新型分化培养基增强HepG2肝细胞功能的研究  

Study on the enhancement of hepatocyte function of HepG2 induced by novel differentiation culture medium

作　　者：王振宇 李伟建 张洪丹 景宏舒 袁天杰 彭媛 鄢和新 翟博[1] WANG Zhen-yu;LI Wei-jian;ZHANG Hong-dan;JING Hong-shu;YUAN Tian-jie;PENG Yuan;YAN He-xin;ZHAI Bo(Department of Interntional Oncology,Renji Hospital,Jiaotong University School of Medicine,Shanghai 200127,China)

机构地区：[1]上海交通大学医学院附属仁济医院肿瘤介入科,200127 [2]上海交通大学医学院附属仁济医院麻醉科,200127 [3]上海赛立维生物科技有限公司

出　　处：《肝脏》2019年第7期761-764,共4页Chinese Hepatology

基　　金：国家科技重大专项课题-艾滋病和病毒性肝炎等重大传染病防治(2017ZX10203206-006-002)

摘　　要：目的探讨自行研发的肝细胞成熟培养基(Hepatic Maturation Medium,HMM)是否能够提高肝肿瘤细胞系HepG2的肝细胞功能,并联合3D培养开发一种更适于HepG2的培养体系。方法从细胞库中取得人类肝癌细胞株HepG2。分别用DMEM及HMM进行细胞的2D及3D培养,通过定量PCR、PAS染色、尿素合成等方法进行功能评价,最后在3D模型下比较DMEM及HMM培养的HepG2与氯丙嗪肝毒性的敏感性,综合评价HMM对HepG2的作用。结果HMM可以提高HepG2的糖原合成能力,并使其CYP3A4、NTCP、ALB、CPS1、G6PC等肝细胞功能基因的表达水平分别上调4.7±0.2、1.3±0.1、1.4±0.1、4.9±0.2、5.6±0.1倍。当HMM联合3D培养后,相对于2D培养HepG2,其CYP3A4、NTCP、CPS1、G6PC等肝细胞功能基因的表达水平及尿素合成能力进一步提高6.5±0.6、2.0±0.03、2.3±0.2、77±1.2、479±196倍。此外,HMM显著改善了3D环境下的HepG2对于氯丙嗪毒性的敏感性。结论不论是在2D还是3D条件下,HMM均可以改善HepG2的肝细胞功能,HMM联合3D培养将有利于HepG2为基础的研究。Objective To investigate whether the novel hepatic maturation medium(HMM)can improve the hepatocyte function of the liver tumor cell line of HepG2,and to develop a more suitable culture system for HepG2 in combination with 3-dimensional(3D)culture technique.Methods Human hepatoma cell line HepG2 was obtained from the cell bank.Subsequently,the cells were cultured both in 2-dimensional(2D)and 3D,using Dulbecco's modified Eagle medium(DMEM)or HMM.Functional evaluation was performed by quantitative polymerase chain reaction(qPCR),periodic acid-schiff staining,urea synthesis assay.Finally,the sensitivity of cultured HepG2 to chlorpromazine was detected in the 3D condition combined with DMEM and HMM,respectively.Results HMM could increase the glycogen synthesis ability of HepG2,and upregulate the expression levels of Cytochrome P450 3A4(CYP3A4),Na+-taurocholate cotransporting polypeptide(NTCP),albumin(ALB),Carbamoyl-phosphase synthetase 1(CPS1)and Glucose-6-phosphatase(G6PC),by 4.7±0.18,1.3±0.10,1.4±0.10,4.9±0.20 and 5.6±0.10 times,respectively.More importantly,in HepG2 cultured using HMM in 3D compared with those in 2D,the expression levels of CYP3A4,NTCP,CPS1,G6PC,and the ability of urea synthesis were further increased by 6.5±0.6,2.0±0.03,2.3±0.2,77±1.2 and 479±196 times,respectively.In addition,HMM significantly improved the sensitivity of HepG2 to chlorpromazine toxicity in 3D culture.Conclusion HMM can improve the hepatocyte function of HepG2 in both 2D and 3D conditions.HMM combined with 3D culture will be helpful in HepG2-based researches.

关 键 词：HEPG2 3D 肝细胞成熟培养基 肝细胞 肝毒性 

分 类 号：R73[医药卫生—肿瘤]