Hix in Enteric Mesenchymal and Neuronal Development

虽然很明显，消化系统的间质或神经元成分的改变在许多人类先天性运动障碍的病理生理中起着重要作用，但对这些成分之间发育相互作用的遗传调控或肠间质分化所需的遗传程序知之甚少。Hlx是一种编码同源盒转录因子的基因，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是肠神经系统（ENS）和肠间质正常发育所必需的。我们的总体假设是，Hlx转录因子调节肠间质分化和ENS发育所需的间质-神经元相互作用。在这项应用中，我们提出了三个具体目标来了解Hlx在ENS发育和肠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嵴细胞正常迁移和肠道定植所必需的假设。在这些实验中，我们将使用多巴胺β -羟化酶- lacz转基因来比较神经嵴细胞在Hlx+/+和Hlx-/-胚胎中的位置。我们的第二个目标是验证Hlx是小鼠发育过程中神经嵴细胞正常增殖和/或分化所必需的假设。为了验证这一假设，我们将使用免疫组织化学特异性肠神经元标记物来评估与Hlx+/+胚胎相比，Hlx-/-胚胎中存在的肠神经元谱系。最后，我们将验证Hlx是原始肠间充质细胞发育为平滑肌细胞、Cajal间质细胞和上皮下肌成纤维细胞所必需的假设。我们将使用两种互补的方法来验证这一假设：免疫组化对Hlx+/+和Hlx-/-胚胎肠间充质细胞系的标记，以及平滑肌分化的转基因标记（由α -平滑肌肌动蛋白和γ -平滑肌肌动蛋白启动子驱动的报告蛋白）。本实验将为肠间质和肠神经元的分化程序及其受Hlx转录因子调控提供新的见解。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转录因子下游靶标的鉴定，特别是那些在发育过程中对间充质-上皮相互作用重要的研究。

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.