Development of humanized transgenic mice for HBV/HIV co-infection studies

摘要 更好地理解对B型肝炎病毒的先天性和获得性免疫应答之间的相互作用 (HBV)在人类免疫缺陷病毒（HIV-1）诱导的免疫系统损伤是至关重要的 开发新的抗病毒免疫疗法。携带人类免疫系统的小鼠， HIV-1在研究HBV/HIV共感染的体内实验模型中具有很大的前景。尽管目前 在人源化小鼠模型的进展中，物种差异阻碍了人类的有效“合作”， 免疫系统在一个外壳的小鼠非免疫细胞。我们建议创建一个新的模型， 淋巴组织发育最关键的人类基因组，即β-光敏素受体 (hLTβR），两种参与T细胞和B细胞区建立的趋化因子（hCCL 21，hCXCL 13），以及一种胸腺细胞趋化因子。 基质淋巴细胞生成素（hTSLP）。在本申请中，我们提出用hLTβR、hCCL 21、 hCXCL 13和hTSLP，以开发适合于未来HBV/HIV共感染研究的新模型。我们会委聘 CRISPR基因编辑，我们最近开发的高度稳健的方法，以创建拟议的小鼠 模型。在这种菌株中，我们期望有效地产生人类滤泡辅助细胞，这是一个关键组成部分， 用于建立针对HBV的适应性免疫。研究HBV免疫应答的能力， HIV-1感染的存在将是HBV/HIV共感染模型的新进展。所有的基因操作 在NOG背景下已经具有人IL-34转基因的小鼠品系上进行。合理 对于NOG-hIL 34转基因小鼠，其有效支持人组织驻留巨噬细胞（包括 库普弗细胞），并引入淋巴组织的人的因素，这将提供强有力的支持， 滤泡辅助性T细胞的发育和对HBV抗原的适应性免疫应答。我们将验证 新的人源化毒株控制HIV-1感染的能力以及感染对免疫应答的影响 HBsAg。这样的模型也将提供一种手段，用于开发治疗和解决 HBV/HIV共感染的病理生物学范例以前不可能在体内研究。 我们的近期目标是改进现有的模型，为研究人员创造一个新的，宝贵的工具， 可以支持广泛的HBV/HIV合并感染研究。通过测试人体免疫系统的功效 我们可以确定是否将所有基因型联合收割机组合成一个 小鼠品系。作为开发的下一步，我们将引入TK-NOG背景，以便高效移植 人类肝细胞。我们的长期目标是促进HBV/HIV-1先天免疫和适应性免疫的研究。 合并感染

ABSTRACT A better understanding of the interplay between innate and adaptive immune responses to hepatitis B virus (HBV) in human immunodeficiency virus (HIV-1)-induced immune system impairment is crucial for the development of new antiviral immune therapeutics. Mice that carry human immune system and are affected by HIV-1 have great promise in in vivo experimental models for studying HBV/HIV co-infection. Despite current progress in humanized mouse models, species differences preclude efficient “collaboration” of the human immune system in a shell of mouse non-immune cells. We propose to create a new model that possesses the most crucial set of human genes for lymphoid tissue development, namely the lymphotoxin beta receptor (hLTβR), two chemokines involved in T- and B-cell zone establishment (hCCL21, hCXCL13), and a thymic stromal lymphopoietin (hTSLP). In this application, we propose replacing mouse genes with hLTβR, hCCL21, hCXCL13, and hTSLP to develop a new model suitable for future HBV/HIV coinfection studies. We will employ Easi-CRISPR gene editing, our recently developed and highly robust method, to create the proposed mouse model(s). In this strain, we expect efficient generation of human follicular helper cells, which are a key component for the establishment of adaptive immunity to HBV. The ability to study the immune response to HBV in the presence of HIV-1 infection will be a new step in modeling HBV/HIV coinfection. All genetic manipulations will be performed on a strain of mice that already has human IL-34 transgene on a NOG background. The rational for NOG-hIL34 transgenic mice is that it efficiently supports human tissue-resident macrophages (including Kupffer cells) and introduces lymphoid tissue organizing human factors, which will provide a strong support for development of follicular helper T cells and the adaptive immune response to HBV antigens. We will validate the ability of the new humanized strain to control HIV-1 infection and the effects of infection on immune responses to HBsAg. Such a model will also provide a means for the development of therapeutics and address pathobiological paradigms of HBV/HIV coinfection previously not possible to study in vivo. Our immediate goal is to improve the existing model to create a new, invaluable tool for researchers that can support a wide range of HBV/HIV coinfection studies. By testing the efficacy of human immune system development and function in all four modifications, we can determine whether to combine all genotypes into one mouse strain. As the next step in development, we will introduce TK-NOG background for efficient transplanting of human hepatocytes. Our long-term goal is to facilitate studies of innate and adaptive immunity in HBV/HIV-1 coinfection.