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

 描述(申请人提供)：B-1细胞是小鼠的B细胞，很容易与传统的B细胞(B-2细胞)区分开来，优先定位于腹膜腔，是天然免疫的一部分，以不依赖T细胞的方式分泌天然抗体，并在抵御细菌或病毒感染、败血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细胞在宿主防御和动态平衡方面很重要，但它们的发育起源尚不清楚。CD5+B-1a细胞可通过胎肝(FL)祖细胞移植重建，但不能由成人骨髓(BM)造血干细胞(HSCs)重建。因此，B-1a细胞被认为是胎儿起源的，在成年生活中维持B-1a细胞而不需要成年HSC补充的机制尚未阐明。该项目的总体目标是确认不依赖于HSC的胚胎B-1祖细胞对出生后B-1血统的贡献，并确定在成年后维持B-1细胞的机制。我们已经报道B-1细胞起源于卵黄囊中的血源性内皮细胞(HECS)，在小鼠胚胎中HSC出现之前。我们还报道了在HSC缺陷小鼠模型中存在B-1祖细胞。因此，我们假设YS来源的(不依赖于HSC的)B-1a细胞在整个围产期和成年生活中都保持着。为了证明这一假设，我们将利用一种谱系追踪小鼠模型，在该模型中，人们可以在HSC出现之前的特定胚胎时间点通过他莫昔芬诱导标记HEC及其后代。我们预计在Aim 1中能够确定胚胎来源的细胞对成人B-1细胞库的贡献。成熟的B-1a细胞显示出可通过二次移植检测到的自我补充能力。在B-1a细胞中维持这种自我补充活性的分子机制尚不清楚。BMI1是一种多梳状蛋白，对HSCs的自我更新至关重要。我们报道了Bmi1-/-B-1a细胞的自我补充能力受损，提出了Bmi1在B-1a细胞自我更新中发挥重要作用的假设。我们将阐明Bmi1调控B-1细胞自我更新的分子机制，在目标2中确定一个新的候选Bmi1靶基因。最后，在目标3中，我们将建立小鼠ES培养系统，以产生具有功能的可移植的小鼠B-1细胞。我们将比较ES来源的B-1祖细胞和YS/FL来源的B-1祖细胞的功能。我们的结果将为B-1细胞的发育和自我更新能力提供新的见解，并将为未来潜在的细胞治疗提供一种新的方法来从小鼠ES细胞中分离出B-1细胞。

 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.