gp73在小鼠肝纤维化肝组织中的表达及机制

基础医学与临床 ›› 2020, Vol. 40 ›› Issue (6): 771-776.

gp73在小鼠肝纤维化肝组织中的表达及机制

马玲玉, 甄一宁, 罗云萍, 段昭君^*

中国医学科学院基础医学研究所北京协和医学院基础学院免疫学系, 北京 100005

收稿日期:2020-03-12 修回日期:2020-04-20 出版日期:2020-06-05 发布日期:2020-05-29
通讯作者: *duanzhaojun@ibms.pumc.edu.cn
基金资助:
国家自然科学基金(81601374)

Expression and mechanism of gp73 in liver tissue of mouse liver fibrosis

MA Ling-yu, ZHEN Yi-ning, LUO Yun-ping, DUAN Zhao-jun^*

Department of Immunology, Institute of Basic Medical Science CAMS, School of Basic Medicine PUMC, Beijing 100005, China

Received:2020-03-12 Revised:2020-04-20 Online:2020-06-05 Published:2020-05-29
Contact: *duanzhaojun@ibms.pumc.edu.cn

摘要/Abstract

摘要： 目的在小鼠肝组织中探讨高尔基体跨膜糖蛋白73(gp73)在肝纤维化进程中的变化及机制。方法腹腔注射CCl₄构建C57BL/6J小鼠肝纤维化模型;用ELISA检测小鼠血清中gp73的水平;用免疫组织化学染色检测肝组织内gp73的表达;用密度梯度离心法分离肝星状细胞(HSC)并检测其中gp73的表达;用免疫组织化学染色法检测肝组织中胰岛素样生长因子2 mRNA结合蛋白3(IGF2BP3)的表达;接下来分别转染IGF2BP3过表达与敲低的质粒,用RT-PCR检测GP73蛋白的编码基因GOLM1的表达变化;流式细胞计量术检测基因敲除鼠肝纤维化模型中HSC内gp73的表达。结果在肝纤维化模型的小鼠血清(P＜0.01)、肝组织(P＜0.05)及原代HSC(P＜0.001)中gp73蛋白表达水平均升高,同时组织中的IGF2BP3蛋白表达也升高(P＜0.05)。细胞系中转入IGF2BP3的过表达及敲低质粒后,成功过表达IGF2BP3(P＜0.001)和敲低(P＜0.01),GOLM1的表达也随之上调(P＜0.001)和下调(P＜0.01)。随后在IGF2BP3敲除鼠的肝纤维化模型的HSC中检测发现gp73表达降低(P＜0.01)。结论在CCl₄诱导的小鼠肝纤维化模型中,由HSC表达的gp73蛋白水平升高,而RNA结合蛋白IGF2BP3是促进其表达的潜在调控因素。

关键词: 肝纤维化, gp73, IGF2BP3

Abstract: Objective To investigate the changes and mechanism of golgi protein 73 (gp73) in mouse with liver fibrosis. Methods The model of hepatic fibrosis was developed by intraperitoneal injection of CCl₄ in C57BL6/J mice. Serum level of gp73 was determined by ELISA. The expression of gp73 in liver tissues was detected by immuno-histochemical staining. Hepatic stellate cell (HSC) was isolated by density gradient centrifugation and gp73 expression was detected. The expression of insulin like growth factor 2 mRNA binding protein 3 (IGF2BP3) in liver tissues was detected by immuno-histochemical staining. Next, IGF2BP3 over-expression and knock down plasmids were transfected, and the changes of GOLM1, which encodes gp73, were detected by RT-PCR. Furthermore, the expression of gp73 in hepatic stellate cells of fibrotic IGF2BP3 knockout mice was detected by flow cytometry. Results A higere expression of gp73 protein increased in serum(P＜0.01), liver (P＜0.05) and primary hepatic stellatecells (P＜0.001) in the fibrotic mice model was found as compared to the control animals. Along with that, the protein level of IGF2BP3 in the liver was also increased (P＜0.05). After over-expressing or knocking down IGF2BP3 by transient transfection, the mRNA expression of GOLM1 was also up-regulated (P＜0.001) and down-regulated (P＜0.01). Respectively, compared with the wide type group, the expression of gp73 was decreased in the hepatic stellate cells of fibrotic IGF2BP3 conditional knockout mice (P＜0.01). Conclusions In the CCl₄ induced liver fibrosis mice, the protein level of gp73 protein in hepatic stellate cells is increased, and RNA binding protein IGF2BP3 is a potential regulatory factor to promote its expression.

Key words: hepatic fibrosis, gp73, IGF2BP3

中图分类号:

R735.7

马玲玉, 甄一宁, 罗云萍, 段昭君. gp73在小鼠肝纤维化肝组织中的表达及机制[J]. 基础医学与临床, 2020, 40(6): 771-776.

MA Ling-yu, ZHEN Yi-ning, LUO Yun-ping, DUAN Zhao-jun. Expression and mechanism of gp73 in liver tissue of mouse liver fibrosis[J]. Basic & Clinical Medicine, 2020, 40(6): 771-776.

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