Embryonic origin and self-renewal of B-1a cells

 描述（由申请人提供）：B-1细胞是鼠B细胞，其易于与常规B细胞（B-2细胞）区分，优先定位于腹膜腔，是先天免疫的一部分，以不依赖于T细胞的方式分泌天然抗体，并且在抵抗细菌或病毒感染、脓毒症和动脉粥样硬化的第一道防线中发挥重要作用。尽管B-1细胞在宿主防御和体内平衡中的重要性，但其发育起源尚不清楚。存在于腹膜腔中的CD 5 +B-1a细胞可以通过移植胎肝（FL）祖细胞而重建，但不能通过成人骨髓（BM）造血干细胞（HSC）重建。因此，B-1a细胞被认为是胎儿起源的，并且在成年生活中维持B-1a细胞而不被成年HSC补充的机制尚未阐明。该项目的总体目标是确认HSC非依赖性胚胎B-1祖细胞对出生后B-1谱系的贡献，并确定在整个成人生活中维持B-1细胞的机制。 我们报道了B-1细胞起源于小鼠胚胎中HSC出现之前卵黄囊（YS）中的生血内皮细胞（HEC）。我们还报道了B-1祖细胞存在于HSC缺陷小鼠模型中。因此，我们假设YS衍生的（HSC非依赖性）B-1a细胞在整个围产期和成年期维持。为了证明这一假设，我们将利用谱系追踪小鼠模型，其中可以通过他莫昔芬诱导在HSC出现之前的特定胚胎时间点标记HEC及其后代。我们预期能够在目标1中确定胚胎来源的细胞对成人B-1细胞池的贡献。成熟的B-1a细胞具有自我补充能力，可通过二次移植检测到。维持B-1a细胞这种自我补充活性的分子机制尚不清楚。Bmi 1是一种多梳组蛋白，对HSC的自我更新至关重要。我们报道了Bmi 1-/- B-1a细胞的自我补充能力受损，提出了Bmi 1在B-1a细胞自我更新中起重要作用的假设。我们将阐明Bmi 1调节B-1细胞自我更新的分子机制，确定一个新的候选Bmi 1靶基因的目标2。最后，本研究的目的三是建立小鼠ES细胞培养体系，以获得具有移植功能的小鼠B-1细胞。我们将比较ES衍生的B-1祖细胞功能与YS/FL衍生的B-1祖细胞功能。我们的研究结果将为B-1细胞的发育和自我更新能力提供新的见解，并将提供一种新的方法，从小鼠ES细胞产生B-1细胞，作为未来潜在的细胞治疗的第一步。

 DESCRIPTION (provided by applicant): B-1 cells are murine B cells that are readily distinguishable from conventional B cells (B-2 cells), that preferentially localize in the peritonel cavity, are part of innate immunity, secrete natural antibodies in a T cell independent manner, and play important roles in the first line of defense against bacterial or viral infections, sepsis and atherosclerosis. Despite the importance of B-1 cells in host defense and homeostasis, their developmental origin is not clear. CD5+B-1a cells residing in the peritoneal cavity can be reconstituted by transplantation of fetal liver (FL) progenitors, but not by adult bone marrow (BM) hematopoietic stem cells (HSCs). Thus B-1a cells are considered to be of fetal origin and the mechanisms that maintain B-1a cells in adult life without being replenished by adult HSCs have yet to be elucidated. The overall goal of the project is to confirm the contribution of HSC-independent embryonic B-1 progenitor cells into postnatal B-1 lineages and define the mechanisms that maintain B-1 cells throughout adult life. We have reported that B-1 cells originate from hemogenic endothelial cells (HECs) in the yolk sac (YS) prior to HSC emergence in the mouse embryo. We have also reported that B-1 progenitor cells are present in an HSC-deficient mouse model. Thus, we hypothesize that YS-derived (HSC independent) B-1a cells are maintained throughout perinatal and adult life. In order to prove this hypothesis, we will utilize a lineage tracing mouse model where one can mark HECs and their progeny at specific embryonic time point before HSC emergence by tamoxifen induction. We anticipate the ability to define the contribution of the embryonic-derived cells to adult B-1 cell pools in Aim 1. Mature B-1a cells display self-replenishing ability that is detectable by secondary transplantation. The molecular mechanisms that sustain this self-replenishing activity in B-1a cells are unknown. Bmi1 is a polycomb group protein that is critical for self-renewal of HSCs. We report that the self- replenishing capacity of Bmi1-/- B-1a cells is impaired, raising the hypothesis that Bmi1 plays an important role in B-1a cell self-renewal. We will elucidate the molecular mechanisms through which Bmi1 regulates B-1 cell self-renewal, identifying a novel candidate Bmi1 target gene in Aim 2. Finally, in Aim 3, we will develop mouse ES culture system to produce functional transplantable mouse B-1 cells. We will compare ES-derived B-1 progenitor cell functions with YS/FL-derived B-1 progenitor cells. Our results will provide new insights into B-1 cell development and self-renewal ability and will provide a novel method to generate B-1 cells from mouse ES cells as a first step of potential cell therapy in the future.