NEFL通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞的侵袭迁移

doi:10.16288/j.yczz.22-019

遗传 ›› 2022, Vol. 44 ›› Issue (4): 322-334.doi: 10.16288/j.yczz.22-019

NEFL通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞的侵袭迁移

范志露¹(), 杨荔艳²(), 张娜¹, 冯丹¹, 郭静¹, 常晨¹, 苑青¹, 蔡岩¹, 张钰¹, 魏文强³, 王明荣¹, 郝佳洁¹()

1. 国家癌症中心/国家肿瘤临床医学研究中心,中国医学科学院北京协和医学院肿瘤医院,分子肿瘤学国家重点实验室,北京 100021
2. 国家癌症中心/国家肿瘤临床医学研究中心,中国医学科学院北京协和医学院肿瘤医院检验科,北京 100021
3. 国家癌症中心/国家肿瘤临床医学研究中心,中国医学科学院北京协和医学院肿瘤医院流行病学中心,北京 100021

收稿日期:2022-01-24 修回日期:2022-02-22 出版日期:2022-04-20 发布日期:2022-03-07
通讯作者: 杨荔艳,郝佳洁 E-mail:18679148679@163.com;yangliyan0729@163.com;hjj8173@126.com
作者简介:范志露,在读硕士研究生,专业方向：细胞生物学。E-mail: 18679148679@163.com
基金资助:
国家自然科学基金项目编号(81972770);中国医学科学院医学与健康科技创新工程编号(2021-I2M-1-018);中国医学科学院肿瘤医院潜力培育计划资助编号(PY2018B01)

NEFL promotes invasion and migration of esophageal squamous carcinoma cells via the EGFR/AKT/S6 pathway

Zhilu Fan¹(), Liyan Yang²(), Na Zhang¹, Dan Feng¹, Jing Guo¹, Chen Chang¹, Qing Yuan¹, Yan Cai¹, Yu Zhang¹, Wenqiang Wei³, Mingrong Wang¹, Jiajie Hao¹()

1. State Key Laboratory of Molecular Oncology, Center for Cancer Precision Medicine, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
2. Department of Clinical Laboratory, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
3. Department of Cancer Epidemiology, Center for Cancer Precision Medicine, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China

Received:2022-01-24 Revised:2022-02-22 Online:2022-04-20 Published:2022-03-07
Contact: Yang Liyan,Hao Jiajie E-mail:18679148679@163.com;yangliyan0729@163.com;hjj8173@126.com
Supported by:
Supported by the National Natural Science Foundation of China No(81972770);Chinese Academy of Medical Sciences (CAMS) Innovation Fund for Medical Sciences No(2021-I2M-1-018);the Potential Development Projects of Cancer Hospital, CAMS No(PY2018B01)

摘要/Abstract

摘要：

为探讨神经丝轻链(neurofilament light chain, NEFL)在食管鳞癌(esophageal squamous cell carcinoma, ESCC)中的表达情况及作用机制,本研究首先对食管鳞癌组织及配对的正常食管上皮中NEFL的mRNA和蛋白表达情况进行了检测。基因表达综合数据库(Gene Expression Omnibus, GEO) RNA表达数据,以及临床标本的实时荧光定量逆转录-聚合酶链式反应(real-time quantitative reverse transcription PCR, qRT-PCR)分析结果显示,与正常食管上皮组织相比,食管鳞癌组织中NEFL基因mRNA水平明显升高。Western blot分析结果显示,与正常食管上皮组织相比,食管鳞癌组织中NEFL蛋白水平也明显升高。进一步采用CCK8法和Transwell法检测NEFL过表达对食管鳞癌细胞恶性表型的影响,结果显示敲降NEFL后,食管鳞癌细胞的侵袭和迁移能力显著减弱。体内裸鼠成瘤实验显示,敲降NEFL可显著抑制肿瘤生长。分子水平检测表明,敲降NEFL显著升高上皮间质转化(epithelial-mesenchymal transition, EMT)相关分子E-钙黏蛋白(E-cadherin)的表达并降低N-钙黏蛋白(N-cadherin)的表达、下游分子表皮生长因子受体(epidermal growth factor receptor, EGFR)的mRNA和蛋白的表达水平,蛋白激酶B (protein kinase B, PKB;又被称为AKT)和核糖体蛋白S6 (ribosomal protein S6)的磷酸化水平也显著降低。敲降NEFL后转染EGFR过表达载体,AKT和S6的磷酸化水平以及食管鳞癌细胞侵袭和迁移表型都得以恢复。以上研究结果表明,NEFL过表达可能通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞EMT,最终促进食管鳞癌细胞的侵袭迁移。

关键词: NEFL, 食管鳞癌, 侵袭迁移, EGFR, 上皮间质转化

Abstract:

To explore the expression, the roles and the underlying mechanism of neurofilament light chain (NEFL) in esophageal squamous cell carcinoma (ESCC), we firstly analyzed the NEFL mRNA and protein expression in ESCC and paired normal tissues by using Gene Expression Omnibus (GEO) database, and real-time quantitative reverse transcription PCR (qRT-PCR). The results showed that NEFL mRNA level was significantly upregulated in ESCC tissues compared with that of normal tissues. Western blot analysis revealed that NEFL protein level was also significantly upregulated in ESCC tissues. CCK8 and transwell assays were performed to analyze the effect of NEFL overexpression on the malignant phenotypes of ESCC cells, and the results showed that NEFL knockdown significantly impaired the ESCC cell invasion and migration in vitro. Xenograft assay in nude mice indicated that NEFL silencing suppressed tumor growth in vivo. At the molecular level, NEFL knockdown significantly upregulated E-cadherin and downregulated N-cadherin expression, suggesting that NEFL overexpression might influence the epithelial-mesenchymal transition (EMT) process. Furthermore, we found that NEFL knockdown significantly reduced the mRNA and protein expression of epidermal growth factor receptor (EGFR) and the phosphorylation levels of protein kinase B (PKB; also known as AKT) and ribosomal protein S6 (S6). Ectopic expression of EGFR after NEFL knockdown significantly restored the phosphorylation levels of AKT and S6 as well as the invasion and migration of ESCC cells. These data indicate that NEFL overexpression might promote the EMT process of ESCC cells via the EGFR/AKT/S6 pathway, ultimately enhancing the invasion and migration of ESCC cells.

Key words: NEFL, esophageal squamous cell carcinoma, invasion and migration, EGFR, EMT

范志露, 杨荔艳, 张娜, 冯丹, 郭静, 常晨, 苑青, 蔡岩, 张钰, 魏文强, 王明荣, 郝佳洁. NEFL通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞的侵袭迁移[J]. 遗传, 2022, 44(4): 322-334.

Zhilu Fan, Liyan Yang, Na Zhang, Dan Feng, Jing Guo, Chen Chang, Qing Yuan, Yan Cai, Yu Zhang, Wenqiang Wei, Mingrong Wang, Jiajie Hao. NEFL promotes invasion and migration of esophageal squamous carcinoma cells via the EGFR/AKT/S6 pathway[J]. Hereditas(Beijing), 2022, 44(4): 322-334.

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NEFL通过激活EGFR/AKT/S6信号通路促进食管鳞癌细胞的侵袭迁移

NEFL promotes invasion and migration of esophageal squamous carcinoma cells via the EGFR/AKT/S6 pathway

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