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）：器官发生始于将器官特有的细胞类型指定、定位和组装成器官原基（原基）。活跃的细胞增殖也会产生器官形态发生和扩张的临界质量。在本提案中，我们将使用脊椎动物的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不同细胞类型之间密切相互作用的结果：间充质细胞（“基本实质”）、侵入内皮细胞和定植造血细胞。在人类中，脾脏在早期造血、免疫和血液过滤中发挥着关键作用，其缺失（如先天性无脾症，一种诊断不足的疾病，通常只有在尸检时才能识别）会导致新生儿和新生儿出现危及生命的细菌感染的高风险。孩子们。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、扩张和流入，因为这些相互关联的器官发生过程对于脾脏功能至关重要，但大多是未知的。使用遗传方法和无脾小鼠品系，我们定义了控制早期脾脏发育的遗传途径中的关键步骤。我们报道了同源盒基因 Pbx1 是脾细胞命运所必需的，并且是 Nkx2.5 和 Hox11（这对于脾脏形成也是必需的）的分层协同调节因子。我们还发现，在侧板中胚层 (LPM) 中，Pbx1 的表达开始时间早于 Nkx2.5 和 Hox11。此外，我们发现 Pbx1 在发育中的脾原基的内皮细胞中表达。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实质规范、形态发生和扩张所必需的，并且内皮细胞中的 Pbx 表达也有助于其在脾形态发生中的功能和扩展。此外，我们假设完整的间充质原基和内皮对于正常的脾造血定植和功能至关重要。使用现有的基因靶向和转基因小鼠品系，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，我们将表征脾间充质实质中 Pbx1 条件性失活的小鼠系的脾形态发生和细胞增殖缺陷，但在内皮细胞中则不然。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，我们将通过表征仅内皮细胞因 Pbx1 基因失活而改变的小鼠系来评估完整的内皮细胞是否对于脾脏形态发生和扩张至关重要。此外，通过 Pbx1 诱导失活，我们将确定脾内皮细胞中 Pbx 的时间需求。第三，我们将从遗传学角度剖析间质和内皮分别在脾脏造血定植、发育和功能中的作用。我们的研究将揭示脾脏发育背后的新型遗传和分子网络，脾脏是一个在个体发育方面被忽视的器官。鉴于间充质脾原基、侵袭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理解产生深远的影响。最后，我们的研究旨在更好地理解先天性无脾的发病机制，因为我们提出了先天性无脾的产前分子诊断的先决条件基本遗传背景。 公共健康相关性：脾脏的复杂结构和功能是不同细胞类型之间密切相互作用的结果：间充质细胞（“基础实质”）、侵入内皮细胞和定植造血细胞。在人类中，脾脏在早期造血、免疫和血液过滤中发挥着关键作用，其缺失（如先天性无脾症，一种诊断不足的疾病，通常只有在尸检时才能识别）会导致危及生命的细菌感染和致命性败血症的高风险在新生儿和儿童中。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：内皮细胞和造血细胞的形态发生、扩张和流入，因为这些相互关联的器官发生过程对于脾脏功能至关重要，并且迄今为止，大多数未知。我们预计这些研究产生的新知识将对脾脏功能的理解产生深远的影响，包括造血和免疫反应。 总之，拟议的工作将揭示脾脏个体发育背后的新型遗传和分子网络，脾脏是一个在其发育过程中被忽视的器官。最后，我们的研究旨在更好地理解先天性无脾的发病机制，因为我们提出了先天性无脾的产前分子诊断的先决条件基本遗传背景，这种情况导致对致命细菌感染的免疫反应严重受损。