Genetic and Transcriptional Control of Spleen Development

脾脏发育的遗传和转录控制

• 批准号: 7986541
• 负责人: Licia Selleri
• 金额: $ 35.07万
• 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/7986541
• 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; Hematopoiesis; Hematopoietic; Homeobox Genes; Human; Immune response; Immunity; Intervention; Invaded; Knowledge; Lateral; Lateral Mesoderm; 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; public health relevance; 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）：器官发生开始于器官特异性细胞类型的描述、定位和组装到器官原基（样本）中。活跃的细胞增殖也会为器官的形态发生和扩张建立一个临界质量。在这个建议中，我们将使用脊椎动物脾脏作为模型来研究这些基本步骤。脾脏复杂的结构和功能是不同类型细胞之间密切相互作用的结果：间充质细胞（“基本实质”）、侵袭的内皮细胞和定植的造血细胞。在人类中，脾脏在早期造血、免疫和血液过滤中起着至关重要的作用，它的缺失（如先天性脾功能不全，一种诊断不清的疾病，通常只有在尸检时才能发现）导致新生儿和儿童发生危及生命的细菌感染的风险很高。我们的长期目标是确定控制脾脏发育连续阶段的遗传途径：即造血细胞和内皮细胞的形态发生、扩张和涌入，因为这些相互关联的器官发生过程对脾脏功能至关重要，但大多未知。利用遗传方法和无脾小鼠品系，我们确定了控制早期脾脏发育的遗传途径的关键步骤。我们报道了同源盒基因Pbx1是脾脏细胞命运所必需的，并且是Nkx2.5和Hox11的分层共同调节剂（它们也是脾脏形成所必需的）。我们还发现Pbx1在侧板中胚层（LPM）中的表达开始时间早于Nkx2.5和Hox11。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中一个独特的pbx1阳性祖细胞亚群是脾脏实质规范、形态发生和扩张所必需的，内皮中Pbx的表达也有助于其在脾脏形态发生和扩张中的功能。此外，我们假设完整的间充质细胞和内皮对于正常的脾脏造血定植和功能是必不可少的。利用现有的基因靶向和转基因小鼠，我们将通过胚胎学、遗传学和分子方法来验证我们的假设。首先，我们将建立控制脾脏形态发生和扩张的遗传和分子途径。为此，我们将描述脾脏形态发生和细胞增殖缺陷的小鼠系，在脾间质实质中，而不是在内皮中，Pbx1条件失活。我们将进一步利用这些胚胎脾脏产生的永生化细胞培养物来确定Pbx在细胞周期调节中的作用。其次，我们将评估完整的内皮对脾脏形态发生和扩张是否必要，通过表征一个小鼠品系，其中只有内皮因Pbx1基因失活而改变。此外，通过诱导Pbx1失活，我们将在脾内皮中建立Pbx的时间需求。第三，我们将从遗传学角度分别剖析间质和内皮在脾脏造血定植、发育和功能中的作用。我们的研究将揭示新的遗传和分子网络，基础上的发展脾脏，一个被忽视的器官，在其个体发生。鉴于脾间充质基质与侵袭性内皮细胞和造血细胞之间的密切相互作用，这项工作产生的新知识将对脾功能的认识产生深远的影响。最后，我们的研究希望提供一个更好的理解先天性脾的发病机制，因为我们提出了先决条件的基本遗传背景产前分子诊断这种情况。