Characterizing transient hematopoietic multipotent and megakaryocyte progenitor cells during postnatal development

产后发育过程中短暂造血多能和巨核祖细胞的特征

• 批准号: 10537794
• 负责人: Ryan Mack
• 金额: $ 3.33万
• 依托单位: LOYOLA UNIVERSITY CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10537794
• 关键词: 4-Hydroxy-Tamoxifen; Address; Adult; Animals; Aorta; Arteries; B-Lymphocytes; Biological; Biological Assay; Biology; Birth; Blood Cells; Blood Island; Blood Platelets; Bone Marrow; Cell Maturation; Cells; Childhood Leukemia; Clonal Evolution; Data; Development; Disease; Embryo; Embryonic Development; Erythrocytes; Fetal Development; Fetal Liver; Flow Cytometry; Future; Gene Expression Profile; Generations; Gonadal structure; Green Fluorescent Proteins; Hematologic Neoplasms; Hematological Disease; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; In Vitro; Infant; Infant Development; Infant Leukemia; Injections; Label; MPP1 gene; MPP2 gene; Megakaryocytes; Mesonephric structure; Modeling; Molecular; Mouse Protein; Multipotent Stem Cells; Mus; Myelogenous; Myeloproliferative disease; Neonatal; Organism; Palmitoylated Membrane Protein 2; Pathogenesis; Pathologic; Perinatal; Phenotype; Ploidies; Population; Process; Property; Reporter; Spleen; T-Lymphocyte; Techniques; Testing; Tomatoes; Transplantation; Validation; Yolk Sac; base; experimental study; fetal; fetal immunity; granulocyte; hemogenic endothelium; in vivo; insight; leukemia; liver function; monocyte; mouse model; neonatal mice; novel; pediatric patients; postnatal; postnatal development; progenitor; protein expression; reconstitution; self-renewal; stem cells; success; transcriptome sequencing; young adult

Abstract Fetal hematopoiesis has properties distinct from adult hematopoiesis, explaining the unique features of infant myeloproliferative disorders and leukemia. Adult-type hematopoietic stem cells (HSCs) are generated in the major arteries in the aorta-gonad-mesonephros region at embryonic day (E)9.5-E11.5 in mice, colonize the fetal liver (FL) and fetal bone marrow (BM), and remain in adult BM for an organism’s lifetime. Previous studies hypothesized that the fetal-to-adult hematopoietic transition is a gradual maturation process of adult type HSCs. However, two fetal-type hematopoietic progenitors (HPCs) originate from the yolk sac hematopoiesis (E7.5 and E8.5) that also colonize the FL and fetal BM, overlap with maturing HSCs, and are the primary contributors to hematopoiesis during fetal development. Increased evidence suggests this transition is a combined process of declining fetal-type HPCs and maturation of adult-type HSCs. However, the cellular and molecular mechanisms regulating the hematopoietic transition are still largely unknown. Green fluorescent protein (GFP) expression in Ctnnal1-GFP reporter mice was shown to specifically label HSCs in young adults, but not downstream multipotent progenitors (MPPs) or megakaryocyte progenitors (MkPs). In addition to HSCs in neonatal mice, I found GFP labels a subset of MPP1, MPP2 and MkPs. These GFP+ HPCs gradually decline until absent by week 4 post-birth, while GFP- MPP1, MPP2, and MkPs remain. As adult HSC maturation occurs during this period, GFP+ subsets represent novel developmentally restricted HPCs with unknown origin, function, and regulatory mechanisms. In vitro study demonstrates that GFP+ MPP1, MPP2 and MkPs are distinct from their GFP- counterparts and display many properties of fetal type HPCs. For example, although both GFP+ and GFP- MPP2s differentiate to monocytes, granulocytes and megakaryocytes (Mk), GFP+ MPP2s are biased to monocyte and Mk differentiation like fetal HPCs, while GFP- MPP2s are biased to granulocytic differentiation. Compared to Mks produced by GFP- MkPs, GFP+ MkPs produce smaller, lower ploidy Mks similar to Mks found in fetal BM. I hypothesize that Ctnnal1-GFP+ MPP1, MPP2, and MkPs are phenotypically and functionally distinct from their Ctnnal1-GFP- counterparts, are derived from fetal type HPCs independent of adult HSCs, and represent novel transient populations that are capable of initiating infant hematological diseases. I intend to address this hypothesis by (1) fully characterizing the biology and function of GFP+ MPP1, MPP2, and MkPs; and (2) determining where they arise and if they are derived from fetal type HPCs independent of adult HSCs. To do so, I will use three mouse models: Ctnna1-GFP mice for in vitro and flow cytometry experiments, Ctnna1-GFP/Rosa26-tdTomato mice for transplantations, and Cdh5Cre-Ert/Ctnnal1- GFP/Rosa26-tdTomato mice to determine the origin of GFP+ cells. Success of this study will not only help to understand the mechanism of fetal to adult hematopoietic transition, but also serve as a model for hypothesis generation related to the development of infant/pediatric leukemia.

