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中国癌症防治杂志

中国癌症防治杂志 ›› 2020, Vol. 12 ›› Issue (6): 650-656.doi: 10.3969/j.issn.1674-5671.2020.12.10

• 基础研究 • 上一篇    下一篇

TRPM8对肝细胞癌Huh7细胞侵袭和转移的影响及其作用机制

  

  1. 830054 乌鲁木齐 新疆医科大学第一附属医院消化二科
  • 出版日期:2020-12-25 发布日期:2021-01-08
  • 作者简介:张志强， E-mail: drzhiqiang@163.com
  • 基金资助:
    省部共建中亚高发病成因与防治国家重点实验室开放课题项目（SKL-HIDCA-2018）

Effect of TRPM8 on the invasion and metastasis of hepatocellular carcinoma Huh7 cells and its mechanism

  1. Department of Gastroenterology, the First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, China
  • Online:2020-12-25 Published:2021-01-08

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摘要： 目的 探讨顺时受体电位8（chronic receptor potential 8，TRPM8）对肝细胞癌（hepatocellular carcinoma，HCC）细胞侵袭和转移的影响及其调控机制。方法 收集2017年3月—2019年12月于本院手术切除的20对配对HCC组织及其癌旁组织，采用免疫组化和qRT-PCR检测TRPM8表达情况。用shTRPM8转染HCC Huh7细胞（shTRPM8组），并设阴性对照组（shcontrol组），采用划痕愈合实验和Transwell小室实验检测细胞迁移和侵袭能力；免疫荧光实验检测E-cadherin和vimentin表达情况；Western blot检测AFP、TRPM8及AKT/GSK-3β/Snail信号途径相关蛋白表达情况。将构建的Huh7-LV-shTRPM8细胞通过尾静脉注入BALB/C裸鼠作为实验组（LV-shTRPM8组），以注射Huh7-LV-shNC细胞建立的裸鼠模型为对照组（LV-shNC组），6周后观察裸鼠肺转移情况。结果 TRPM8在HCC组织和细胞系中均高表达（P<0.01）；与shcontrol组比较，沉默TRPM8后 Huh7细胞迁移和侵袭能力减弱（P<0.01），E-cadherin表达增加（P<0.001），vimentin表达降低（P<0.001），TRPM8和Snail蛋白表达水平下调（P<0.001），而AKT和GSK-3β蛋白磷酸化程度降低（P<0.001）。沉默TRPM8使裸鼠肺转移率、肺转移结节数量及肺组织中AFP和TRPM8表达下降（P<0.01）。结论 TRPM8可能通过调控AKT/GSK-3β/Snail信号途径抑制HCC Huh7细胞侵袭和转移。

关键词: 肝细胞癌, TRPM8, AKT/GSK-3β/Snail信号途径, 侵袭, 转移

Abstract: Objective  To investigate the effect of chronic receptor potential 8 (TRPM8) on the invasion and metastasis of hepatocellular carcinoma(HCC) cells and its related regulation mechanism. Methods A total of 20 pairs of matched HCC tissues and their adjacent tissues in our hospital from March 2017 to December 2019 were collected, and the expression of TRPM8 in HCC tissues was detected by immunohistochemistry and qRT-PCR. HCC Huh7 cells were transfected with shTRPM8(shTRPM8 group), and set up a negative control group (shcontrol group). The ability of cell migration and invasion were detected by scratch healing assay and Transwell chamber assay, respectively; the expression of E-cadherin and vimentin were detected by immunofluo-rescence experiment; the expression of AFP, TRPM8 and AKT/GSK-3β/Snail signal pathway related proteins were detected by Western blot. The constructed Huh7-LV-shTRPM8 HCC cells were injected into BALB/C nude mice through the tail vein as the experimental group(LV-shTRPM8 group), while the nude mouse model established by the injection of Huh7-LV-shNC cells were used as the control group(LV-shNC group). The lung metastasis of nude mice was observed after injection for 6 weeks. Results TRPM8 was highly expressed in HCC tissues and cell lines(P<0.01). Compared with the shcontrol group, the migration and invasion of Huh7 cells after TRPM8 silence were decreased (P<0.01);the expression of E-cadherin increased(P<0.001), while the expression of vimentin decreased (P<0.001); the expression of TRPM8 and Snail proteins were down-regulated(P<0.001), and the phosphorylation degree of AKT and GSK-3β proteins were decreased (P<0.001). TRPM8 silencing decreased the lung metastasis rate, the number of lung metastatic nodules and the expression of AFP and TRPM8 in lung tissues in nude mice (P<0.01). Conclusion TRPM8 can inhibit the invasion and metastasis of HCC Huh7 cells by regulating the AKT/GSK-3β/Snail signal pathway.

Key words: Hepatocellular carcinoma, TRPM8, AKT/GSK-3β/Snail signaling pathway, Invasion, Metastasis

中图分类号: 

  • R735.7