干扰CASC9表达对结直肠癌细胞增殖、凋亡的影响及其与miR-195-5p/CCND1轴的关系  

Effects of interfering with CASC9 expression on proliferation and apoptosis of colorectal cancer cells and their relationship with miR-195-5p/CCND1 axis

作　　者：梁峰[1] 张玮[2] 王胜杰 黄晓梦 岳磊 武雪亮[1] LIANG Feng;ZHANG Wei;WANG Shengjie;HUANG Xiaomeng;YUE Lei;WU Xueliang(Department of General Surgery,The First Affiliated Hospital of Hebei North University,Zhangjiakou 075000,China;不详)

机构地区：[1]河北北方学院附属第一医院普通外科,河北张家口075000 [2]河北北方学院附属第一医院小儿外科 [3]河北北方学院附属第一医院医务处

出　　处：《山东医药》2021年第26期1-5,共5页Shandong Medical Journal

基　　金：河北省卫计委医学科学重点课题计划(20210282)。

摘　　要：目的探讨长链非编码RNA癌症易感性候选基因9(CASC9)对结直肠癌细胞增殖、凋亡的影响,并探讨其作用机制与miR-195-5p细胞周期蛋白D1(CCND1)轴的关系。方法取人正常结肠黏膜上皮细胞系NCM460和结直肠癌细胞系SW480、SW620、HCT-116,采用实时荧光定量PCR法检测四种细胞中的CASC9的表达量。将CASC9表达量最高的结直肠癌细胞分为CASC9干扰组和干扰对照组,CASC9干扰组转染shRNA-CASC9,干扰对照组转染sh-NC。采用CCK-8法检测两组细胞增殖情况,流式细胞术检测两组细胞凋亡率,实时荧光定量PCR法检测细胞中miR-195-5p、CCND1 mRNA表达。使用StarBase数据库预测CASC9与miR-195-5p、miR-195-5p与CCND1的潜在结合位点,采用双荧光素酶报告基因检测法进行验证。结果与NCM460细胞相比,SW480、SW620、HCT-116细胞CASC9表达升高,其中SW480细胞CASC9表达高于SW620、HCT-116细胞(P均<0.01)。选择SW480细胞用于后续实验,CASC9干扰组细胞增殖能力较干扰对照组降低(P<0.01),细胞凋亡率较干扰对照组升高(P<0.05),miR-195-5p表达升高、CCND1 mRNA表达降低(P均<0.01)。StarBase数据库分析表明,CASC9可靶向吸附miR-195-5p;CCND1为miR-195-5p下游靶蛋白。荧光素酶报告基因结果显示,CASC9可与miR-195-5p特异性结合(P<0.01)。结论CASC9在结直肠癌细胞中表达升高,其机制可能是CASC9作为miR-195-5p吸附海绵靶向调控CCND1表达,从而促进结直肠癌细胞的增殖并抑制其凋亡。Objective To investigate the effects of long non-coding RNA cancer susceptibility candidate 9(CASC9)on the proliferation and apoptosis of colorectal cancer cells,and to explore the relationship between its mechanism and miR-195-5p cyclin D1(CCND1)axis.Methods The expression levels of CASC9 in the normal colon epithelial cell line NCM460 and colorectal cancer cell lines SW480,SW620 and HCT-116 were detected by real-time fluorescence quan⁃titative PCR.Colorectal cancer cells with the highest expression of CASC9 were divided into the CASC9 interference group and control group.The cells in the CASC9 interference group were transfected with SHRNA-CASC9,and the control group with SH-NC.Cell proliferation of the two groups was detected by CCK-8,apoptosis rates of the two groups were detected by flow cytometry,and mRNA expression levels of miR-195-5p and CCND1 were detected by real-time quantitative PCR.The potential binding sites of CASC9 and miR-195-5p,and miR-195-5p and CCND1 were predicted by StarBase database,and were verified by double luciflucase reporter assay.Results Compared with NCM460 cells,the expression levels of CASC9 in SW480,SW620 and HCT-116 cells increased,and the expression level of CASC9 in SW480 cells was higher than those in SW620 and HCT-116 cells(all P<0.01).SW480 cells were selected for subsequent experiments.Compared with the control group,the proliferation ability of CASC9 cells decreased(P<0.01),the apoptosis rate increased(P<0.05),and the expression levels of miR-195-5p and CCND1 mRNA increased in the interference group(both P<0.01).StarBase database analysis showed that CASC9 could target and adsorb miR-195-5p.CCND1 was the downstream target protein of miR-195-5p.Luciferase reporter gene results showed that CASC9 could specifically bind with miR-195-5p(P<0.01).Conclusion CASC9 is highly expressed in colorectal cancer tissues and cells,and its mechanism may be that CASC9,as a miR-195-5p adsorption sponge,regulates the expression of CCND1 and promotes the proliferation of colorec⁃tal canc

关 键 词：结直肠癌 长链非编码RNA 癌症易感性候选基因9 miR-195-5p 内源竞争RNA 

分 类 号：R735.3[医药卫生—肿瘤]