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.

抽象的 胎儿造血的特性与成人造血不同，解释了 婴儿骨髓增生性疾病和白血病。成人型造血干细胞（HSC）在 在胚胎日（E）9.5-E11.5的主动脉 - 贡纳德 - 孔孔氏菌区域的主要动脉，在小鼠中殖民 胎儿肝（FL）和胎儿骨髓（BM），并保留在成年BM中，以终生生命周期。先前的研究 假设胎儿到成年的造血过渡是成人型HSC的成熟过程。 但是，两个胎儿造血祖细胞（HPC）来自蛋黄囊造血（E7.5和 e8.5）也将FL和胎儿BM定居，与成熟的HSC重叠，并且是造成的主要贡献者 胎儿发育过程中的造血。越来越多的证据表明，这种过渡是一个合并过程 胎儿型HPC的下降和成人型HSC的成熟。但是，细胞和分子机制 调节造血过渡仍然是未知的。 CTNNAL1-GFP报告基因小鼠中的绿色荧光蛋白（GFP）表达被证明是特异性标记的 年轻人的HSC，但不是下游多元祖细胞（MPP）或巨核细胞祖细胞 （MKP）。除了新生儿小鼠中的HSC外，我还发现GFP标记了MPP1，MPP2和MKP的子集。这些 GFP+ HPC逐渐下降，直到出生后第4周缺席，而GFP-MPP1，MPP2和MKP仍保持不变。作为 成人HSC成熟发生在此期间，GFP+子集代表了开发的新型HPC 具有未知的起源，功能和调节机制。体外研究表明GFP+ MPP1，MPP2 MKP与他们的GFP-对应物不同，并显示了胎儿型HPC的许多特性。例如， 尽管GFP+和GFP-MPP2都与单核细胞，粒细胞和巨核细胞（MK），GFP+分开 MPP2偏向单核细胞和MK分化，例如胎儿HPC，而GFP-MPP2则偏向于 粒细胞分化。与GFP-MKP产生的MK相比，GFP+ MKP产生的较小，较低 类似于胎儿BM中发现的MK的倍性MK。我假设CTNNAL1-GFP+ MPP1，MPP2和MKP是 表型和功能上与其CTNNAL1-GFP-对应物不同，源自胎儿型HPC 独立于成人HSC，代表能够引发婴儿的新型瞬态种群 血液学疾病。我打算通过（1）充分表征这一假设 GFP+ MPP1，MPP2和MKP； （2）确定它们是从胎儿类型派生的位置 HPC独立于成人HSC。为此，我将使用三种鼠标模型：CTNNA1-GFP小鼠进行体外和 流式细胞仪实验，用于移植的CTNNA1-GFP/ROSA26-TDTOMATO小鼠，CDH5CRE-ERT/CTNNAL1- GFP/ROSA26-TDTOMATO小鼠确定GFP+细胞的起源。这项研究的成功不仅会有所帮助 了解胎儿对成人造血过渡的机制，但也是假设的模型 与婴儿/小儿白血病的发展有关的一代。