lncRNA COX10-AS1在肾癌组织中的表达及对肾癌细胞增殖和迁移的影响

Expression of lncRNA COX10-AS1 in renal cell carcinoma tissues and its effect on proliferation and migration of renal carcinoma cells

目的探讨长链非编码RNA(lncRNA)COX10-AS1在肾癌组织和细胞系中的表达及其对肾癌细胞增殖和迁移的影响。方法荧光实时定量PCR(qRT-PCR)检测经病理确诊为肾癌组织和癌旁组织的手术标本、肾癌细胞系(786-O、CaKi-1、A498、ACHN)和正常肾小管上皮细胞系(HK-2)中COX10-AS1的表达。以表达最低的ACHN细胞为研究对象,分为对照组和实验组,对照组细胞转染载有无意义序列的阴性对照质粒,实验组细胞转染载有COX10-AS1表达序列的质粒。qRT-PCR检测两组细胞中COX10-AS1的表达水平,MTS法和细胞划痕实验检测ACHN细胞的增殖和迁移能力。qRT-PCR检测两组细胞中MFN2 mRNA的表达水平,免疫印迹法(Western blotting)检测两组细胞中MFN2、Ras-NF-κB信号通路蛋白的表达情况。计量资料以均数±标准差(Mean±SD)表示,两组间比较采用t检验,多组间比较采用单因素方差分析。结果肾癌组织COX10-AS1表达明显低于癌旁组织(P<0.01),肾癌细胞系COX10-AS1表达明显低于肾小管上皮细胞(P<0.05),ACHN细胞中COX10-AS1的表达最低(P<0.01),上述差异均具有统计学意义。与对照组细胞相比,实验组ACHN细胞中COX10-AS1的表达明显增加,差异具有统计学意义(P<0.01)。与对照组细胞相比,实验组ACHN细胞增殖活力明显降低,细胞迁移能力明显降低,差异均具有统计学意义(P<0.01)。与对照组细胞相比,实验组ACHN细胞中MFN2 mRNA的表达明显增加(P<0.01),MFN2蛋白表达明显增加(P<0.01),Ras-NF-κB信号通路蛋白表达明显降低(P<0.05),上述差异均具有统计学意义。结论COX10-AS1在肾癌组织和细胞系中表达降低,COX10-AS1可能通过促进MFN2基因的表达,抑制ACHN细胞的增殖和迁移能力。

Objective To investigate the expression of long non-coding RNA(lncRNA)COX10-AS1 in renal cell carcinoma tissues and cell lines and its effect on proliferation and migration of renal cancer cells.Methods Fluorescence real-time quantitative PCR(qRT-PCR)was used to detect the expression of COX10-AS1 in surgical specimens that have been diagnosed as renal cancer tissues and adjacent tissues by pathology,renal cancer cell lines(786-O,CaKi-1,A498,ACHN)and normal renal tubular epithelium cell line(HK-2).The ACHN cells with the lowest expression were divided into a control group(transfected with a negative control plasmid carrying nonsense sequences)and an experimental group(transfected with a plasmid carrying COX10-AS1 sequences).The expression level of COX10-AS1 was detected by qRT-PCR in two groups of cells.The proliferation and migration ability of ACHN cells were detected by MTS assay and cell scratch assay.The expression of MFN2 mRNA was detected by qRT-PCR.The expressions of MFN2 and Ras-NF-κB signaling pathway proteins were detected by Western blotting.The measurement data were expressed as mean±standard deviation(Mean±SD),the comparison between the two groups used the t-test,and the comparison among multiple groups adopts the one-way analysis of variance.Results The expression of COX10-AS1 in renal cell carcinoma was significantly lower than that in adjacent tissues(P<0.01),The expression of COX10-AS1 in renal cell carcinoma cells was significantly lower than that in renal tubular epithelial cells(P<0.05),the expression of COX10-AS1 was the lowest in ACHN cells(P<0.01),the above differences were statistically significant compared with the control group,the expression of COX10-AS1 in ACHN cells of experimental group was significantly increased(P<0.01),the above differences were statistically significant compared with the control cells,the proliferation of ACHN cells in the experimental group was significantly decreased(P<0.05),and the cell migration ability was significantly decreased(P<0.01).Compared with the control cells,the expression of MFN2 mRNA in ACHN cells of experimental group was significantly increased(P<0.01).The expression levels of MFN2 were significantly up-regulated(P<0.01),and Ras-NF-κB signaling pathway proteins were significantly down-regulated(P<0.05),the above differences were statistically significant.Conclusions The expression of COX10-AS1 is decreased in renal cell carcinoma tissues and cell lines.COX10-AS1 may inhibit the proliferation and migration of ACHN cells by promoting the expression of MFN2 gene.