LncRNA KCNQ1OT1降表达对顺铂耐药卵巢癌细胞耐药性的影响及其作用机制  

Effect of knockdown of LncRNA KCNQ1OT1 expression on resistance of cisplatin-resistant ovarian cancer cells to cisplatin

作　　者：沈艳丽[1] 冯文广[1] 杨静[1] 伊婷 易金玲[1] SHEN Yanli;FENG Wenguang;YANG Jing;YI Ting;YI Jinling(Department of Gynecology,The Fifth Affiliated Hospital of Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学第五附属医院妇科,乌鲁木齐830011

出　　处：《山东医药》2022年第17期12-16,共5页Shandong Medical Journal

基　　金：新疆维吾尔自治区自然科学基金青年基金项目(2018D01C309)。

摘　　要：目的观察长链非编码RNA(LncRNA)KCNQ1OT1降表达对顺铂(DDP)耐药卵巢癌细胞耐药性的影响并分析其作用机制。方法选用SKOV-3人卵巢癌细胞,经DDP反复间歇冲击诱导法构建DDP耐药卵巢癌细胞株SKOV3/DDP;以不同浓度DDP作用SKOV3、SKOV3/DDP细胞,CCK-8法检测细胞增殖情况得到50%细胞生长的药物浓度(IC50)并计算SKOV3/DDP细胞耐药指数(RI);采用qRT-PCR法检测SKOV-3、SKOV3/DDP细胞中的KCNQ1OT1。将SKOV3/DDP细胞分为三组,KCNQ1OT1降表达组和转染对照组中分别转染KCNQ1OT1 siRNA、NC-siRNA 48 h,对照组不转染,采用qRT-PCR法检测KCNQ1OT1观察转染效果;以不同浓度DDP作用各组细胞,以CCK-8法检测细胞增殖情况得到IC50并计算RI观察耐药性变化;取转染后的三组细胞,采用流式细胞术检测细胞周期,Western blotting法检测细胞p-AKT、p-mTOR蛋白。结果DDP对SKOV-3、SKOV-3/DDP细胞IC50分别为1.224、4.920μg/mL,SKOV-3/DDP细胞RI为4.020。与SKOV-3细胞比较,SKOV-3/DDP细胞KCNQ1OT1表达量增高(P<0.05)。KCNQ1OT1降表达组细胞中KCNQ1OT1表达量均低于转染对照组及对照组(P均<0.05),转染对照组与对照组间差异无统计学意义。随DDP浓度增加,各组细胞增殖抑制率均升高(P均<0.05);在相同DDP浓度下,KCNQ1OT1降表达组细胞增殖抑制率均高于转染对照组和对照组(P均<0.05)。DDP对KCNQ1OT1降表达组细胞IC50为3.613μg/mL,RI为0.734;DDP对转染对照组、对照组细胞IC50分别为9.737、8.461μg/mL,RI分别为1.979、1.720。与转染对照组和对照组比较,KCNQ1OT1降表达组G0/G1期细胞比例、p-mTOR蛋白表达量升高,S期和G2/M期细胞比例、p-AKT蛋白表达量降低(P均<0.05);转染对照组与对照组各期细胞比例及p-AKT、p-mTOR蛋白表达量差异均无统计学意义。结论干扰KCNQ1OT1表达可有效逆转卵巢癌细胞对DDP的耐药,其机制可能与阻滞耐药细胞于G0/G1期并调控AKT/mTOR信号通路活性有关。Objective To investigate the effect and mechanism of inhibiting LncRNA KCNQ1OT1 expression on the cisplatin resistance of cisplatin-resistant ovarian cancer cells.Methods The cisplatin-resistant ovarian cancer cell line(SKOV3/DDP)from the original SKOV-3 human ovarian cancer cells was established using DDP repeated intermittent shock induction.The relative proliferation was determined using the cell counting Kit-8(CCK-8)assay to obtain the IC50 value.The reactivity index(RI)of the cell was calculated.The qRT-PCR method was used to detect the expression level of LncRNA KCNQ1OT1 in SKOV-3 and SKOV3/DDP cells.The SKOV3/DDP cells were randomly divided into the three groups:the knockdown KCNQ1OT1 group,negative control(NC)group,and control group.The cells in the knockdown KCNQ1OT1 group and NC group were transfected with KCNQ1OT1 siRNA and NC-siRNA for 48 h,respectively.The cells in the control group were incubated under the same conditions but were not transfected.KCNQ1OT1 was detected by qRT-PCR to observe the transfection effect;cells in each group were treated with different concentrations of DDP,the cell proliferation was detected by CCK-8 method to obtain IC50 and RI was calculated to observe the change of drug resistance;we took cells from the three groups after transfection,the cell cycle was detected by flow cytometry,and the protein expression of p-AKT and p-mTOR was detected by Western blotting.Results IC50 Values of SKOV-3 and SKOV3/DDP cells treated with DDP were 1.224μg/mL and 4.920μg/mL,respectively.The RI was 4.020.Compared with SKOV-3 cells,the relative expression of KCNQ1OT1 in SKOV-3/DDP cells significantly increased(P<0.05).The relative expression of KCNQ1OT1 in the knockdown KCNQ1OT1 group was lower than those of the NC group and control group,while there was no significant difference between the NC group and control group.With the increase of DDP concentrations,the inhibition rate of cell proliferation in each group increased(P<0.05);at the same DDP concentration,the inhibition rate of cell pro

关 键 词：卵巢癌 KCNQ1OT1 AKT/mTOR信号通路 肿瘤耐药 顺铂 细胞周期 

分 类 号：R73-3[医药卫生—肿瘤]