利用食管类器官研究c-Myc在食管癌发生中的作用  

Using esophagus organoid to explore the role of c-Myc in esophageal cancer initiation

作　　者：陈友红 杨文豪 倪超 Youhong Chen;Wenhao Yang;Chao Ni(School of Life Sciences Fudan University,Shanghai 200433,China;Department of Pediatrics,West China Second University Hospital,Sichuan University,Chengdu 610041,China)

机构地区：[1]复旦大学生命科学学院,上海200433 [2]四川大学华西第二医院小儿呼吸免疫科,成都610041

出　　处：《遗传》2021年第6期601-614,I0003,共15页Hereditas（Beijing)

基　　金：国家自然科学基金青年项目(编号:31900581)资助。

摘　　要：c-Myc基因在食管癌等多种恶性肿瘤中异常高表达,但其参与癌症发生发展机制尚不完全清楚。为探究c-Myc在食管癌发生中的作用,本文成功构建了食管类器官作为研究模型,首先制备表达c-Myc的慢病毒并通过高效侵染方法获得了稳定过表达c-Myc的食管类器官。以正常食管类器官作为对照,使用ImageJ软件分析感染7代后食管类器官的形态,显示食管类器官的上皮形态并未出现异常。随后利用免疫荧光染色实验和CCK8试剂检测食管类器官的细胞增殖状态,结果显示细胞增殖速度也没有显著改变。最后通过实时荧光定量PCR实验(quantitative real-time PCR,qPCR)检测细胞周期、细胞代谢以及常见的在食管癌中高表达基因的表达情况,结果显示相关基因表达均未显著升高。这些结果初步表明在食管中单独过表达c-Myc基因不足以诱导食管上皮细胞癌化。本文建立了食管类器官研究模型,通过高效的慢病毒过表达体系研究了原癌基因c-Myc对食管类器官发育和增殖的潜在影响。这对于食管类器官模拟食管发育和食管癌发生相关研究具有一定参考意义。C-Myc gene is aberrantly highly expressed and participates in cancer initiation and development in various malignant tumors including esophageal cancer,while the underlying mechanism(s)still remains unclear.In order to explore the role of c-Myc in the occurrence of esophageal cancer,we successfully established the esophageal organoids(EOs)as the research model.By constructing a lentivirus overexpressing c-Myc and developing more effective infection method,EOs with stable overexpression of c-Myc were efficiently obtained.The morphologies of EOs with or without overexpressing c-Myc were first analyzed with ImageJ,which showed no difference between two groups during continuous subculture.Subsequently,we applied immunofluorescence and CCK8 assays to evaluate the cell proliferation,and the results showed no change in the c-Myc-overexpressed group as compared to control EOs.Furthermore,qPCR was used to detect the expression of genes that are related to cell cycle,cell metabolism as well as esophageal cancer.The results indicated the expression of these genes was not significantly increased in the c-Myc overexpressing EOs.In conclude,we discovered that overexpression of c-Myc gene alone in the esophagus organoid is not sufficient to induce carcinogenesis in esophageal carcinoma.In this study,we successfully established an esophagus organoid culture system and together with efficient lentivirus-infection method for investigation on the effects of overexpressing c-Myc in esophageal cancer.Our work demonstrated a promising research model for the study of esophagus development and esophageal cancer.

关 键 词：食管癌 食管类器官 c-Myc过表达 慢病毒侵染 

分 类 号：R735.1[医药卫生—肿瘤]