|本期目录/Table of Contents|

[1]刘璐洁,杨 悦,余双庆,等.C 基因型多重耐药乙型肝炎病毒稳定复制表达小鼠模型的建立[J].传染病信息,2018,06:532-536.
 LIU Lu-jie,YANG Yue,YU Shuang-qing,et al.Establishment of a mouse model stably replicating and expressing genotype C multidrug resistant hepatitis B virus[J].Infectious Disease Information,2018,06:532-536.
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C 基因型多重耐药乙型肝炎病毒稳定复制表达小鼠模型的建立(PDF)

《传染病信息》[ISSN:1007-8134/CN:11-3886/R]

期数:
2018年06期
页码:
532-536
栏目:
论 著
出版日期:
2018-12-30

文章信息/Info

Title:
Establishment of a mouse model stably replicating and expressing genotype C multidrug resistant hepatitis B virus
作者:
刘璐洁杨 悦余双庆许智慧赵 丽刘新光吴小兵徐东平董小岩刘 妍
523808 东莞,广东医科大学广东省医学分子诊断重 点实验室(刘璐洁、刘新光);100039 北京,中国人民解放军总 医院第五医学中心临床研究管理中心(刘璐洁、杨悦、许智慧、赵丽、 徐东平、刘妍);100176 北京,五加和分子医学研究所有限公司(余 双庆、吴小兵、董小岩)
Author(s):
LIU Lu-jie YANG Yue YU Shuang-qing XU Zhi-hui ZHAO Li LIU Xin-guang WU Xiao-bing XU Dong-ping DONG Xiao-yan* LIU Yan*
Guangdong Provincial Key Laboratory of Medical Molecular Diagnostics, Guangdong Medical University, Dongguan 523808, China Research Center for Clinical Medicine, the Fifth Medical Center of Chinese PLA General Hospital, Beijing 100039, China
关键词:
多重耐药突变乙型肝炎病毒病毒复制抗原表达小鼠模型
Keywords:
multidrug resistant mutation hepatitis B virus viral replication antigen expression mouse model
分类号:
R373.2
DOI:
10.3969/j.issn.1007-8134.2018.06.009
文献标识码:
A
摘要:
目的 建立高嗜肝性重组8 型腺相关病毒(recombinant adeno-associated virus 8, rAAV8)介导的C 基因型多 重耐药乙型肝炎病毒(multidrug resistant hepatitis B virus, MDR HBV)稳定复制表达小鼠模型。方法 分别构建rAAV8-1.3 倍C 基因型多重耐药型(HBV 逆转录酶区含有rtL180M+S202G+M204V+N236T 位点)和野生型HBV 重组质粒,然后包 装纯化并筛选高滴度重组病毒(rAAV8-1.3HBV-C-MDR 和rAAV8-1.3HBV-C-WT),将病毒经尾静脉注射至6 ~ 8 周龄的 C57BL/6 小鼠,建立C 基因型多重耐药型(实验组,n=6)和野生型(对照组,n=6)HBV 稳定复制表达小鼠模型,监测病 毒复制和抗原表达至9 周并于第9 周末处死小鼠,取肝脏组织进行HE 染色和免疫组化染色,观察肝组织病理改变并分析 HBsAg 和HBcAg 的表达。结果 小鼠注射重组病毒后第2、3、5、7、9 周,血清HBV DNA 表达较稳定,实验组和对照组 小鼠血清HBV DNA 波动范围分别为3.67 ~ 4.06 lg IU/ml 和4.36 ~ 5.11 lg IU/ml。血清HBsAg 和HBeAg 在观察期间均高表达, 第2 周实验组和对照组血清HBsAg 和HBeAg OD 值分别为3.501±0.230 和2.989±0.250,3.967±0.230 和3.384±0.230。2 组小 鼠肝组织未见明显的炎性细胞浸润及组织结构异常,但可检测到HBsAg 和HBcAg 蛋白表达。结论 利用高嗜肝性rAAV8 载体携带1.3 倍C 基因型MDR HBV 基因组体内转导C57BL/6 小鼠,成功建立了稳定复制并持续表达C 基因型MDR HBV 的小鼠模型,为后续评价抗MDR HBV 药物疗效提供实验平台。  
Abstract:
Objective To establish a mouse model stably replicating and expressing genotype C multidrug resistant hepatitis B virus (MDR HBV) mediated by recombinant adeno-associated virus 8 (rAAV8). Methods The recombinant adeno-associated virus plasmids containing 1.3 copies of wild or MDR HBV (HBV reverse transcriptase region included rtL180M+S202G+M204V+N236T site) genotypes C genome were constructed, then the recombinant virus was packaged and purified. High-titer recombinant viruses rAAV8- 1.3HBV-C-MDR and rAAV8-1.3HBV-C-WT were screened and injected via the tail vein into C57BL/6 mice at the age of 6-8 weeks, to establish mouse models model stably replicating and expressing genotype C MDR HBV (experimental group, n=6) and WT HBV (control group, n=6). Viral replication and antigen expression were monitored until 9 weeks after injection, then mice were sacrificed at the end of the ninth weeks. Liver tissue was sampled for hematoxylin-eosin (HE) staining and immunohistochemistry staining. The pathological changes of liver tissue were observed, the HBsAg and HBcAg expression in liver tissue was analyzed. Results At week 2, 3, 5, 7, 9 after recombinant virus injection, serum HBV DNA load maintained stable. The serum HBV DNA load of experimental group and control group fluctuated within 3.67-4.06 lg IU/ml and 4.36-5.11 lg IU/ml, respectively. While, serum HBsAg and HBeAg were both highly expressed during the observation period. The OD values of serum HBsAg and HBeAg in experimental group and control group at week 2 were 3.501±0.230 and 2.989±0.250, 3.967±0.230 and 3.384±0.230, respectively. No obvious infiltration of inflammatory cells or abnormal structure of liver tissue was observed, while HBsAg and HBcAg expression in the liver tissue were detected for both groups. Conclusions By in vivo transduction with recombinant virus rAAV8-1.3HBV-C-MDR, a C57BL/6 mouse model that stably replicated and expressed genotype C MDR HBV is successfully established, providing an experimental platform for further evaluation of anti-MDR HBV drug efficacy.     

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备注/Memo

备注/Memo:
[ 基金项目] 国家自然科学基金面上项目(81371852)
[ 作者单位] 523808 东莞,广东医科大学广东省医学分子诊断重 点实验室(刘璐洁、刘新光);100039 北京,中国人民解放军总 医院第五医学中心临床研究管理中心(刘璐洁、杨悦、许智慧、赵丽、 徐东平、刘妍);100176 北京,五加和分子医学研究所有限公司(余 双庆、吴小兵、董小岩)
[ 通信作者] 董小岩,E-mail: dong-xy@vip.sina.com;刘妍,E-mail: liuyan5360@163.
更新日期/Last Update: 2018-12-30