lncRNA TNK2‐AS1调控miR‐802表达对结直肠癌细胞增殖与迁移的影响  

Effects of lncRNA TNK2⁃AS1 on proliferation and migration of colorectal cancer cells by regulating miR⁃802 expression

作　　者：温舒淼 李红菊[2] 陈晓杨[1] 隋楠[1] WEN Shu-miao;LI Hong-ju;CHEN Xiao-yang;SUI Nan(Endoscopy Room of the Third Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Shenyang,Liaoning 110040,China;Department of Gastroenterology,Liaoning Provincial People's Hospital,Shenyang,Liaoning Province 110040,China)

机构地区：[1]辽宁中医药大学附属第三医院腔镜室,辽宁沈阳110040 [2]辽宁省人民医院消化内科,辽宁沈阳110040

出　　处：《解剖学研究》2024年第6期523-532,539,共11页Anatomy Research

基　　金：国家自然科学基金项目(82174370)。

摘　　要：目的探讨长链非编码RNA TNK2反义RNA 1(lncRNA TNK2‐AS1,简称TNK2‐AS1)调控微小RNA‐802(miR‐802)表达对结直肠癌HT29细胞增殖、迁移、侵袭的作用与机制。方法qRT‐PCR检测TNK2‐AS1和miR‐802表达;双荧光素酶报告基因实验、RNA pull down和RNA结合蛋白免疫沉淀(RIP)实验验证TNK2‐AS1和miR‐802的靶向关系;将细胞分为NC组、pcDNA‐NC组、pcDNA‐TNK2‐AS1组、si‐NC组、si‐TNK2‐AS1组、si‐TNK2‐AS1+inhibitor‐NC组、si‐TNK2‐AS1+miR‐802 inhibitor组;Western blot检测蛋白表达;MTT法和EdU法分别检测细胞增殖;Transwell检测细胞迁移与侵袭;检测裸鼠移植瘤的质量和体积及TNK2‐AS1和miR‐802表达。结果与癌旁组织比较,结直肠癌组织中TNK2‐AS1表达升高(分别为1.00±0.05、2.83±0.21,P<0.05),miR‐802表达下降(分别为1.00±0.06、0.44±0.03,P<0.05);与正常结肠上皮细胞NCM460中TNK2‐AS1、miR‐802表达(1.00±0.05)比较,结直肠癌细胞系SW480、SW620、HCT116和HT‐29中TNK2‐AS1表达升高(分别为1.64±0.19、1.37±0.36、1.48±0.30、1.92±0.25,均P<0.05),miR‐802表达下降(分别为0.65±0.08、0.53±0.04、0.69±0.05、0.41±0.02,均P<0.05);TNK2‐AS1和miR‐802具有靶向关系;过表达TNK2‐AS1促进HT29细胞增殖、迁移侵袭,下调miR‐802表达(P<0.05);与si‐NC组比较,si‐TNK2‐AS1组HT29细胞TNK2‐AS1表达、增殖能力、CyclinD1表达、细胞迁移、侵袭数目及MMP‐2、MMP‐9表达明显下降,miR‐802表达明显上升(P<0.05);抑制miR‐802表达逆转了沉默TNK2‐AS1表达对HT29细胞增殖、迁移及对裸鼠移植瘤的抑制作用(P<0.05)。结论沉默TNK2‐AS1表达可能靶向上调miR‐802表达,抑制结直肠癌细胞的增殖、迁移及移植瘤生长。Objective To investigate the effect and mechanism of long chain non‐coding RNA TNK2 antisense RNA 1(lncRNA TNK2‐AS1,TNK2‐AS1 for short)regulating the expression of microRNA‐802(miR‐802)on the proliferation,migration and invasion of colorectal cancer HT29 cells.Methods The expression of TNK2‐AS1 and miR‐802 in rectal cancer tissues and cells was detected by qRT‐PCR;the dual luciferase reporter gene experiment,RNA pull down and RNA‐binding protein immunoprecipitation(RIP)assay were applied to verify the targeting relationship between TNK2‐AS1 and miR‐802;cells were grouped into NC group,pcDNA‐NC group,pcDNATNK2‐AS1 group,si‐NC group,si‐TNK2‐AS1 group,si‐TNK2‐AS1+inhibitor NC group,and si‐TNK2‐AS1+miR‐802 inhibitor group;Western blot was applied to detect protein expression;cell proliferation was detected by MTT and EdU methods,respectively;Transwell was applied to detect cell migration and invasion;detect the mass and volume of transplanted tumors in nude mice,as well as the expression of TNK2‐AS1 and miR‐802.Results Compared with adjacent tissues,TNK2‐AS1 expression was increased(1.00±0.05,2.83±0.21 respectively,P<0.05)in colorectal cancer tissues,and miR‐802 expression was decreased(1.00±0.06,0.44±0.03 respectively,P<0.05);Compared with normal colon epithelial cells NCM460,TNK2‐AS1 expression(1.00±0.05)in colorectal cancer cell lines SW480,SW620,HCT116 and HT‐29 was increased(1.64±0.19,1.37±0.36,1.48±0.30,1.92±0.25 respectively,all P<0.05),miR‐802 expression decreased(0.65±0.08,0.53±0.04,0.69±0.05,0.41±0.02 respectively,all P<0.05);The double luciferase reporter gene experiment showed that TNK2‐AS1 and miR‐802 had a targeting relationship;overexpression of TNK2‐AS1 promoted HT29 cell proliferation,migration,and invasion,while downregulating miR‐802 expression(P<0.05);Compared with si‐NC group,the expression of TNK2‐AS1,proliferative ability,expression of CyclinD1,cell migration,invasion number,and expression of MMP‐2 and MMP‐9 in

关 键 词：长链非编码RNA TNK2反义RNA 1 微小RNA‐802 结直肠癌 增殖 迁移 

分 类 号：R735.34[医药卫生—肿瘤]