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

Abstract

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

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.

References

[1] 中华医学会肝病学分会, 中华医学会感染病学分会. 慢性乙型肝炎防治指南(2015更新版). 肝脏, 2015, 20: 915-932.
[2] Kosinska AD, Liu J, Lu M, et al. Therapeutic vaccination and immunomodulation in the treatment of chronic hepatitis B: preclinical studies in the woodchuck. Med Microbiol Immunol. 2015, 204: 103-114.
[3] Korba BE, Cote PJ, Menne S, et al. Clevudine therapy with vaccine inhibits progression of chronic hepatitis and delays onset of hepatocellular carcinoma in chronic woodchuck hepatitis virus infection. Antivir Ther, 2004, 9: 937-952.
[4] Jamall I, Finelli V, Que Hee S. A simple method to determine nanogram levels of 4-hydroxyproline in biological tissues. Anal Biochem, 1981, 112: 70-75.
[5] 王宝菊, 朱彬, 郭伟娜, 等. HBV感染与复制模型的建立及应用. 临床肝胆病杂志, 2017, 33: 1458-1464.
[6] Qing Y, Chen M, Zhao J, et al. Construction of an HBV DNA vaccine by fusion of the GM-CSF gene to the HBV-S gene and examination of its immune effects in normal and HBV-transgenic mice. Vaccine, 2010, 28: 4301-4307.
[7] Ding C, Wei H, Sun R, et al. Hepatocytes proteomic alteration and seroproteome analysis of HBV-transgenic mice. Proteomics, 2009, 9: 87-105.
[8] Huang LR, Gabel YA, Graf S, et al. Transfer of HBV genomes using low doses of adenovirus vectors leads to persistent infection in immune competent mince. Gastroenterology, 2012, 142: 1447-1450.
[9] Chisari FV, Pinkert CA, Milich DR, et al. A transgenic mouse model of the chronic hepatitis B surface antigen carrier state. Science, 1985, 230: 1157-1160.
[10] Guidotti LG, Matzke B, Schaller H, et al. High-level hepatitis B virus repliction in transgenic mice. J Virol, 1995, 69: 6185-6189.
[11] Larkin J, Clayton M, Sun B, et al. Hepatitis B virus transgenic mouse model of chronic liver disease. Nat med, 1999, 5: 907-912.
[12] Jin Z, Sun R, Wei H, et al. Accelerated liver fibrosis in hepatitis B virus transgenic mice: involvement of natural killer T cells. Hepatology, 2011, 53: 219-229.
[13] 陈迪, 田枫, 康爱军, 等. C57BL/6J-HBV转基因小鼠随增龄乙肝表面抗原表达及转氨酶水平变化探讨. 四川动物, 2011, 30: 186-188.
[14] 徐军, 訾晓渊, 余宏宇, 等. C57 TgN(HBV adr2.0)SMMU转基因小鼠肝脏的组织病理学观察. 第二军医大学学报, 2005, 26: 283-285.
[15] 骆抗先, 朱幼芙. 乙型肝炎:临床与活体组织病理. 北京: 科学出版社, 2001, 162-179.
[16] Tsuchida T, Friedman SL. Mechanisms of hepatic stellate cell activation. Nat Rev Gastroenterol Hepatol, 2017, 14: 397-411.