lncRNA AC068768.1靶向miR-21-5p调控肾癌细胞周期和增殖

lncRNA AC068768.1 regulates the cycle and proliferation of renal cancer cells by targeting miR-21-5p

目的探讨长链非编码RNA(lncRNA)AC068768.1对肾癌细胞周期和增殖能力的影响及分子机制。方法采用实时定量聚合酶链反应(qPCR)检测AC068768.1在肾癌细胞系中的表达情况。以AC068768.1表达最低的OS-RC-2细胞为转染对象,设转染阴性对照质粒的OS-RC-2为对照组,转染AC068768.1质粒为AC068768.1组。qPCR检测转染OS-RC-2细胞中AC068768.1的表达。通过流式细胞术和四甲基偶氮唑蓝比色(MTT)增殖实验检测AC068768.1对OS-RC-2细胞周期和增殖能力的影响。生物信息学方法预测AC068768.1可能结合的微小RNA(miRNA)。qPCR检测miRNA及下游基因mRNA的表达,Western blotting法检测下游基因蛋白的表达。计量资料以均数±标准差(Mean±SD)表示,组间比较采用独立样本t检验,多组间比较采用单因素方差分析。结果与正常肾小管上皮细胞相比,AC068768.1在肾癌细胞系中的表达显著降低,差异明显具有统计学意义(P<0.01)。AC068768.1组OS-RC-2细胞AC068768.1的表达显著高于对照组,差异明显具有统计学意义(P<0.01)。上调AC068768.1表达可抑制肾癌细胞周期(P<0.05)和增殖能力(P<0.05)。miR-21-5p可能是AC068768.1的功能性靶基因。上调AC068768.1可明显抑制miR-21-5p表达(P<0.01),促进金属蛋白酶组织抑制因子3(TIMP3)基因表达(P<0.01)。结论AC068768.1通过调节miR-21-5p表达,促进TIMP3基因表达,进而抑制肾癌OS-RC-2细胞周期和增殖能力。

Objective To explore the effect of long non-coding RNA(lncRNA)AC068768.1 on the cycle and proliferation of renal cancer cells and its molecular mechanism.Methods Real-time quantitative polymerase chain reaction(qPCR)was used to detect the expression of AC068768.1 in renal cancer cell lines.The OS-RC-2 cells with the lowest expression of AC068768.1 were used as the transfection objects,OS-RC-2 transfected with the negative control plasmid was set as the control group,and the cells transfected with the AC068768.1 plasmid were set as the AC068768.1 group.qPCR was used to detect the expression of AC068768.1 in transfected OS-RC-2 cells.The effects of AC068768.1 on the cell cycle and proliferation of OS-RC-2 were detected by flow cytometry and tetramethylazazole blue colorimetric(MTT)proliferation experiments.Using bioinformatics methods to predict the microRNA(miRNA)that AC068768.1 may bind.qPCR was used to detect the expression of miRNA and downstream gene mRNA,and Western blot was used to detect the expression of downstream gene protein.The measurement data were expressed as mean±standard deviation(Mean±SD),the comparison between the two groups adopts the t-test,and the comparison among multiple groups adopts the One-way analysis of variance.Results Compared with normal renal tubular epithelial cells,the expression of AC068768.1 in renal cancer cell lines was significantly reduced,the difference was statistically significant(P<0.01).The expression of AC068768.1 in OS-RC-2 cells in the AC068768.1 group was significantly higher than that in the control group,the difference was statistically significant(P<0.01).Up-regulating the expression of AC068768.1 can inhibit the cycle(P<0.05)and proliferating ability(P<0.05)of renal cancer cells.miR-21-5p may be the functional target gene of AC068768.1.Up-regulation of AC068768.1 can significantly inhibit the expression of miR-21-5p(P<0.01)and promote the expression of tissue inhibitor of metalloproteinase 3(TIMP3)(P<0.01).Conclusion AC068768.1 promotes the expression of TIMP3 gene by regulating the expression of miR-21-5p,thereby inhibiting the cell cycle and proliferation of renal cancer OS-RC-2 cells.