Optimization of humanized mouse models using mobilized peripheral blood stem cells

项目摘要 本提交文件是对NOT-AI-19-040“管理补充文件 人源化免疫系统小鼠的表征和进一步开发。这 特殊利益机会对我们的母公司R24的结果特别重要 由于动员外周血（mPB）植入的不一致性， 单位，以及缺乏发展的所有组成部分的人类免疫系统， 老鼠我们的R24赠款目标3的目标是使我们的新免疫菌株人源化。 缺陷型HD小鼠，以评估人类免疫系统对 疾病的进程。如果mPB单位失败，我们浪费资金和重要的鼠标。如果 小鼠有一个不完整的人类免疫系统，我们的R24赠款结果的影响 会不那么引人注目。在本补充中，我们将测试两个假设，使用NRG和 NSG.A2小鼠; 1）可获得动员的外周血（mPB）单位的最佳供体。 在动员前通过血液检查主动识别， 用于HIV和其他研究的多批人源化小鼠，和2）共移植 具有人诱导多能干细胞衍生内皮细胞的人干细胞移植物 经工程化以分泌IL-7和IL-15将增强TCR库并拓宽TCR的表达。 人类免疫系统的发展。通过验证HLA类型，CD 47水平 （负责小鼠吞噬人细胞）、选择素、岩藻糖基化水平和 供体mPBSC单位的其他特征，这些特征提供最一致和稳健的 移植水平，我们可以帮助建立最佳的供体标准领域。为 计划的共移植研究，与iPSC衍生的IL-7/IL-15相比， 为了产生内皮细胞，我们将使用从相同细胞扩增的内皮祖细胞， mPB单元用于CD 34+细胞HSC分离。内皮细胞将移植到 新生小鼠幼仔胸腺、粘膜和其他淋巴器官，以允许更好的TCR 从由移植的HSC发育的T细胞祖细胞中选择。持续 IL-7和IL-15的存在将促进整个人淋巴样细胞的发育。我们 目的是显著提高新生儿免疫植入的一致性 缺乏人CD 34+造血干细胞的小鼠幼崽， 外周血单位，增加人TCR发育的范围，以及 扩大了小鼠体内发育的人类细胞的免疫反应。 这一目标与我们自己的工作以及该领域许多其他人的工作高度相关。

Project Abstract This submission is in response to NOT-AI-19-040, “Administrative Supplements for Characterization and Further Development of Humanized Immune System Mice. This special interest opportunity is of particular importance to the outcome of our parent R24 grant, due to the inconsistency of engraftment of mobilized peripheral blood (mPB) units, and the lack of development of all components of the human immune system in the mice. The goal of Aim 3 in our R24 grant is to humanize our new strain of immune deficient HD mice in order to evaluate the role that the human immune system has on the course of the disease. If mPB units fail we waste funding and important mice. If the mice have an incomplete human immune system the impact of our R24 grant outcomes will be less striking. In this supplement we will test two hypotheses, using NRG and NSG.A2 mice; 1) Optimal donors of mobilized peripheral blood (mPB) units can be proactively identified by a blood test prior to mobilization, giving rise to consistent batches of humanized mice for HIV and other studies, and 2) Co-transplantation of the human stem cell graft with human induced pluripotent stem cell-derived endothelial cells engineered to secrete IL-7 and IL-15 will enhance the TCR repertoire and broaden human immune system development. By validating the HLA type, CD47 levels (responsible for mouse phagocytosis of human cells), selectins, fucosylation levels and other characteristics of the donor mPBSC units that give the most consistent and robust engraftment levels, we can help the field by establishing optimal donor criteria. For the planned co-transplantation studies, in comparison to the iPSC-derived IL-7/IL-15 producing endothelial cells, we will use endothelial progenitors expanded from the same mPB unit used for CD34+ cell HSC isolation. The endothelial cells will engraft into the newborn mouse pup thymus, mucosa, and other lymphoid organs to allow better TCR selection from T cell progenitors developing from the transplanted HSCs. The sustained presence of IL-7and IL-15 will enhance overall human lymphoid cell development. Our goal is to significantly improve the consistency of engraftment of newborn immune deficient mouse pups with human CD34+ hematopoietic stem cells from mobilized peripheral blood units, increasing the range of human TCR development, and broadening immune responses by the human cells that have developed in the mice. This goal is highly relevant to our own work and to that of many others in the field.