摘要
目的探索在肝癌的发生发展过程中,O-N-乙酰葡萄糖胺(O-GlcNAc)糖基化水平与促血管生成素-2(Ang-2)表达的关系.方法采用二乙基硝胺(DEN)诱导C57BL/6小鼠肝癌模型,检测O-GlcNAc糖基化、Ang-2及内皮细胞黏附分子(CD31)在肝癌中表达;采用N-乙酰氨基葡萄糖转移酶(OGT)抑制剂及N-乙酰氨基葡萄糖苷酶(OGA)抑制剂干预HepG2细胞,检测Ang-2的表达是否随O-GlcNAc糖基化水平的变化而改变;采用MTr检测O-GlcNAc糖基化的水平与肝癌细胞增殖的关系;采用Transwell法检测HUVEC细胞中O-GlcNAc糖基化水平与细胞迁移侵袭能力的关系.结果在DEN诱导的肝癌模型中,小鼠肝脏体积明显增大,肝脏上可见大小不一的肿瘤.免疫组织化学结果显示,与对照组相比,O-GlcNAc糖基化水平与Ang-2的表达在模型组小鼠肝脏中均明显上升(P<0.05),且肝癌组织中CD31染色的微血管密度明显上升(P<0.01);qRT-PCR检测结果显示,通过检测OGT、OGA、Ang-2 mRNA水平,得出O-GlcNAc糖基化水平影响Ang-2的表达水平;Western blotting结果显示,采用OGT抑制剂干预HepG2细胞后,Ang-2的表达随着O-GlcNAc糖基化水平的降低而下调;采用OGA抑制剂干预HepG2细胞后,Ang-2的表达随着O-GlcNAc糖基化水平的上升而升高.MTT结果显示,O-GlcNAc糖基化水平与肝癌细胞的增殖有密切关系,当O-GlcNAc糖基化水平高时,肝癌细胞增殖能力较强,当O-GlcNAc糖基化水平较低时,肝癌细胞的增殖能力明显受到抑制.Transwell结果显示,HUVEC细胞迁移侵袭能力与O-GlcNAc糖基化水平呈正相关.结论在肝癌的发生发展过程中,O-GlcNAc糖基化水平逐渐上升,且Ang-2的表达水平受到O-GlcNAc糖基化的正调控,从而促进肿瘤新生血管的生成.
Objective To explore the relationship between O-linked N-acetylglucosamine(O-GlcNAc)glycosylation and Angiopoietin-2(Ang-2)expression in the development of hepatocellular carcinoma.Methods C57BL/6 mice model of hepatocellular carcinoma was induced by Diethylnitroamine(DEN)to detect the changes of O-GlcNAc glycosylation,Ang-2 and cell adhesion molecule-1(CD31)expression in hepatocellular carcinoma.HepG2 cells were intervened with O-linked Glc NAc-transferase(OGT)inhibitors and O-Glc NAcase(OGA)inhibitors to detect whether the expression of Ang-2 changed with the changes of O-GlcNAc glycosylation level.MTT was used to detect the relationship between O-GlcNAc glycosylation level and proliferation of hepatocellular carcinoma cells.Transwell method was used to detect relationship between the level of O-GlcNAc glycosylation and cell migration and invasion in HUVEC cells.Results In the DEN-induced hepatocarcinoma model,the liver volume of mice increased significantly,and tumors of different sizes could be seen in the liver.The results of immunohistochemistry showed that the glycosylation level of O-GlcNAc and the expression of Ang-2 increased significantly in the liver of the model group compared with the control group(P<0.05).The results of qRT-PCR showed that the level of O-GlcNAc glycosylation affected the expression of Ang-2 by detecting the levels of OGT,OGA and Ang-2.Western blotting showed that the expression of Ang-2 decreased with the decrease of the glycosylation level of O-GlcNAc after the intervention of OGT inhibitors on HepG2 cells.After the intervention of HepG2 cells with OGA inhibitors,the expression of Ang-2 increased with the decrease of the glycosylation level of O-GlcNAc.MTT results showed that the glycosylation level of O-GlcNAc was closely related to the proliferation of hepatocellular carcinoma cells.The proliferation ability of hepatocellular carcinoma cells was stronger when the glycosylation level of O-GlcNAc was higher,.The proliferation ability of hepatocellular carcinoma cells was significantly inhibited,when the glycosylation level of O-GlcNAc was low,.Transwell results showed that HUVEC cell migration and invasion ability was positively correlated with O-GlcNAc glycosylation level.Conclusion During the development of hepatocellular carcinoma,the glycosylation level of O-GlcNAc increases gradually,and the expression level of Ang-2 positively regulated by the glycosylation of O-GlcNAc,thus promoting neovascularization.
作者
王爱红
王明全
杜娟
WANG Aihong;WANG Mingquan;DU Juan(Key Laboratory of Cancer Prevention and Treatment Research,Yan'an University Medical College,Yan'an 716000,China;Department of Interventional Radiology,Yan'an UniversiR AffiliatedHospital,Yan'an 716000,China)
出处
《实验动物与比较医学》
CAS
2020年第1期39-46,共8页
Laboratory Animal and Comparative Medicine
基金
延安市科技局项目(2017KS-19)。
