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[免疫抑制剂地塞米松可通过水动力转染法显著延长乙肝病毒小鼠模型中乙肝病毒抗原的表达]

[Immunosuppressant dexamethasone can significantly extend the expression of hepatitis B virus antigens in the HBV mouse model by hydrodynamic transfection method].

作者信息

Guo Yan-Ju, Wang Wen, Sun Shi-Hui, Zeng Dao-Bing, Zhao Guang-Yu, Yu Hong, Guo Yan, Tan Wen-Jie, Lu Shi-Chun, Zhou Yu-Sen

机构信息

State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Science, Beijing 100071, China.

出版信息

Bing Du Xue Bao. 2010 Jan;26(1):20-6.

Abstract

To develop a HBV infection mouse model by hydrodynamic-based transfection and further to optimize the method of development of HBV infection mouse model. We first developed a construct which contained inverted terminal repeat elements (ITR) of adeno-associated virus (AAV) and 1. 3 copies of HBV genome (ayw subtype). The pAAV-HBV1. 3 DNA was then injected hydrodynamically into the tail veins of C57BL/6 mice in 5 seconds. The virus load in serum and liver was assayed by ELISA and Real-time PCR. The expression of virus antigen and the pathologic changes of liver were analyzed by HE and immunohistochemical staining. Meanwhile, to develop HBV transfected immunosuppressied mouse, mice were injected intraperitoneally triple with 0.2 ml dexamethason (50 mg/kg) every two days before HBV transfection. The levels of HBsAg and HBeAg were assayed by ELISA. Our data showed: (1) HBsAg and HBeAg were positive (100%) in serum and liver of experimental normal mouse at day 10 after HBV transfection, and became negative at day 30 and day 60. Meanwhile the viral load in serum and liver in experimental group was significantly higher than that in control group at day 10, 30 and 60 after HBV transfection (P < 0.01, P < 0.05, respectively). (2) HBsAg and HBeAg in serum in immunosuppressed mouse model were positive until 60 days. In conclusion, a HBV infection mouse model was developed successfully by hydrodynamic-based transfection. By suppressing the immune status of mice injected with dexamethasone, the expression of HBV antigens was extended longer than that in normal adult mice. These models pave a way for HBV research and evaluation of HBV vaccine and drug development.

摘要

通过基于流体动力学的转染建立乙型肝炎病毒(HBV)感染小鼠模型,并进一步优化HBV感染小鼠模型的建立方法。我们首先构建了一个包含腺相关病毒(AAV)反向末端重复元件(ITR)和1.3拷贝HBV基因组(ayw亚型)的载体。然后在5秒内通过流体动力学方法将pAAV-HBV1.3 DNA注入C57BL/6小鼠的尾静脉。通过酶联免疫吸附测定(ELISA)和实时聚合酶链反应(Real-time PCR)检测血清和肝脏中的病毒载量。通过苏木精-伊红(HE)染色和免疫组织化学染色分析病毒抗原的表达和肝脏的病理变化。同时,为了建立HBV转染的免疫抑制小鼠模型,在HBV转染前每两天给小鼠腹腔注射0.2 ml地塞米松(50 mg/kg)三次。通过ELISA检测乙肝表面抗原(HBsAg)和乙肝e抗原(HBeAg)的水平。我们的数据显示:(1)在HBV转染后第10天,实验正常小鼠血清和肝脏中的HBsAg和HBeAg均为阳性(100%),在第30天和第60天变为阴性。同时,在HBV转染后第10天、第30天和第60天,实验组血清和肝脏中的病毒载量显著高于对照组(分别为P < 0.01,P < 0.05)。(2)免疫抑制小鼠模型血清中的HBsAg和HBeAg在60天内均为阳性。总之,通过基于流体动力学的转染成功建立了HBV感染小鼠模型。通过用地塞米松抑制小鼠的免疫状态,HBV抗原的表达比正常成年小鼠延长。这些模型为HBV研究以及HBV疫苗和药物开发的评估铺平了道路。

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