长链非编码RNA LINC00339在结直肠癌中的表达和作用机制

摘要

目的 研究长链非编码RNA LINC00339在结直肠癌患者中的表达情况及其对结直肠癌细胞增殖和凋亡的作用与机制.方法 回顾性分析2015年8月至2017年1月河南省肿瘤医院收治的经病理确证的158例结直肠癌患者.采用实时荧光定量PCR方法检测结直肠癌组织和癌旁组织中LINC00339的表达,分析LINC00339的表达与患者临床病理特征和微RNA(miR)-218表达的关系.双荧光素酶报告系统验证LINC00339与miR-218的关系.在高表达LINC00339的结直肠癌细胞LoVo和HCT116中使用siRNA干扰技术下调LINC00339,实时荧光定量PCR检测miR-218的表达、MIT检测细胞活力、流式细胞术检测细胞凋亡.另外,在LoVo和HCT116细胞中共转染LINC00339 siRNA和miR-218拮抗剂(anti-miR-218),检测下调miR-218对LINC00339 siRNA影响的细胞增殖和凋亡的变化.结果 LINC00339在结直肠癌组织的表达显著高于癌旁组织(4.69±1.52比1.02±0.38,P＜0.05).LINC00339的表达与患者的年龄和性别无关(P＞0.05),而与患者的TNM分期、淋巴结转移、肿瘤长径和分化程度相关(均P＜0.05).在结直肠癌组织中LINC00339的表达与miR-218的表达呈负相关性.miR-218模拟物能够显著降低野生型LINC00339质粒的荧光强度(P=0.001),但是对突变型质粒的荧光强度无影响(P=0.88).在LoVo和HCT116细胞中下调LINC00339,细胞增殖能力受到抑制,细胞凋亡率增加(均P＜0.05);与仅转染LINC00339 siRNA相比,下调miR-218后LoVo和HCT116细胞的增殖能力增加、凋亡率降低(均P＜0.05).结论 LINC00339在结直肠癌中表达升高,与患者的临床病理特征密切相关.LINC00339通过下调miR-218表达促进结直肠癌细胞增殖,抑制凋亡.%Objective To investigate the expression of long non-coding RNA LINC00339 in colorectal cancer patients and its effect and mechanism on proliferation and apoptosis of colorectal cancer cells.Methods A retrospective analysis of 158 pathology-confirmed colorectal cancer patients,who were enrolled from August 2015 to January 2017,was performed.LINC00339 expression in colorectal cancer tissues and adjacent colorectal sampleswas detected by Real-time PCR.The correlation between LINC00339 expression and clinicopathological features as well as the relationship between LINC00339 and microRNA (miR)-218 expression was assayed.The interaction between LINC00339 and miR-218 was further confirmed by dual luciferase report system.Downregulation of LINC00339 was performed by siRNA interference technology in LoVo and HCT116 cells.Real-time PCR was used to detect miR-218 expression.3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) analysis was carried out to examine cell viability.Flow cytometry was used to determine cell apoptosis.Additionally,LINC00339 siRNA and miR-218 antagomirs (anti-miR-218) were co-transfected into LoVo and HCT116 cells,and then cell viability and apoptosis were detected.Resets LINC00339 expression was significantly increased in colorectal cancer tissues compared with adjacent colorectal tissues (4.69± 1.52 vs 1.02 ±0.38,P＜0.05).LINC00339 expression was not related to the age and gender of patients (P＞0.05),but was associated with TNM stage,lymphatic metastasis,tumor maximum diameters,and differentiation degree (all P＜0.05).LINC00339expression was negatively correlated with miR-218 expression in colorectal cancer tissues (P＜0.05).miR-218 mimics remarkably suppressed the fluorescence intensity of wild-type LINC00339 plasmid (P=0.001),but did not affect the fluorescence intensity of the mutant ones(P=0.88).Knockdown of LINC00339remarkably inhibited proliferation,but promoted apoptosis of LoVo and HCT116 cells (all P＜0.05).Compared with cells transfected with LINC00339siRNA only,downregulation of miR-218 elevated proliferation and decreased apoptosis of LoVoand HCT116 cells.Conclusions LINC00339 expression is upregulated in colorectal cancer tissues and correlated with patients' clinicopathological features.LINC00339 promotes proliferation,and suppresses apoptosis of colorectal cancer cells via downregulating miR-218.