C57BL/6N-Tg(1.28HBV)/Vst乙肝病毒转基因小鼠肝脏炎症与纤维化的病理特点

肝脏 ›› 2018, Vol. 23 ›› Issue (1): 26-30.

C57BL/6N-Tg(1.28HBV)/Vst乙肝病毒转基因小鼠肝脏炎症与纤维化的病理特点

孙鑫, 黄恺, 赵志敏, 吕靖, 彭渊, 陶艳艳, 刘成海

201203 上海中医药大学附属曙光医院肝病研究所上海市中医临床重点实验室

收稿日期:2017-10-03 发布日期:2020-04-30
通讯作者: 刘成海,Email:chenghailiu@hotmail.com
基金资助:
上海中医药大学研究生创新培育项目(Y201826);国家自然科学基金资助项目(81473404,81603467,81730109);上海市三年行动计划建设项目(ZY3-CCCX-2-1003)

Pathological features of liver inflammation and fibrosis in hepatitis B virus transgenic mice C57BL/6N-Tg(1.28HBV)/Vst

SUN Xin, HUANG Kai, ZHAO Zhi-min, LV Jing, PENG Yuan, TAO Yan-yan, LIU Cheng-hai

Institute of Liver Diseases, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China;Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China;E-institute of TCM Internal Medicine, Shanghai Municipal Education Commission, Shanghai 201203, China

Received:2017-10-03 Published:2020-04-30
Contact: LIU Cheng-hai,Email:chenghailiu@hotmail.com

摘要/Abstract

摘要： 目的观察不同周龄C57BL/6N-Tg(1.28HBV)/Vst乙肝病毒转基因(HBV-Tg)小鼠肝脏炎症与纤维化病理特点。方法采集12、24、36周龄HBV-Tg小鼠血清和肝组织样本,并以36周龄野生型C57BL/6小鼠为对照。采用ELISA检测血清HBsAg、HBeAg水平,荧光定量PCR检测血清HBV DNA,试剂盒检测血清ALT、AST和碱性磷酸酶(ALP)水平,盐酸水解法检测肝组织羟脯氨酸(Hyp)含量;苏木精-伊红(HE)染色、天狼星红染色与网状纤维染色以观察肝组织病理及胶原沉积情况,免疫组化染色观察肝组织HBsAg、HBcAg、α-平滑肌肌动蛋白(α-SMA)表达。结果各周龄HBV-Tg小鼠血清HBsAg、HBeAg及肝组织HBsAg与HBcAg表达均呈阳性,血清HBV DNA载量均大于(1.0×10⁷) IU/mL;血清ALT、AST水平及肝组织Hyp含量、炎症、胶原面积、α-SMA表达均随HBV-Tg小鼠周龄逐渐升高,12周龄时ALP活性最高,36周龄时肝组织炎症损伤及胶原沉积最为明显。结论随着HBV-Tg小鼠周龄增加,肝脏逐渐呈现自发炎症和纤维化,可作为慢性乙型肝炎和乙型肝炎肝纤维化动物模型,为探究其病理机制和药物研发提供基础。

关键词: 乙型肝炎病毒, 转基因动物, 炎症, 肝纤维化, 病理组织学

Abstract: Objective To investigate the pathological features of liver inflammation and fibrosis in hepatitis B virus transgenic (HBV-Tg) mice C57BL/6N-Tg(1.28HBV)/Vst at different ages. Methods HBV-Tg mice at different ages (12, 24 and 36-week old) and wild-type C57BL/6 mice at week 36 were enrolled. Levels of serum hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) were detected using enzyme-linked immunosorbent assay (ELISA). HBV DNA in serum was detected using reverse transcription polymerase chain reaction (RT-PCR). The in situ expression of HBsAg and HBcAg in liver tissue was detected using immunohistochemical staining. Alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) levels in serum were tested using reagent kit. Hydroxyproline (Hyp) level in liver tissue was measured by hydrochloric acid hydrolysis method. Hematoxylin eosin (HE), sirius red and reticular fiber staining were used to observe morphological changes and collagen deposition in liver tissue, and α-SMA immunohistochemical staining was used to observe activation of hepatic stellate cells. Results HBsAg and HBeAg were positive in serum and liver tissues, and serum HBV DNA was more than 1.0×10⁷ IU/mL in all HBV-Tg mice at different ages. Serum ALT and AST levels gradually increased as age increasing. Compared with that in wild type, ALP level in HBV-Tg mice at different weeks was significantly higher and peaked at week 12. Besides, hydroxyproline content, collagen deposition and activation of hepatic stellate cells in liver tissue of HBV-Tg mice were gradually increased. Liver tissue inflammation and fibrosis aggravated gradually increased, and peaked at week 36. Conclusion The liver performed spontaneous inflammation and fibrosis as age increasing in HBV-Tg mice. HBV-Tg mice might be more suitable for the establishment of chronic HBV persistent infection model and liver fibrosis model after HBV infection. It provides technical supports for further mechanism research, as well as development of novel anti-fibrosis and HBV drug.

Key words: Hepatitis B virus, Transgenic animal, Inflammation, Hepatic fibrosis, Histopathology

孙鑫, 黄恺, 赵志敏, 吕靖, 彭渊, 陶艳艳, 刘成海. C57BL/6N-Tg(1.28HBV)/Vst乙肝病毒转基因小鼠肝脏炎症与纤维化的病理特点[J]. 肝脏, 2018, 23(1): 26-30.

SUN Xin, HUANG Kai, ZHAO Zhi-min, LV Jing, PENG Yuan, TAO Yan-yan, LIU Cheng-hai. Pathological features of liver inflammation and fibrosis in hepatitis B virus transgenic mice C57BL/6N-Tg(1.28HBV)/Vst[J]. Chinese Hepatolgy, 2018, 23(1): 26-30.

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