摘要 胎儿造血具有不同于成人造血的特性，这解释了胎儿造血的独特特征。 婴儿骨髓增生性疾病和白血病。成人型造血干细胞（HSC）是在 在胚胎9.5-E11.5天的小鼠中，腹主动脉-性腺-中肾区域的主要动脉， 胎儿肝脏（FL）和胎儿骨髓（BM），并在生物体的一生中保留在成人BM中。以前的研究 假设胎儿向成人造血转化是成人型HSC的逐渐成熟过程。 然而，两种胎儿型造血祖细胞（HPC）起源于卵黄囊造血（E7.5和E7.5）。 E8.5），其也定殖于FL和胎儿BM，与成熟的HSC重叠，并且是造血干细胞的主要贡献者。 胚胎发育过程中的造血。越来越多的证据表明，这种转变是一个综合过程， 下降的胎儿型HPC和成熟的成人型HSC。然而，细胞和分子机制 调节造血转变的机制在很大程度上仍是未知的。 Ctnnal 1-GFP报告小鼠中的绿色荧光蛋白（GFP）表达显示特异性标记 年轻成人中的HSC，但不是下游多能祖细胞（MPP）或巨核细胞祖细胞 （MkPs）。除了新生小鼠的HSC外，我还发现GFP标记了MPP 1，MPP 2和MkPs的一个子集。这些 GFP+ HPC逐渐下降，直到出生后第4周不存在，而GFP-MPP 1、MPP 2和MkPs仍然存在。作为 成体HSC成熟发生在此期间，GFP+亚群代表新的发育受限的HPC 起源、功能和调节机制未知。体外研究表明，GFP+ MPP 1、MPP 2 和MkPs与它们的GFP-对应物不同，并显示出胎儿型HPC的许多特性。比如说， 虽然GFP+和GFP-MPP 2都能分化成单核细胞、粒细胞和巨核细胞（Mk），但GFP + MPP 2和GFP-MPP 2都能分化成单核细胞、粒细胞和巨核细胞（Mk）。 MPP 2像胎儿HPC一样偏向于单核细胞和Mk分化，而GFP-MPP 2偏向于单核细胞和Mk分化。 粒细胞分化与由GFP-MkPs产生的Mk相比，GFP+ MkPs产生更小、更低的Mk。 倍性Mks与胎儿BM中发现的Mks相似。我假设Ctnnal 1-GFP+ MPP 1、MPP 2和MkPs是 在表型和功能上不同于它们的Ctnnal 1-GFP-对应物，衍生自胎儿型HPC 独立于成年HSC，代表了能够启动婴儿HSC的新的瞬时群体， 血液病我打算通过（1）充分表征生物学和功能来解决这个假设， GFP+ MPP 1、MPP 2和MkPs;和（2）确定它们在哪里产生以及它们是否来源于胎儿型 HPC独立于成体HSC。为此，我将使用三种小鼠模型：Ctnna 1-GFP小鼠用于体外实验， 流式细胞术实验，Ctnna 1-GFP/Rosa 26-tdTomato小鼠移植，和Cdh 5Cre-Ert/Ctnna 1- GFP/Rosa 26-tdTomato小鼠以确定GFP+细胞的来源。这项研究的成功不仅有助于 了解胎儿向成人造血过渡的机制，也可作为假说的模型 与婴儿/儿童白血病发展相关的一代。