作者：杨光谱，叶锦宁，陈剑辉，陈斯乐，叶志君，许开武，王昭，陈创奇，蔡世荣，马晋平

单位：中山大学附属第一医院 胃肠外科中心，广东 广州 510080

Authors:  YANG Guangpu, YE Jinning, CHEN Jianhui, CHEN Sile, YE Zhijun, XU Kaiwu, WANG Zhao, CHEN Chuangqi,  CAI Shirong, MA Jinping

Unit:  Department of Gastrointestinal Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, Guangdong, China

摘要：

目的 探讨circ-ITGA7是否通过上调ASXL1抑制结直肠癌细胞增殖。方法 采用RT-qPCR检测结直肠癌及癌旁组织中circ-ITGA7和ASXL1 mRNA表达水平,并用皮尔森相关性分析检测其相关性。RT-qPCR检测结直肠癌细胞株与正常肠道粘膜细胞株circ-ITGA7表达差异。采用质粒转染法在HCT116细胞中过表达circ-ITGA7,以RT-qPCR和western blot检测ASXL1的mRNA与蛋白表达水平。Cck-8对比circ-ITGA7过表达与否的HCT116细胞7天内的增殖活性。通过转染siRNA与质粒,以HCT116细胞集落形成实验对比过表达circ-ITGA7和沉默ASXL1对HCT116细胞集落形成数量的的影响。结果 RT-qPCR结果显示,circ-ITGA7在结直肠癌组织与细胞株中表达显著低于癌旁组织和正常肠道粘膜组织细胞株,且其在结直肠癌组织中与ASXL1的表达呈正相关性。过表达circ-ITGA7组的ASXL1蛋白及mRNA的表达水平显著高于对照组。Cck-8检测示,过表达circ-ITGA7组的吸光度于第5、6、7天显著低于对照组。细胞集落形成实验示,过表达circ-ITGA7组HCT116细胞的集落形成数量显著低于对照组,circ-ITGA7过表达且si-ASXL1沉默组与对照组差异无统计学意义。结论 Circ-ITGA7通过上调ASXL-1抑制结直肠癌细胞增殖。

关键词：  Circ-ITGA7; ASXL-1; 结直肠癌

Abstract：

Objective  This study aims to verify if circ-ITGA7 inhibits proliferation of HCT116 cells via upregulating ASXL1. Methods  RT-qPCR was employed to compare the RNA expression level of circ-ITGA7 and ASXL1 in colorectal cancer and para-cancerous tissue. Pearson’s Correlation Test was used to assess their correlation. RT-qPCR was used to test the circ-ITGA7 expression levels in HCT116、HT29、DLD1、SW480 and NCM460 cells. circ-ITGA7 vectors were transfected to build over-expressed HCT116 cells. Protein and mRNA expression were tested by RT-qPCR and western blot. The effect of circ-ITGA7 on proliferation in HCT116 cells was tested by cck-8 kit. Colonies formation assay was employed to assess the effect of circ-ITGA7 over-expression and ASXL1 silence on the colony formation of HCT116 cells. Results  Compared with para-cancerous tissue and NCM460 cells, the expression of circ-ITGA7 is significantly lower in colorectal cancer and in in-vitro colorectal cancer cell lines respectively. Cric-ITGA7 is positively corelated with the mRNA expression of ASXL1. Overexpression of circ-ITGA7 statistically generates more ASXL1 mRNA and protein than blank group. Overexpression of circ-ITGA7 generates significantly more OD on 5th, 6th and 7th day than control group. The number of colonies formatted by the circ-ITGA7 overexpressed cells was significantly lower than control group. Co-transfection of circ-ITGA7 and si-ASXL1 makes more colonies than transfection of circ-ITGA7 only. Conclusion  circ-ITGA7 inhibits HCT116 pro liferation via upregulating ASXL1.

Key Words:  circ-ITGA7; ASXL1; Colorectal cancer