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

摘要 更好地了解乙肝病毒先天免疫反应和获得性免疫反应之间的相互作用 在人类免疫缺陷病毒(HIV-1)诱导的免疫系统损伤中， 新的抗病毒免疫疗法的开发。携带人类免疫系统并受 HIV-1在体内研究乙肝病毒/艾滋病混合感染的实验模型中具有很大的前景。尽管目前 人源化小鼠模型的进展，物种差异排除了人类有效的“合作” 免疫系统中的一种小鼠非免疫细胞。我们建议创建一种新的模型，它具有 人类淋巴组织发育最关键的一组基因，即淋巴毒素β受体 (hLTβR)，参与T和B细胞区建立的两个趋化因子(hCCL21，hCXCL13)，以及一个胸腺 基质淋巴生成素(HTSLP)。在这一应用中，我们建议用hLTβR，hCCL21， HCXCL13和hTSLP，以开发一种新的模型，适用于未来的乙肝病毒/艾滋病毒共感染研究。我们将聘用 EASI-CRISPR基因编辑，我们最近开发的高度健壮的方法，以创建建议的小鼠 模特(S)。在这种菌株中，我们希望高效地产生人类卵泡辅助细胞，这是一个关键的组成部分 用于建立对乙肝病毒的适应性免疫。研究儿童对乙肝病毒的免疫反应的能力 HIV-1感染的存在将是建立HBV/HIV混合感染模型的新步骤。所有的基因操作都会 在NOG背景下已经有人IL-34转基因的小鼠身上进行了实验。理性的人 对于NOG-hIL34转基因小鼠来说，它有效地支持了人类组织驻留的巨噬细胞(包括 Kupffer细胞)，并引入淋巴组织组织人类因子，这将为 滤泡辅助性T细胞的发育与对乙肝病毒抗原的适应性免疫反应我们将验证 新型人源化毒株控制HIV-1感染的能力及其对免疫应答的影响 对乙肝表面抗原的影响。这种模式还将提供一种手段来发展治疗学和解决 乙肝病毒/艾滋病病毒混合感染的病理生物学范式以前不可能在体内进行研究。 我们的直接目标是改进现有模型，为研究人员创建一种新的、无价的工具， 可以支持广泛的乙肝病毒/艾滋病毒共感染研究。通过测试人类免疫系统的功效 发育和功能在所有四种修饰中，我们可以确定是否将所有基因组合为一种 老鼠品系。作为下一步的开发，我们将介绍TK-NOG背景，以实现高效移植 人类肝细胞。我们的长期目标是促进对乙肝病毒/HIV-1先天免疫和获得性免疫的研究 混合感染。

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.