Genetic and Transcriptional Control of Spleen Development

脾脏发育的遗传和转录控制

• 批准号: 8278674
• 负责人: Licia Selleri
• 金额: $ 33.66万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-15 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8278674
• 关键词: Architecture; Autopsy; Bacterial Infections; Biological Assay; Cell Culture Techniques; Cell Cycle Regulation; Cell Proliferation; Cell Therapy; Cells; Child; Complex; Comprehension; Defect; Development; Diagnosis; Disease; Embryo; Endothelial Cells; Endothelium; Gene Targeting; Generations; Genetic; Genetically Engineered Mouse; Growth; Health; Hematopoiesis; Hematopoietic; Homeobox Genes; Human; Immune response; Immunity; Intervention; Invaded; Knowledge; Lateral; Life; Light; Liver; Mammals; Mesenchymal; Mesenchymal Stem Cells; Mesenchyme; Mesoderm; Modeling; Molecular; Molecular Diagnosis; Molecular Genetics; Morphogenesis; Mouse Strains; Mus; Newborn Infant; Organ; Organogenesis; Pancreas; Pathogenesis; Pathway interactions; Population; Positioning Attribute; Primordium; Process; Proteins; Public Health; Relative (related person); Reporting; Research; Role; Sepsis; Spleen; Spleen Development; Staging; Stem cells; System; Testing; Tissues; Transcriptional Regulation; Transgenic Mice; Transgenic Organisms; Work; base; blood filter; cell type; established cell line; high risk; immortalized cell; mouse model; neglect; novel; prenatal; programs; reconstitution; repaired; research study

DESCRIPTION (provided by applicant): Organogenesis begins with the specification, positioning and assembly of the cell types specific to an organ into the organ primordium (anlage). Active cell proliferation also takes place to build a critical mass for organ morphogenesis and expansion to occur. In this proposal, we will use the vertebrate spleen as a model to investigate these fundamental steps. The complex architecture and functions of the spleen result from intimate interactions among different cell types: mesenchymal cells ("basic parenchyma"), invading endothelial cells and colonizing hematopoietic cells. In humans, the spleen has critical roles in early hematopoiesis, immunity and blood filtering and its absence (as in congenital asplenia, an under-diagnosed disorder often recognized only at autopsy) results in a high risk for life-threatening bacterial infections in newborns and children. Our long-term objective is to identify genetic pathways that control the successive stages of spleen development: i.e. morphogenesis, expansion, and influx of hematopoietic and endothelial cells, since these interrelated organogenetic processes are of utmost importance to spleen function and yet mostly unknown. Using genetic approaches and asplenic mouse strains, we defined key steps in the genetic pathways that govern early spleen development. We reported that the homeobox gene Pbx1 is required for spleen cell fates and is a hierarchical co-regulator of Nkx2.5 and Hox11 (which are also essential for spleen formation). We also found that Pbx1 expression commences earlier than that of both Nkx2.5 and Hox11 in the Lateral Plate Mesoderm (LPM). Additionally, we uncovered that Pbx1 is expressed in the endothelium of the developing spleen anlage. In view of these findings, our hypothesis is that a distinct sub-population of Pbx1-positive progenitor cells within the LPM is required for spleen parenchyma specification, morphogenesis, and expansion and that Pbx expression in the endothelium also contributes to its function in spleen morphogenesis and expansion. In addition, we hypothesize that both an intact mesenchymal anlage and endothelium are essential for normal spleen hematopoietic colonization and function. Using available lines of gene-targeted and transgenic mice, we will test our hypothesis through embryologic, genetic, and molecular approaches. First, we will establish genetic and molecular pathways that control spleen morphogenesis and expansion. To this end, we will characterize the spleen morphogenesis and cellular proliferation defects in a mouse line with conditional inactivation of Pbx1 in the spleen mesenchymal parenchyma, but not in the endothelium. We will further utilize immortalized cell cultures generated from these embryonic spleens to determine the roles of Pbx in cell cycle regulation. Second, we will assess whether an intact endothelium is essential for spleen morphogenesis and expansion by characterizing a mouse line in which only the endothelium is altered by genetic inactivation of Pbx1. Also, by Pbx1 inducible inactivation, we will establish Pbx temporal requirements in the spleen endothelium. Third, we will genetically dissect the role of the mesenchyme and endothelium, respectively, in spleen hematopoietic colonization, development, and function. Our studies will shed light on novel genetic and molecular networks that underlie the development of the spleen, a neglected organ in regard to its ontogeny. In light of the intimate interactions among the mesenchymal spleen anlage, invading endothelial cells and hematopoietic cells, the new knowledge generated from this work will have a deep impact on the understanding of spleen function. Lastly, our studies aspire to provide a better comprehension of the pathogenesis of congenital asplenia, as we put forth the prerequisite basic genetic background towards prenatal molecular diagnosis of this condition. PUBLIC HEALTH RELEVANCE: The complex architecture and functions of the spleen result from intimate interactions among different cell types: mesenchymal cells ("basic parenchyma"), invading endothelial cells and colonizing hematopoietic cells. In humans, the spleen has critical roles in early hematopoiesis, immunity and blood filtering and its absence (as in congenital asplenia, an under-diagnosed disorder often recognized only at autopsy) results in a high risk for life-threatening bacterial infections and fatal sepsis in newborns and children. Our long-term objective is to identify genetic pathways that control the successive stages of spleen development: morphogenesis, expansion, and influx of endothelial and hematopoietic cells, since these interrelated organogenetic processes are of utmost importance to spleen function and, as of yet, mostly unknown. We anticipate that the new knowledge generated from these studies will have a deep impact on the understanding of spleen function, including hematopoiesis and immune response. In summary, the proposed work will shed light on novel genetic and molecular networks that underlie the ontogeny of the spleen, a neglected organ in regard to its development. Lastly, our studies aspire to provide a better comprehension of the pathogenesis of congenital asplenia, as we put forth the prerequisite basic genetic background towards prenatal molecular diagnosis of this condition, which results in heavily impaired immune response to deadly bacterial infections.

