Genes, environment & neural stem cell transplantation in the gut

描述(申请人提供)：功能肠神经元的丧失会导致临床疾病，如失弛缓症、胃瘫、神经病理性假性梗阻、肠神经元发育不良、结肠无力和先天性巨结肠。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潜能神经前体细胞(NPC)。包括我们自己在内的几位研究人员已经证明，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再生。我们现在已经证明，纵肌肌间神经丛的肠神经节在培养中表现出神经再生，这在完全分离的鼻咽癌标本中更加明显。我们已经在体内和体外鉴定了肠神经前体细胞，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”。此外，当将IV型胶原添加到LMMP制剂中时，可以对神经发生产生深刻的抑制作用。因此，假设体内和体外情况之间的差异可以用调节PTEN的微环境因素来解释是合乎逻辑的。因此，在这项建议中，我们打算研究IV型胶原及其下游通路，包括整合素、FAK和RhoA/ROCK，它们可能推动PTEN活性，检查体内肠神经发生，通过以下特定目的：特定目的1：确定外源性(胶原)和内在(PTEN)刹车在健康和疾病中对神经发生的相互作用2：检测对抗内源性和外源性刹车对健康和疾病体内肠道神经发生的影响3：检查操纵这些通路中选定的关键成分对同种异体肠神经干细胞(ENSC)移植存活和功能的影响我们的长期目标是开发神经元移植作为治疗人类肠道神经系统疾病的方法，并刺激这些疾病中的神经再生。通过确定PTEN和相关通路的作用，拟议的研究将为新的治疗干预措施提供关键的基础。

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.