复制型HBV-Tg小鼠原代肝细胞的培养.pdf

垡壶国堕匡堂盘查垫!!生!旦垫旦箜堑鲞箜!塑丛!堕堕』墨唑!№!!堑!堕坠；!垒P旦垫!!!!! ·97· ·实验研究· 复制型HBV—Tg小鼠原代肝细胞的培养 陶鑫，易学瑞，袁有成，刘搌扬，张欣蕊，孔祥平 【摘要】目的建立复制型HBV-Tg原代肝细胞体外培养方法，观察HBv-Tg小鼠原代肝细胞的氧化应激敏感 性。方法改良的两步灌注法分离、纯化、培养HBV-Tg小鼠原代肝细胞，24孔板培养7天，每天换液，收集上清，检测 乳酸脱氢酶(1actatedehydrogenase，LDH)、丙氨酸转移酶(alanine trans- 原代肝细胞从铺板第3天开始，LDH、AL，T、√蟠T水平达到平衡，膜稳定性恢复正常；在第3～5天ALB水平稳定于较高 伤，浓度2％时，野生型Balb／c鼠肝细胞损伤程度明显低于HBV-Tg小鼠，浓度≥2％时，两种鼠肝细胞的损伤程度相 当。结论复制型HBV-Tg小鼠原代肝细胞培养可以在体外稳定维持肝细胞活性与功能，表达高水平的HBV-DNA和 HBsAg，为体外研究乙型肝炎病毒发病机制提供新的模型。 【关键词】乙型肝炎病毒；活性氧簇；酒精；细胞模型；原代肝细胞 【中图分类号】R329．2 【文献标识码】A CIIllureofHepat雠—妯嚣inReplicative卸阶，一T童Mice 了、A0Xin，Y了Xue-rui，H舱N Xin-rui，K0』、『G You-cheng，LjUZhen-yang，ZHA^『GXiang—ping．CenterofHe抛斑 Diseases，No．458 510602，China Hospital author：舯NG Corresponding Xiang-ping，E-mail：xiangping_kon艘hotmail．C077q Toestablishamethodin invitroin miceandobserve [AbstractlObjective culturingprimaryhepatocytesHBV-Tg theroleofoxidativestressinthe ofHBV．MethodsMice wereharvested amodified pathogenesis hepatocytes byusing two isolated wereinoculatedintoafreshmedium 24hoursfor stepscollagenasepeffusion．Thehepatocytes changedevery 7 lactate transaminase(AST)andalbumin days．Thedehydrogenase(LDH)，alaninetransaminase(AL，1、)，aspartate of in (ALB)levelsand HgsAg，HBV-DNAthecollected were virologicalindicators supernatants mice wereincubatedin24一well for24hours theisolationof1ivercellsfrom andwild