Genes, environment & neural stem cell transplantation in the gut

描述（由申请人提供）：功能性肠神经元的丧失导致临床疾病，如失弛缓症、胃轻瘫、神经性假性梗阻、肠神经元发育不良、结肠惰性和先天性巨结肠病。尽管在这些情况下，体内缺乏显著的神经发生，但越来越多的证据表明，肠道中存在大量的多能性神经前体细胞（npc）。包括我们在内的几位研究人员已经表明，这些细胞具有一些典型的神经胶质细胞的表型标记，能够在体外进行强大的神经发生。我们现在已经证明，纵肌肌丛（LMMP）制剂中的肠神经节显示神经发生，这在完全分离的鼻咽癌培养制剂中更为明显。我们已经在体内和体外鉴定了肠内神经前体，我们的初步研究表明，这些细胞中的磷酸酶和紧张素同源物（PTEN）是体内神经发生的主要“刹车”。进一步将IV型胶原添加到LMMP制剂中，可以对神经发生产生深远的抑制作用。因此，假设体内和离体情况之间的差异可以通过调节PTEN的微环境因素来解释是合乎逻辑的。因此，在本提案中，我们打算通过以下具体目标，研究胶原IV和下游途径（包括整合素、FAK和RhoA/ROCK）的影响，这些途径可能驱动PTEN活性，检查体内肠道神经发生：具体目标1：确定健康和疾病中神经发生的外在（胶原）和内在（PTEN）抑制之间的相互作用研究在健康和疾病中对抗内在和外在抑制对体内肠道神经发生的影响特异性目标3：研究操纵这些途径的选定关键组分对同种异体肠内NSC （ENSC）移植的存活和功能的影响我们的长期目标是发展神经元移植作为人类肠道神经系统疾病的治疗方法，并刺激这些疾病中的神经再生。通过确定PTEN和相关途径的作用，拟议的研究将为新的治疗干预提供关键的基础。

DESCRIPTION (provided by applicant): Loss of functional enteric neurons results in clinical disorders such as achalasia, gastroparesis, neuropathic forms of pseudoobstruction, enteric neuronal dysplasia, colonic inertia and Hirschsprung's disease. Despite the lack of significant neurogenesis in vivo in these conditions, there is growing evidence to suggest the existence of multipotent neural precursor cells (NPCs) in abundant numbers in the gut. Several investigators including ourselves have shown that these cells, which have some phenotypical markers typical of glia, are capable of robust neurogenesis in vitro. We have now shown that enteric ganglia in longitudinal muscle myenteric plexus (LMMP) preparations display neurogenesis and this is even more pronounced in completely dissociated NPC preparations in culture. We have identified enteric neural precursors in vivo and in vitro and our preliminary studies implicate phosphatase and tensin homolog (PTEN) in these cells as the major "brake" on neurogenesis in vivo. Further collagen IV when added to LMMP preparations can have a profound inhibitory effect on neurogenesis. Thus it is logical to hypothesize that the discrepancy between the in vivo and ex vivo situation can be explained by factors in the microenvironment that regulate PTEN. In this proposal we therefore intend to examine the effects of collagen IV and downstream pathways including integrin, FAK and RhoA/ROCK which may drive PTEN activity, checking enteric neurogenesis in vivo, via the following specific aims: Specific Aim 1: To determine the interactions between extrinsic (collagen) and intrinsic (PTEN) brakes on neurogenesis in health and disease Specific Aim 2: To examine the effects of countering intrinsic and extrinsic brakes on enteric neurogenesis in vivo in health and disease Specific Aim 3: To examine the effects of manipulating selected key components of these pathways on survival and function of allogeneic enteric NSC (ENSC) transplants Our long-term objective is to develop neuronal transplantation as a treatment for human diseases of the enteric nervous system as well as stimulate neural regeneration in these disorders. By establishing a role for PTEN and related pathways the proposed studies will provide the critical groundwork for novel therapeutic interventions.