沉默lncRNA RNF185-AS1通过靶向miR-637/HMGA1抑制胶质瘤细胞增殖和侵袭

Silencing the expression of lncRNA RNF185-AS1 inhibits the proliferation and invasion of glioma cells by targeting miR-637/HMGA1

目的研究长链非编码RNA(lncRNA)RNF185-AS1调控微小RNA-637(miR-637)对胶质瘤细胞增殖、侵袭的作用机制。方法通过GEPIA数据库在线分析胶质瘤组织和癌旁组织中RNF185-AS1的表达水平。qRT-PCR检测胶质瘤细胞系(SNB-19、LN382、U87MG、U251)和脑星型胶质正常细胞HEB中RNF185-AS1的表达水平。以U251细胞为研究对象,分别转染sh-Con对照质粒(sh-Con组)和sh-RNF185-AS1质粒(sh-RNF185-AS1组)。qRT-PCR验证RNF185-AS1的沉默效率。MTT实验和Transwell侵袭实验检测沉默RNF185-AS1后U251细胞增殖活力和侵袭情况。qRT-PCR与双荧光素酶报告基因实验检测RNF185-AS1和miR-637的靶向关系。qRT-PCR和Western blot检测高迁移率族蛋白A1(HMGA1)基因的表达。结果胶质瘤组织中RNF185-AS1的表达水平明显高于癌旁组织(P<0.01)。胶质瘤细胞系中RNF185-AS1的表达水平明显高于脑星型胶质正常细胞(P<0.01),U251细胞中RNF185-AS1表达升高最明显(P<0.01)。与sh-Con组相比,sh-RNF185-AS1组U251细胞中RNF185-AS1的表达水平显著下降(P<0.01),U251细胞增殖活力明显降低(P<0.05),侵袭细胞数目明显较少(P<0.01)。RNF185-AS1与miR-637存在互补结合的核苷酸序列(P<0.01)。与sh-Con组相比,sh-RNF185-AS1组U251细胞中miR-637的表达水平明显增加(P<0.01),HMGA1基因的表达水平明显降低(P<0.01)。结论RNF185-AS1在胶质瘤组织和细胞系中高表达,沉默RNF185-AS1通过靶向上调miR-637表达、下调HMGA1基因表达,抑制胶质瘤U251细胞的增殖和侵袭能力。

Objective To study the mechanism of long non-coding RNA(lncRNA)RNF185-AS1 affecting the proliferation and invasion of glioma cells by regulating microRNA-637(miR-637).Methods The GEPIA database was used to analyze the expression level of RNF185-AS1 in glioma tissues and adjacent tissues.qRT-PCR was used to detect the expression level of RNF185-AS1 in glioma cell lines(SNB-19,LN382,U87 MG,U251)and normal brain astrocytes HEB.U251 cells were used as the research object,and sh-Con control plasmid(sh-Con group)and sh-RNF185-AS1 plasmid(sh-RNF185-AS1 group)were transfected respectively.qRT-PCR verified the silencing efficiency of RNF185-AS1.MTT experiment and Transwell invasion experiment were used to detect the proliferation and invasion of U251 cells after silencing RNF185-AS1.qRT-PCR and dual luciferase reporter gene experiment were used to detect the targeting relationship between RNF185-AS1 and miR-637.qRT-PCR and Western blot were used to detect the expression of high mobility group A1(HMGA1)gene.Results The expression level of RNF185-AS1 in glioma tissue was significantly higher than that in adjacent tissues(P<0.01).The expression level of RNF185-AS1 in the glioma cell line was significantly higher than that in normal brain astrocytes(P<0.01),and the expression of RNF185-AS1 in U251 cells increased the most significantly(P<0.01).Compared with the sh-Con group,the expression level of RNF185-AS1 in U251 cells in the sh-RNF185-AS1 group significantly decreased(P<0.01),the proliferation activity of U251 cells was significantly reduced(P<0.05),and the number of invaded cells was significantly less(P<0.01).RNF185-AS1 and miR-637 had complementary nucleotide sequences(P<0.01).Compared with the sh-Con group,the expression level of miR-637 in U251 cells in the sh-RNF185-AS1 group significantly increased(P<0.01),and the expression level of HMGA1 gene was significantly reduced(P<0.01).Conclusion RNF185-AS1 is highly expressed in glioma tissues and cell lines.Silencing RNF185-AS1 can inhibit the proliferation and invasion of glioma U251 cells by targeting up-regulation of miR-637 expression and down-regulation of HMGA1 gene expression.