描述（由申请人提供）：器官发生始于特定于器官的细胞类型的规范，定位和组装到器官原始中（Anlage）。还发生了活跃的细胞增殖，以建立一个临界质量，以实现器官形态发生和扩张。在此提案中，我们将使用脊椎动物脾脏作为模型来研究这些基本步骤。脾脏的复杂结构和功能是由于不同细胞类型之间的紧密相互作用而引起的：间充质细胞（“基本实质”），入侵的内皮细胞和定居造血细胞。在人类中，脾脏在早期造血，免疫力和血液过滤及其缺失（如先天性的阿斯普氏症中，这种诊断不足的疾病通常仅在尸检时识别）会导致对新生儿和儿童威胁生命的细菌感染的高风险。我们的长期目标是确定控制脾发育连续阶段的遗传途径：即造血和内皮细胞的形态发生，扩张和涌入，因为这些相互关联的有机遗传学过程对脾脏功能至关重要，但却大多是未知。我们使用遗传方法和无酶小鼠菌株，定义了控制早期脾发育的遗传途径中的关键步骤。我们报道说，同源基因PBX1是脾细胞命运所必需的，是NKX2.5和HOX11的分层共同调节剂（对于脾脏形成也是必不可少的）。我们还发现，PBX1的表达比侧板中胚层（LPM）中NKX2.5和HOX11的表达更早。此外，我们发现PBX1在发育中的脾脏内皮中表达。鉴于这些发现，我们的假设是，LPM内的PBX1阳性祖细胞的独特亚群是脾脏实质的规范，形态发生和扩张所必需的，并且在内皮中PBX表达也有助于其在脾形成和扩张中的功能。此外，我们假设完整的间充质和内皮对于正常的脾脏造血定植和功能至关重要。使用靶向基因和转基因小鼠的可用线，我们将通过胚胎学，遗传和分子方法检验我们的假设。首先，我们将建立控制脾形态发生和扩张的遗传和分子途径。为此，我们将表征小鼠线中脾形态发生和细胞增殖缺陷，在脾间质实质中有条件pBx1的条件失活，但在内皮中却没有。我们将进一步利用这些胚胎脾产生的永生细胞培养物来确定PBX在细胞周期调节中的作用。其次，我们将通过表征只有PBX1遗传失活而改变的小鼠线来评估完整的内皮对于脾形态发生和扩张是否至关重要。同样，通过PBX1诱导的失活，我们将在脾内皮中建立PBX时间要求。第三，我们将分别剖析间质和内皮的作用，分别在脾脏造血定植，发育和功能中。我们的研究将揭示出新的遗传和分子网络，这些网络是脾发育的发展，这是其本体发育的被忽视器官。鉴于间充质脾脏，入侵的内皮细胞和造血细胞之间的紧密相互作用，这项工作产生的新知识将对对脾脏功能的理解产生深远的影响。最后，我们的研究渴望更好地理解先天性阿斯普氏症的发病机理，因为我们提出了对这种疾病产前分子诊断的先决条件基本遗传背景。 公共卫生相关性：脾脏的复杂结构和功能是由于不同细胞类型之间的亲密相互作用而引起的：间充质细胞（“基本实质”），入侵的内皮细胞和造血细胞定居。在人类中，脾脏在早期造血，免疫力和血液过滤及其缺失（如先天性的阿斯普氏症中，这种诊断不足的疾病通常仅在尸检时识别出来）会导致对新生儿和儿童造成生命威胁性细菌感染的高风险和致命的衰退。我们的长期目标是确定控制脾发育连续阶段的遗传途径：内皮和造血细胞的形态发生，扩张和涌入，因为这些相互关联的有机遗传学过程对于脾脏功能至关重要，而且尚未清楚。我们预计，这些研究产生的新知识将对对脾脏功能的理解产生深远的影响，包括造血和免疫反应。总而言之，拟议的工作将揭示出新的遗传和分子网络，这些网络是脾脏的个体发育，这是其发育中被忽视的器官。最后，我们的研究渴望更好地理解先天性阿斯普氏症的发病机理，因为我们提出了对这种疾病产前分子诊断的先决条件基本遗传背景，从而导致对致命细菌感染的免疫力受损。