PR2）对卵巢上皮性癌（卵巢癌）细胞增殖能力的影响及其作用机制。方法（1）设计针对S1PR2基因的小分子干扰RNA（siRNA，命名为si-S1PR2），以及与S1PR2基因无关的siRNA（作为阴性对照）；以卵巢癌细胞系SKOV3细胞作为研究对象，实验分为3组，分别为si-S1PR2转染组、阴性对照组和空白对照组，采用蛋白印迹（western blot）法检测3组SKOV3细胞中S1PR2蛋白的表达。（2）体外实验：分组同前，采用活细胞计数（CCK-8）法检测转染后不同时间点（分别为24、48、72 h）3组SKOV3细胞增殖的抑制率；流式细胞仪检测转染后3组SKOV3细胞的细胞周期比例；western blot法检测转染后3组SKOV3细胞中磷酸化细胞外调节蛋白激酶1/2（p-ERK1/2）蛋白的表达水平。（3）体内实验：将SKOV3细胞接种于裸鼠腹腔，构建卵巢癌裸鼠腹腔移植瘤模型。实验分为两组，分别为S1PR2抑制剂组和对照组（每组8只裸鼠），两组裸鼠在腹腔接种SKOV3细胞后7 d开始腹腔给药，分别给予S1PR2抑制剂——JTE-013和磷酸盐缓冲液（PBS），每周2次共8次。28 d后处死并解剖两组裸鼠，观察腹腔移植瘤的生长情况。结果（1）western blot法检测显示，si-S1PR2转染组SKOV3细胞中S1PR2蛋白的表达水平为0.24±0.04，与阴性对照组（1.07±0.13）、空白对照组（1.10±0.14）分别比较，差异均有统计学意义（P均〈0.01）。（2）CCK-8法检测显示，转染24、48、72 h后，si-S1PR2转染组SKOV3细胞增殖的抑制率[分别为（26.6±3.3）%、（35.0±3.4）%、（34.0±2.8）%]明显高于阴性对照组[分别为（1.7±0.9）%、（2.5±0.5）%、（2.4±1.1）%；P均〈0.01]及空白对照组（均为0；P均〈0.01）。流式细胞仪检测显示，转染48 h后，si-S1PR2转染组细胞的G0/G1期比例为（70.9±2.8）%，明显高于空白对照组及阴性对照组[分别为（61.7±2.4）%、（62.1Objective To study the effect and mechanism of S1PR2 inhibition on epithelial ovarian cancer SKOV3 cell proliferation in vitro and in vivo.Methods （1） A pair of S1PR2 gene small interference RNA （siRNA） , namely si-S1PR2, and a pair of negative control siRNA were designed. Western blot methods were used to detect the silence efficiency of the S1PR2 in the si-S1PR2 group, blank control group and negative control group. （2） Study in vitro： the experiment included three groups, namely si-S1PR2 group, blank control group and negative control group. Cell counting kit-8 （CCK-8） assay was used to detect the proliferation inhibition rates of the transfected cells. The cell cycles of the transfected cells were measured by flow cytometry. Western blot was used to detect the levels of phosph-extracellular regulated protein kinase 1/2 （p-ERK1/2） of the transfected cells. （3） Study in vivo：to establish intraperitoneal transplantation models, 8 mice in each group were intraperitoneally injected with 5×106 SKOV3 cells. Phosphate buffered saline （PBS） or JTE-013 were administered into mice twice per week starting on day 7 after the injection of the cancer cells. Twenty-eight days after nude mice intraperitoneal injection with JTE-013 or PBS, the mice were sacrificed and the number and the weight of visible tumors were calculated.Results （1） The results of western blot showed that the relative S1PR2 protein expression levels were 0.24±0.04 in the si-S1PR2 group, which was lower than that in the blank control group （1.10±0.14, P〈0.01） and negative control group （1.07±0.13, P〈0.01） . （2） The results of CCK-8 assay indicated that after transfected for 24, 48 and 72 hours, the proliferation inhibition rate of si-S1PR2 group were respectively （26.6±3.3） %, （35.0±3.4） %, and （34.0±2.8） %, significantly lower than those in the blank control group （all 0; all P〈0.01） and negative control group [ （1.7±0.9） %, （2.5±0.5） %,and （2.4±1.1） % respectivel

关 键 词：卵巢肿瘤 受体 鞘磷脂 细胞系 肿瘤 细胞增殖 

分 类 号：R737.31[医药卫生—肿瘤]