lncRNA SNHG16在结直肠癌组织和细胞中表达及其调控结肠癌细胞中GPAM表达的机制  

Expression of lncRNA SNHG16 in colorectal cancer tissues and cells and its regulationon expression of GPAM in colon cancer cells

作　　者：周云松[1] 温小辉 张琦[3] 寇炜[1] ZHOU Yunsong;WEN Xiaohui;ZHANG Qi;KOU Wei(Clinical Skill Training Center,Medical School of Northwest University for Nationalities,Lanzhou 730030,Gansu,China;Clinical College,Gansu University of Traditional Chinese Medicine,Lanzhou 730000,Gansu,China;Department of Endocrinology,Gansu Provincial People's Hospital,Lanzhou 730000,Gansu,China)

机构地区：[1]西北民族大学医学院临床技能中心,甘肃兰州730030 [2]甘肃中医药大学临床学院,甘肃兰州730000 [3]甘肃省人民医院内分泌科,甘肃兰州730000

出　　处：《中国肿瘤生物治疗杂志》2019年第1期58-66,共9页Chinese Journal of Cancer Biotherapy

基　　金：国家自然科学基金资助项目(No.81460677)~~

摘　　要：目的:探讨长链非编码RNA(long non-coding RNA,lncRNA)SNHG16在结直肠癌(colorectal cancer,CRC)组织和细胞中的表达及其通过海绵吸附miR-128-3p调控结肠癌细胞线粒体甘油3磷酸酰基转移酶基因(mitochondrial glycerol-3-phosphate acyltransferase,GPAM)表达的分子机制。方法:收集2014年1月至2017年1月甘肃省人民医院肛肠科手术切除的60例CRC患者的癌及癌旁组织标本,以及结直肠癌细胞系SW480、SW620、HCT116、Caco-2、DLD-1、HT29和结肠上皮细胞CCD841,用q PCR法检测CRC组织和细胞系中SNHG16的表达,分析SNHG16表达与CRC患者临床病例特征的关系。分别用miR-128-3p模拟物、miR-128-3p抑制剂、SNHG16敲降载体转染SW480细胞后,用qPCR法检测细胞中miR-128-3p及SNHG16 mRNA的表达,用Western blotting法检测GPAM蛋白的表达,用CCK-8法、克隆形成实验及细胞凋亡实验、Transwell小室法检测细胞的增殖、凋亡及侵袭。用双荧光素酶报告基因法和RNA免疫共沉淀实验验证SNHG16和miR-128-3p mRNA靶向结合。构建小鼠SW480细胞移植瘤模型,观察敲降SNHG16对移植瘤生长的影响。结果:CRC组织及细胞系中SNHG16高表达(均P<0.01),其表达水平与CRC淋巴结转移、Duke’s分期及患者生存期相关(均P<0.01)。敲降SNHG16可显著抑制SW480细胞的增殖及侵袭能力,并诱导细胞凋亡(均P<0.01);敲降SNHG16后小鼠移植瘤瘤体显著小于对照组(P<0.05)。双荧光素酶报告基因检测及RNA免疫沉淀反应结果显示,miR-128-3p与SNHG16相互作用,且在CRC患者中miR-128-3p与SNHG16负相关(P<0.01)。SNHG16通过内源性竞争海绵吸附miR-128-3p影响其下游靶基因GPAM的表达。结论:SNHG16在CRC细胞中可通过海绵吸附miR-128-3p调控GPAM表达,SNHG16及miR-128-3p可作为CRC诊断及治疗的潜在靶点。Objective: To investigate the expression of long non-coding RNA SNHG16(lncRNA SNHG16) in colorectal cancer(CRC)tissues and cells, and to explore the mechanism of its regulation on the expression of mitochondrial glycerol-3-phosphate acyltransferase(GPAM) via sponging miR-128-3 p. Methods: Sixty pairs of colorectal cancerous tissues and para-cancerous tissues that resected from CRC patients, who underwent surgery in the Department of Anorectal Surgery, Gansu Provincial People’s Hospital during Jan.2014 and Jan. 2017, were collected for this study;In addition, CRC cell lines(SW480, SW620, HCT116, Caco-2,DLD-1, HT29) and colonic epithelial cell line CCD841 were also collected for the study. The expression of SNHG16 in collected tissues and cell lines was determined by Real-time quantitative PCR(qPCR), and its correlation to the clinicopathological features of CRC patients was also analyzed. SW480 cells were transfected with miR-128-3 p mimic, miR-128-3 p inhibitor, and si-SNHG16, respectively, and then the mRNA expressions of miR-128-3 p and SNHG16 were detected by qPCR, the protein expression of GPAM was determined by Western blotting,and the cell proliferation, apoptosis and invasion were detected by CCK-8 assay, colony formation assay, cell apoptosis assay and Transwell chamber assay, respectively. The binding between SNHG16 and miR-128-3 p was validated with dual luciferase reporter gene assay and RNA Immunoprecipitation assay. For in vivo experiment, mouse model of SW480 cell exnograft was constructed, and the ef-fect of SNHG16 knockdown on the growth of exnograft was observed. Results: SNHG16 was found to highly expressed in human CRC tissues and cell lines(all P<0.01), and SNHG16 expression level was associated with lymph node metastasis, Duke’s stage and patients’ survival(all P<0.01). Knockdown of SNHG16 significantly inhibited CRC cell proliferation and invasion, and induced apoptosis(all P<0.01);After SNHG16 knockdown, the volume of exnograft was obviously reduced(P<0.05). Dual luciferase reporter g

关 键 词：结直肠癌 SW480细胞 SNHG16 miR-128-3p 线粒体甘油3磷酸酰基转移酶基因 海绵 

分 类 号：R735.3[医药卫生—肿瘤] R730.5[医药卫生—临床医学]