Hix in Enteric Mesenchymal and Neuronal Development

Hix 在肠间充质和神经元发育中的作用

• 批准号: 6698518
• 负责人: MICHAEL D. BATES
• 金额: $ 22.2万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-02-01 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6698518
• 关键词: beta galactosidase; cell differentiation; cell migration; confocal scanning microscopy; developmental genetics; developmental neurobiology; dopamine beta monooxygenase; embryo /fetus; embryogenesis; enteric nervous system; gastrointestinal epithelium; gene expression; gene targeting; genetic regulation; genetically modified animals; histogenesis; immunocytochemistry; in situ hybridization; laboratory mouse; mammalian embryology; mesenchyme; neural crest; protein localization; protein structure function; smooth muscle; transcription factor

Although it is clear that alterations in mesenchymal or neuronal components of the digestive system play important roles in the pathophysiology of a number of congenital motility disorders of humans, little is known about the genetic regulation of the developmental interplay between these components, or about the genetic programs required for intestinal mesenchymal differentiation. Hlx, a gene encoding a homeobox transcription factor, is required for normal development and growth of the digestive system. We have found in preliminary experiments that Hlx is required for normal development of the enteric nervous system (ENS) and intestinal mesenchyme. Our overarching hypothesis is that the Hlx transcription factor regulates intestinal mesenchymal differentiation and mesenchymal-neuronal interactions required for ENS development. In this application, we propose three specific aims to understand the role of Hlx in ENS development and intestinal mesenchymal differentiation. First, we will test the hypothesis that Hlx is required for normal migration and intestinal colonization of neural crest cells in mouse development. For these experiments we will use the dopamine beta-hydroxylase-LacZ transgene to compare the position of neural crest cells in Hlx+/+ and Hlx-/- embryos. Our second aim is to test the hypothesis that Hlx is required for normal proliferation and/or differentiation of neural crest cells in mouse development. To test this hypothesis, we will use immunohistochemistry for specific enteric neuronal markers to assess the lineages of enteric neurons present in Hlx-/- as compare to Hlx+/+ embryos. Finally, we will test the hypothesis that Hlx is required for development of primitive intestinal mesenchymal cells into smooth muscle cells, interstitial cells of Cajal, and subepithelial myofibroblasts. We will use two complementary approaches to test this hypothesis: immunohistochemistry for markers of intestinal mesenchymal cell lineages in Hlx+/+ and Hlx-/- embryos, and transgene markers of smooth muscle differentiation (reporter proteins driven by alpha- smooth muscle actin and gamma-smooth muscle actin promoters). The proposed experiments will provide novel insights into the differentiation program of intestinal mesenchyme and enteric neurons and their regulation by the Hlx transcription factor. This research is complementary to separately funded research focusing on regulation of Hlx gene expression and the identification of downstream targets of the Hlx transcription factor, particularly those important for mesenchymal-epithelial interactions in development.

虽然消化系统间充质或神经元成分的改变在许多人类先天性动力障碍的病理生理学中起着重要作用，但对这些成分之间发育相互作用的遗传调节，或肠道间充质分化所需的遗传程序，我们知之甚少。HLX是一种编码同源框转录因子的基因，是消化系统正常发育和生长所必需的。我们在初步实验中发现，HLX是肠道神经系统(ENS)和肠间充质正常发育所必需的。我们的主要假设是，HLX转录因子调节ENS发育所需的肠道间充质分化和间充质-神经元相互作用。在这一应用中，我们提出了三个特定的目标，以了解HLX在ENS发育和肠道间充质分化中的作用。首先，我们将验证HLX是小鼠发育过程中神经脊细胞正常迁移和肠道定植所必需的假设。对于这些实验，我们将使用多巴胺β-羟基酶-LacZ转基因来比较神经脊细胞在HLX+/+和HLX-/-胚胎中的位置。我们的第二个目标是验证这一假设，即HLX是小鼠发育中神经脊细胞正常增殖和/或分化所必需的。为了验证这一假设，我们将使用特定肠神经标志物的免疫组织化学方法来评估HLX-/-与HLX+/+胚胎中存在的肠神经元的谱系。最后，我们将验证HLX是原始肠间充质细胞发育成平滑肌细胞、Cajal间质细胞和上皮下肌纤维母细胞所必需的假说。我们将使用两种互补的方法来检验这一假设：免疫组织化学检测HLX+/+和HLX-/-胚胎中的肠道间充质细胞系标记物，以及平滑肌分化的转基因标记物(由α-平滑肌肌动蛋白和伽马-平滑肌肌动蛋白启动子驱动的报告蛋白)。这些实验将为肠道间充质和肠神经元的分化程序以及HLX转录因子对它们的调控提供新的见解。这项研究是对单独资助的研究的补充，重点是HLX基因表达的调控和HLX转录因子下游靶点的确定，特别是那些在发育过程中对间充质-上皮相互作用至关重要的靶点。