The developmental pathway of fetal-derived B cells

摘要 干细胞理论指出，所有的血细胞都来源于造血干细胞（HSC），是造血干细胞的核心。 血液学和免疫学的概念。骨髓移植治疗血液病是植根于 这一概念认为，供体HSC可以取代受体体内的所有血细胞。然而，最近的研究 挑战了干细胞理论，表明一些组织驻留免疫细胞是从内皮细胞发育而来的， 这些免疫细胞可以以HSC非依赖性的方式被胚胎的细胞（EC）所替代，并且这些免疫细胞可能不会被HSC所替代。 此外，据报道，显著百分比的常规B细胞（B-2细胞）也来源于 从胚胎中的胚胎EC祖细胞开始，并持续到老年小鼠。这些惊人的结果突出了一个巨大的 血液学和免疫学领域的知识空白，因为HSC非依赖性B-2细胞的发育 尚未被认可。由于B细胞在保护免受感染和在炎症的病理学中起重要作用， 自身免疫性疾病和其他疾病，了解HSC的发育途径至关重要， 与HSC衍生的对应物相比，独立的B细胞及其功能。然而，这是具有挑战性的， 澄清这些B细胞的起源，因为胚胎中EC有多波造血， 其中HSC和HSC非依赖性血液祖细胞似乎是同时产生的。我们的初步数据 其他研究表明，B细胞发育至少有三个阶段：1）先天免疫B-1细胞 发育，2）先天B-1和常规B-2细胞的共同祖细胞，和3）B-2显性祖细胞 来源于HSC。我们的目标是根据B细胞的起源和来源来确定它们的功能差异， 建立一个修订的B细胞发育图，从胚胎到成人，使用各种谱系的组合， 追踪和体内条形码化小鼠模型。为了实现这些目标，Aim 1将利用HSC标记小鼠（100%）。 实现HSC的标记）和HSC衍生的淋巴样细胞耗竭模型，使得HSC非依赖性和 将分离依赖性B细胞亚群。然后，这些HSC非依赖性和依赖性B细胞亚群将 分选并检查1）抗体和细胞因子分泌，体外刺激后的类别转换，和 2）在体内对S.肺炎感染，和3）对肺炎球菌 预防针在目标2中，初步数据鉴定了早期免疫缺陷病毒中最早的先天免疫限制性B祖细胞。 胎肝此外，新生儿脾脏的scRNA测序显示了三个独立的B祖细胞簇， 支持我们的假设因此，可以标记HSC非依赖性血液的EC谱系示踪小鼠模型 将祖细胞与体内条形码结合，这将使克隆关系可视化 HSC与胎儿和HSC衍生的B祖细胞之间的关系。从这一建议中获得的结果将修订 目前的B细胞发展的范例，并将用于了解病理以前没有认识到 在没有认识到各种胎儿来源的B细胞亚群的存在的情况下。

Abstract Stem cell theory states that all the blood cells are derived from hematopoietic stem cells (HSCs) and is a central concept of hematology and immunology. Bone marrow transplant therapy for blood disorders is rooted in the concept that donor HSCs can replace all blood cells in the recipient's body. However, recent research has challenged the stem cell theory by showing that some tissue-resident immune cells develop from endothelial cells (EC) of the embryo in an HSC-independent manner and these immune cells may not be replaced by HSCs. Furthermore, it has been reported that a significant percentage of conventional B cells (B-2 cells) also originate from fetal EC progenitors in the embryo and persist into old mice. These striking results highlight a huge knowledge gap in the hematology and immunology fields because HSC-independent B-2 cell development has yet to be recognized. Since B cells play important roles in protecting against infections and in the pathology of autoimmune and other diseases, it is critically important to understand the developmental pathways of HSC- independent B-cells and their functions compared to HSC-derived counterparts. However, it is challenging to clarify the origins of these B-cells since there are multiple waves of hematopoiesis from ECs in the embryo, in which HSCs and HSC-independent blood progenitors seem to be produced simultaneously. Our preliminary data and other studies suggest that there are at least three waves of B-cell development; 1) innate-immune B-1 cell development, 2) common progenitors for innate B-1 and conventional B-2 cells, and 3) B-2 dominant progenitors derived from HSCs. Our objective is to determine the functional differences of B-cells based on their origins and to establish a revised B-cell development map from embryo to adult, using combinations of various lineage tracing and in vivo barcoding mouse models. To pursue these goals, Aim 1 will utilize HSC-labeling mice (100% labeling of HSCs is achieved) and the HSC-derived lymphoid cell depletion model so that HSC-independent and dependent B-cell subsets will be separated. Then, these HSC-independent and dependent B-cell subsets will be sorted and examined for 1) antibody and cytokine secretion, class switching upon in vitro stimulation, and their IgM repertoire, 2) in vivo repsonse to S. pneumoniae infection, and 3) response to pneumococcal vaccination. In Aim 2, preliminary data identified the earliest innate-immune restricted B-progenitors in the early fetal liver. Also, scRNA-sequencing of the neonatal spleen revealed three separate B-progenitor clusters, supporting our hypothesis. Thus, the EC-lineage tracing mouse model that can mark HSC-independent blood progenitors will be combined with in vivo barcoding, which will enable visualization of the clonal relationships between HSCs and fetal- and HSC-derived B-progenitors. The results obtained from this proposal will revise the current paradigm of B-cell development and will be utilized to understand pathology not previously recognized in the absence of recognizing the existance of various fetal-derived B cell subsets.