摘要
目的:探讨长链非编码RNA(lncRNA)SUCLG2-AS1对非小细胞肺癌细胞增殖和免疫逃逸的影响及对微小RNA-491-5p(miR-491-5p)的调节作用。方法:通过Cancer LncRNA Census数据库分析SUCLG2-AS1在非小细胞肺癌组织中的表达及SUCLG2-AS1表达量与非小细胞肺癌患者生存期的关系。通过实时荧光定量PCR(qRT-PCR)检测SUCLG2-AS1在非小细胞肺癌株A549、PG49、H1975、H2170和人正常肺支气管上皮细胞BEAS-2B中表达量的变化。培养H1975细胞并分为si-NC组和si-SUCLG2-AS1组。集落实验和流式细胞术检测H1975细胞的增殖能力和细胞周期。蛋白质印迹(Western blot)检测H1975细胞免疫相关蛋白细胞程序性死亡-配体1(PD-L1)、维甲酸相关孤儿受体γt(RORγt)和细胞周期相关蛋白细胞周期依赖性蛋白激酶3(Cdk3)、细胞周期蛋白C(Cyclin C)、细胞周期依赖性蛋白激酶2(Cdk2)的表达。LncRMap生物信息学软件预测SUCLG2-AS1可结合的微小RNA。RNA pull-down实验验证SUCLG2-AS1与miR-491-5p的直接结合。通过qRT-PCR检测SUCLG2-AS1对miR-491-5p表达的调节作用。结果:与正常肺组织比较,非小细胞肺癌组织中SUCLG2-AS1表达升高(P<0.01)。SUCLG2-AS1高表达的非小细胞肺癌患者生存期短于SUCLG2-AS1低表达患者(P<0.01)。与BEAS-2B细胞比较,非小细胞肺癌株中SUCLG2-AS1表达升高,H1975细胞中SUCLG2-AS1表达升高最明显(均P<0.05)。与si-NC组比较,si-SUCLG2-AS1组H1975细胞增殖能力降低,G0/G1期细胞比例上升,S期细胞比例减少,G2/M期细胞比例减少(均P<0.01)。与si-NC组比较,si-SUCLG2-AS1组H1975细胞中免疫相关蛋白PD-L1、RORγt表达减少,细胞周期相关蛋白Cdk3、Cyclin C、Cdk2表达减少(均P<0.01)。SUCLG2-AS1靶向直接结合miR-491-5p(均P<0.01)。与si-NC组比较,si-SUCLG2-AS1组H1975细胞中miR-491-5p表达上升(P<0.01)。结论:沉默SUCLG2-AS1可能通过上调miR-491-5p表达抑制非小细胞肺癌细胞的增殖和免疫逃逸。
Objective:To explore the effect of long non-coding RNA(lncRNA)SUCLG2-AS1 on the proliferation and immune evasion of non-small cell lung cancer cells and its regulatory effect on miR-491-5p.Methods:The Cancer LncRNA Census database was used to analyze the expression of SUCLG2-AS1 in non-small cell lung cancer tissues and the relationship between SUCLG2-AS1 expression and the survival of patients with non-small cell lung cancer.The expression changes of SUCLG2-AS1 in non-small cell lung cancer lines A549,PG49,H1975,H2170 and human normal lung bronchial epithelial cells BEAS-2B were detected by qRT-PCR.H1975 cells were cultured and divided into si-NC group and si-SUCLG2-AS1 group.Colony experiment and flow cytometry were used to detect the proliferation ability and cell cycle of H1975 cells.Western blot was used to detect the expression of immune-related proteins programmed cell death 1 ligand 1(PD-L1),RAR-related orphan receptor(RORγt)and cell cycle-related proteins cyclin dependent kinase3(Cdk3),Cyclin C,and cyclin dependent kinase2(Cdk2)in H1975 cells.LncRMap bioinformatics software was used to predict microRNA that SUCLG2-AS1 can bind.RNA pull-down experiments was used to verify the direct binding of SUCLG2-AS1 to miR-491-5p.The regulatory effect of SUCLG2-AS1 on the expression of miR-491-5p was detected by qRT-PCR.Results:Compared with normal lung tissue,the expression of SUCLG2-AS1 was increased in non-small cell lung cancer tissue(P<0.01).The survival time of non-small cell lung cancer patients with high SUCLG2-AS1 expression was shorter than that of patients with low SUCLG2-AS1 expression(P<0.01).Compared with BEAS-2B cells,the expression of SUCLG2-AS1 was increased in non-small cell lung cancer lines,and the expression of SUCLG2-AS1 was most significantly increased in H1975 cells(all P<0.05).Compared with the si-NC group,the proliferation ability of H1975 cells in the si-SUCLG2-AS1 group decreased,the proportion of cells in G 0/G 1 phase increased,the proportion of cells in S phase decreased,and the proportion of cells in G 2/M phase decreased(all P<0.01).Compared with the si-NC group,the expression of immune-related proteins PD-L1 and RORγt in H1975 cells in the si-SUCLG2-AS1 group was reduced,and the expression of cell cycle-related proteins Cdk3,Cyclin C,and Cdk2 was reduced.SUCLG2-AS1 targets and directly binds to miR-491-5p(all P<0.01).Compared with the si-NC group,the expression of miR-491-5p in H1975 cells in the si-SUCLG2-AS1 group increased(P<0.01).Conclusion:Silencing SUCLG2-AS1 may inhibit the proliferation and immune evasion of non-small cell lung cancer cells by upregulating the expression of miR-491-5p.
作者
陈娟
郭金莲
周慧会
刘琳
杨柳
张华
CHEN Juan;GUO Jinlian;ZHOU Huihui;LIU Lin;YANG Liu;ZHANG Hua(Department of Respiratory Medicine,Jingmen People’s Hospital,Jingmen 448000,China)
出处
《陕西医学杂志》
CAS
2024年第7期884-889,共6页
Shaanxi Medical Journal
基金
国家自然科学基金资助项目(81860413)。
