RPL30在肝细胞肝癌中的表达及对其进展的影响

The expression of RPL30 in hepatocellular carcinoma and its effects on hepatocellular carcinoma progression

目的 研究核糖体蛋白RPL30在肝细胞肝癌(Hepatocellular carcinoma,HCC)发生发展中的作用.方法 利用GEPIA网站预测RPL30在HCC组织中的表达水平,通过qRT-PCR、蛋白免疫印迹及免疫组织化学染色实验验证其表达.构建稳定敲低和过表达RPL30的MHC97H细胞系并利用该细胞系进行体外实验.通过CCK-8和平板克隆实验探究RPL30对HCC细胞增殖能力的影响;通过划痕和transwell实验研究其对细胞迁移和侵袭能力的改变;利用干细胞成球实验和流式细胞术检测HCC细胞肿瘤干性;最后通过蛋白免疫印迹实验确定RPL30的表达对ERK通路的影响.结果 GEPIA数据库的分析结果提示,RPL30在HCC组织高表达,通过qRT-PCR、westem blot及免疫组织化学染色实验对本中心收集的HCC样本检测后得到了一致的结果.CCK-8实验结果显示,RPL30的过表达的细胞增殖速度显著高于对照组.平板克隆实验结果显示,过表达RPL30的MHC97H细胞克隆形成数目明显增多.划痕实验结果表明,RPL30的过表达的HCC细胞迁移距离明显大于转染对照质粒的细胞.Transwell实验提示,过表达RPL30后细胞穿过小室的数目显著增加.与对照组相比,RPL30过表达的MHC97H细胞在干细胞成球实验中成球数目明显增加,CD133的阳性比例升高.此外,RPL30的过表达导致MEK1/2和ERK的磷酸化水平升高,ERK通路激活.结论 RPL30在HCC组织中高表达,其表达水平的升高会增强HCC细胞增殖、迁移、侵袭能力以及肿瘤干性,上调ERK通路的水平.

Objective The aim of this study is to find out the role of ribosomal protein RPL30 in the occurrence and develop-ment of hepatocellular carcinoma(HCC).Method The GEPIA database was used to predict the expression of RPL30 in HCC tis-sues,and its expression was verified through qRT-PCR,western blot,and immunohistochemistry staining.We constructed a stable knockdown and overexpression of RPL30 MHC97H cell line and used these cells for in vitro experiments.CCK-8 and colony formation experiments were used to determine the effect of RPL30 on the proliferation ability of HCC cells.Wound healing and transwell assays were applied to study the RPL30 influence on migration and invasion abilities of HCC cell.The stemness of HCC cells was determined by spheroid formation assay and flow cytometry.Finally,western blot was used to determine the influence of RPL30 on the ERK path-way.Result The analysis results of the GEPIA database indicated that RPL30 was upregulated in HCC tissues.The HCC samples collected in our department were detected using qRT-PCR,western blot,and immunohistochemical staining,and consistent results were obtained.The CCK-8 experiment results showed that the proliferation ability of RPL30 overexpressed cells was significantly promo-ted compared with control groups.The results of the colony formation experiment showed that the number of clones formed in MHC97H cells with RPL30 overexpression significantly increased.The wound healing assay results showed that the migration distance of HCC cells overexpressed with RPL30 was dramatically greater than that of cells transfected with the control plasmid.The results of transwell experiment suggested that overexpression of RPL30 significantly increased the number of cells passing through chambers.Compared with the control groups,MHC97H cells overexpressed with RPL30 showed a significant increase in the number of spheroids in stem cell spheroid formation experiments and a higher proportion of CD133+cells.In addition,overexpression of RPL30 leads to upregulated phosphorylation levels of MEK1/2 and ERK.Conclusion RPL30 is upregulated in HCC tissues,and its increased expression level enhances the proliferation,migration,invasion ability,and tumor stemness of HCC cells,activating the ERK pathway.