下调HMGB2表达对肝癌LM3细胞上皮-间质转化的抑制作用及其AKT/mTOR信号通路机制

Inhibitory effect of downregulating HMGB2 expression on epithelial-mesenchymal transition of liver cancer LM3 cells and its AKT/m TOR signaling pathway mechanism

魏雁虹 ¹杨晨雪 ¹杨广民 ¹宋帅 ¹李明 ¹杨海娇 ¹魏海峰²

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作者信息

1. 吉林省人民医院医学检验中心,吉林长春 130021
2. 吉林省人民医院生物治疗与基因诊断重点实验室,吉林长春 130021
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摘要

目的:探讨下调肝癌细胞中高迁移率族框蛋白2(HMGB2)表达对肝癌细胞生物学行为及上皮-间质转化(EMT)进程的影响,并阐明其作用机制.方法:对数生长期的人肝癌LM3细胞分为阴性对照组和HMGB2 RNA干扰组(HMGB2 siRNA组),分别以Lipofectamin 2000为载体转染无关序列的RNA 寡核苷酸(RNA oligo)和敲除HMGB2序列的RNA oligo.采用实时荧光定量PCR(RT-qPCR)法和Western blotting法检测2组细胞中HMGB2 mRNA和蛋白表达水平,分别采用细胞划痕实验和Transwell小室实验检测2组细胞的迁移和侵袭能力,采用Western blotting法检测2组细胞中 E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、波形蛋白(Vimentin)和蛋白激酶 B(AKT)/哺乳动物雷帕霉素靶蛋白(mTOR)通路相关蛋白表达水平.结果:与阴性对照组比较,HMGB2 siRNA组细胞中HMGB2 mRNA和蛋白表达水平均明显降低(P<0.05),HMGB2 siRNA 组细胞划痕愈合率明显降低(P<0.01),侵袭细胞数明显减少(P<0.01),细胞中E-cadherin蛋白表达水平明显升高(P<0.01),N-cadherin、Vimentin、mTOR、AKT和磷酸化AKT(p-AKT)蛋白表达水平明显降低(P<0.05或P<0.01).结论:下调HMGB2的表达可降低肝癌LM3细胞迁移和侵袭能力并抑制EMT,其作用机制可能与参与调节AKT/mTOR通路相关蛋白表达有关.

Abstract

Objective:To discuss the effect of downregulating of high mobility group box protein 2(HMGB2)expression on the biological behavior of the liver cancer cells and the epithelial-mesenchymal transition(EMT)process,and to clarify its mechanism.Methods:The human liver cancer LM3 cells at logarithmic growth phase were divided into negative control group and HMGB2 RNA interference group(HMGB2 siRNA group);the cells in two groups were transfected with RNA oligonucleotides(RNA oligos)with irrelevant sequences and RNA oligos designed to knock down HMGB2,and the Lipofectamine 2000 was regarded as the vector.The expression levels of HMGB2 mRNA and protein in the cells in two groups were detected by real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods;cell scratch assay and Transwell chamber assay were used to detect the migration and invasion abilities of the cells in two groups;the expression levels of E-cadherin,N-cadherin,and Vimentin proteins and protein kinase B(AKT)/mammalian target of rapamycin(mTOR)pathway related proteins in the cells in two groups were detected by Western blotting method.Results:Compared with negative control group,the expression levels of HMGB2 mRNA and protein in the cells in HMGB2 siRNA group were significantly decreased(P<0.05),the cell scratch healing rate was significantly decreased(P<0.01),the number of invasion cells was significantly decreased(P<0.01),and the expression level of E-cadherin protein in the cells was significantly increased(P<0.01),while the expression levels of N-cadherin,Vimentin,mTOR,AKT,and phosphorylated AKT(p-AKT)proteins in the cells were significantly decreased(P<0.05 or P<0.01).Conclusion:Downregulating the expression of HMGB2 can reduce the migration and invasion abilities of the liver cancer LM3 cells and inhibit the EMT,and its mechanism may be related to regulating the expression of the AKT/mTOR pathway related proteins.

关键词

肝肿瘤/高迁移率族框蛋白2/上皮-间质转化/细胞迁移/细胞侵袭/蛋白激酶B/哺乳动物雷帕霉素靶蛋白

Key words

Liver carcinoma/High mobility group box protein 2/Epithelial-mesenchymal transition/Cell migration/Cell invasion/Protein kinase B/mammalian target of rapamycin

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基金项目

吉林省科技厅自然科学基金项目(20200201346JC)

出版年

2024

吉林大学学报(医学版)

吉林大学

吉林大学学报(医学版)

CSTPCDCSCD北大核心

影响因子：0.909

ISSN：1671-587X

参考文献量2

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