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该文探讨了RORα(retinoid acid receptor related orphan receptorα)高表达对人胃癌MGC803细胞Wnt/β-catenin信号通路靶基因的作用。采用MTT检测了MGC803细胞增殖。采用RT-PCR、Western blot与免疫共沉淀检测了Wnt/β-catenin信号通路相关分子与靶基因表达。荧光素酶报告基因方法检测c-Myc基因启动子活性。MTT结果显示,RORα高表达人胃癌MGC803细胞的增殖能力较对照组明显减弱(P<0.05)。RT-PCR与Western blot结果显示,RORα高表达组Wnt1m RNA与蛋白质水平较对照组下调(P<0.05),而β-catenin m RNA与蛋白质水平无差异(P>0.05)。免疫共沉淀结果显示,RORα高表达组RORα与β-catenin结合明显增加(P<0.05)。RORα高表达可显著下调核内β-catenin水平(P<0.05),同时可显著下调TCF-4(T cell factor-4)蛋白质水平(P<0.05)。RORα高表达可显著下调Axin、c-Myc、c-Jun m RNA与蛋白质水平(P<0.05)。荧光素酶报告基因实验结果显示,RORα高表达c-Myc启动子活性明显降低(P<0.05)。以上结果表明,RORα高表达可通过调控Wnt/β-catenin信号通路相关分子基因表达来抑制人胃癌细胞增殖。
This article explored the effect of high expression of RORα (retinoid acid receptor related orphan receptorα) on Wnt / β-catenin signaling pathway target genes in human gastric cancer MGC803 cells. The proliferation of MGC803 cells was detected by MTT assay. RT-PCR, Western blot and co-immunoprecipitation were used to detect the expression of Wnt / β-catenin signaling pathway related molecules and target genes. Luciferase reporter assay for c-Myc promoter activity. MTT results showed that the proliferation ability of RORα overexpressing human gastric cancer MGC803 cells was significantly weaker than that of the control group (P <0.05). The results of RT-PCR and Western blot showed that Wnt1m RNA and protein levels in RORα-overexpression group were significantly lower than those in control group (P <0.05), while β-catenin mRNA and protein levels were not significantly different (P> 0.05). Co-immunoprecipitation showed that RORα and β-catenin increased significantly in RORα-overexpressing group (P <0.05). RORα overexpression significantly decreased the level of β-catenin in the nucleus (P <0.05) and significantly decreased the protein level of TCF-4 (P <0.05). RORα overexpression significantly down-regulated Axin, c-Myc, c-Jun mRNA and protein levels (P <0.05). The results of luciferase reporter assay showed that RORα overexpression c-Myc promoter activity was significantly decreased (P <0.05). The above results show that RORα overexpression can inhibit the proliferation of human gastric cancer cells by regulating the gene expression of Wnt / β-catenin signaling pathway.