乙型肝炎病毒表面抗原对细胞脂质合成作用的研究  

Transcriptomics analysis reveals the up regulation of lipid synthesis by HBsAg

作　　者：荆沙 马泽林 赵超[1,2] JING Sha;MA Zelin;ZHAO Chao(Key Laboratory of Medical Molecular Virology(MOE/NHC/CAMS),School of Basic Medical Sciences,Shanghai Medical College,Fudan University,Shanghai 200030,China;National Clinical Research Center on Aging and Medicine,Huashan Hospital,Shanghai 200040,China)

机构地区：[1]复旦大学上海医学院基础医学院教育部、卫健委、医科院医学分子病毒学重点实验室,上海200032 [2]国家老年疾病临床医学研究中心(复旦大学附属华山医院),上海200040

出　　处：《微生物与感染》2020年第6期360-369,共10页Journal of Microbes and Infections

基　　金：国家科技重大专项(艾滋病和病毒性肝炎等重大传染病防治,2018ZX10301208-001-003)。

摘　　要：乙型肝炎病毒(hepatitis B virus,HBV)合成的蛋白调节细胞脂质代谢的研究不断被报道,但乙型肝炎病毒表面抗原(hepatitis B virus surface antigen,HBsAg)与脂质代谢的相互调控研究较少,且机制尚不明确。本研究通过对细胞转录组学的分析,揭示HBsAg对脂质代谢的调控机制。选用稳定表达HBsAg的细胞系HepG2-S-G2与其对照细胞系HepG2-neo-F4进行转录组学分析。利用定量聚合酶链反应(polymerase chain reaction,PCR)、蛋白质印迹法(Western blot,WB)分别检测重要差异基因OXCT1和CYP4F3在mRNA水平和蛋白水平的表达差异。为验证HBsAg促进脂质合成上调的表型,对两种细胞系进行油红O染色并检测细胞脂肪酸、总胆固醇水平。进一步对稳定转染HBV的细胞系HepG2.2.15进行降脂处理,以观察细胞上清液中HBsAg与脂质合成之间是否存在相互调控。结果显示,参与脂质代谢的差异基因发生显著变化,提示HBsAg引起了宿主细胞脂质合成途径的上调和消耗途径下调。定量PCR结果显示,相对于HepG2-neo-F4细胞,HepG2-S-G2细胞的3-酮酸辅酶A转移酶1(3-oxoacid CoA-transferase 1,OXCT1)mRNA水平升高约9倍,与转录组测序结果基本一致;CYP4F3基因在HepG2-S-G2细胞中转录相对下调。WB结果显示,OXCT1和CYP4F3蛋白表达均出现相应的显著上调或下调,并且趋势与转录组分析一致。油红O染色以及细胞脂肪酸、总胆固醇水平检测结果证实HepG2-S-G2细胞中脂滴更明显,且游离脂肪酸和总胆固醇均显著升高。降脂处理结果显示细胞上清液中HBsAg显著降低。上述结果表明,HBsAg可上调脂质代谢、促进脂质合成,提示降脂可能成为抑制HBsAg的潜在有效途径。Although potential association of hepatitis B virus(HBV)gene products and cellular lipid metabolism have been reported,the molecular interaction between hepatitis B surface antigen(HBsAg)and lipid metabolism remains unclear.This study revealed a regulatory mechanism of HBsAg on lipid metabolism through the analysis of cell transcriptome.The cell line HepG2-S-G2 stably expressing HBsAg and the control cell line HepG2-neo-F4 were selected for transcriptome analysis.The results showed that the genes involved in lipid metabolism were significantly changed,suggesting up-regulation of lipid synthesis pathways and down-regulation of lipid degradation pathways in host cells by HBsAg.The expression difference of two key differential genes,OXCT1 and CYP4F3 were confirmed by quantitative PCR for mRNA and western blot for proteins.To verify the phenotype on lipids,the two cell lines were stained with oil red O and the levels of fatty acid and total cholesterol were detected.The result demonstrated that the lipid droplet in HepG2-S-G2 cells was more obvious while the non-esterified fatty acid and total cholesterol were significantly increased.We also found that the extracellular HBsAg was dramatically reduced when stably transfected cells with HBV were treated with lipid-lowering method.These results revealed that HBsAg could up-regulate lipid metabolism and promote lipid synthesis,suggesting that the decrease of lipid may be an effective way to inhibit HBsAg.

关 键 词：乙型肝炎病毒 乙型肝炎病毒表面抗原 脂质代谢 转录组学 

分 类 号：R373.21[医药卫生—病原生物学]