Genes, Environment & Neural Stem Cell Transplantation in the Gut

基因、环境

• 批准号: 8094452
• 负责人: PANKAJ J PASRICHA
• 金额: $ 34.17万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-15 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8094452
• 关键词: Apoptotic; Biological Factors; Congenital Megacolon; Constipation; Data; Deglutition; Development; Disease; Disease model; Embryo; Enteral; Enteric Nervous System; Environment; Enzymes; GDNF gene; Gastrointestinal Motility; Gastrointestinal tract structure; Gastroparesis; Genes; Genetic; Goals; Health; Immune response; Inflammatory; Intestinal Obstruction; Knowledge; Laboratories; Movement; Mus; Muscle; Nausea and Vomiting; Nerve; Nervous System Physiology; Neuraxis; Neuronal Differentiation; Neurons; Neuropathy; Nitric Oxide Synthase Type I; Outcome; Pathway interactions; Peripheral; Pharmaceutical Preparations; Phenotype; Relaxation; Role; Signal Transduction; Societies; Source; Stem cell transplant; Stomach; Symptoms; Techniques; Testing; Therapeutic; Therapeutic Uses; Transplantation; Validation; Work; adult stem cell; burden of illness; clinically relevant; gastrointestinal function; human PTCH2 protein; human disease; improved; mutant; nerve stem cell; neurotrophic factor; receptor; research study

DESCRIPTION (provided by applicant): Neural stem cell (NSC) transplantation is an emerging technique with immense promise for the treatment of disorders of the peripheral (PNS) and central nervous system (CNS). More recently, our laboratory has begun investigating the use of NSC as a viable therapeutic strategy for disorders of gastrointestinal motility. Using a strategy of transplanting CNS-derived NSC (CNS-NSC) in mice with a genetic deletion of the neuronal nitric oxide synthase (nNOS) enzyme, we have shown that CNS-NSC can successfully engraft and differentiate into nNOS expressing neurons, can restore nitrergic relaxation and "rescue" gastric function (emptying) in these mice. We have also shown that intrinsic apoptotic signaling is more important in determining the survival of transplanted NSC than the host immune response. The results of these experiments have generated considerable confidence in the viability of this strategy as a potential therapy. Further validation of this approach will require a greater knowledge of the mechanisms responsible for differentiation and survival of NSC in the gut. In this regard, we have initially focused on the most well characterized pathway responsible for the normal development of the enteric nervous system, glial derived neurotrophic factor or GDNF, and its cognate receptor, RET. Our preliminary data, using CNS-NSC transfected with a constitutively active RET mutant (RET/PTC2) suggest that activation of this pathway results in greater proliferation, survival and neuronal differentiation of NSC. Our overall hypothesis is that activation of RET is sufficient to improve the survival and differentiation of CNS-NSC after transplantation into the gut and that this can be an effective therapeutic strategy to replace enteric nervous system function. To test this hypothesis, we will pursue the following specific aims using NSC isolated from the CNS (CNS-NSC) and the gut (ENS-NSC): Specific Aim 1: To use an inducible, constitutively active RET/PTC2 gene construct to determine the duration of RET activation required for an optimal outcome after transplantation. Specific Aim 2: To determine if RET-activation can produce a similar phenotype in either CNS or ENS derived NSC. Specific Aim 3: To determine if RET-activated CNS and ENS-derived NSC can effectively restore enteric neuronal function in clinically relevant models of disease such as Hirschsprung's disease and inflammatory neuropathies. Our long-term objective is to develop neuronal transplantation as a treatment for human diseases of the enteric nervous system. To establish the optimal source of stem cells (adults versus embryonic; CNS versus ENS etc.) currently remains an elusive goal and will need several more years of work by many laboratories across the world. By establishing a permissive and necessary role of RET, the proposed studies will provide the critical groundwork for the therapeutic use of neuronal precursors from a variety of sources. PUBLIC HEALTH RELEVANCE: Disorders of gastrointestinal motility are common and pose a significant burden of disease on our society. These disorders results from disturbances in the function of the nerves and muscle responsible for orderly movement within the digestive tract and result in symptoms such as severe constipation, intestinal obstruction, nausea and vomiting from delayed gastric emptying or difficulty in swallowing. Several of these conditions are caused by loss or abnormal function of gastrointestinal nerves and cannot be treated satisfactorily by drugs alone. An ideal strategy for these disorders is to restore nerve function by transplanting neural stem cells. It is the purpose of this proposal to study this approach experimentally and determine the biological factors that can optimize the use of neural stem cells for the treatment and perhaps cure of these diseases.

描述(申请人提供)：神经干细胞(NSC)移植是一种新兴技术，在治疗周围神经系统(PNS)和中枢神经系统(CNS)疾病方面前景广阔。最近，我们的实验室开始研究NSC作为胃肠动力障碍的一种可行的治疗策略。在神经元型一氧化氮合酶(NNOS)基因缺失的小鼠体内移植CNS-NSC(CNS-NSC)的策略表明，CNS-NSC可以成功地移植并分化为表达nNOS的神经元，并能恢复氮能松弛和“挽救”这些小鼠的胃功能(排空)。我们还表明，在决定移植的神经干细胞存活方面，内在的凋亡信号比宿主免疫反应更重要。这些实验的结果使人们对这一策略作为一种潜在疗法的可行性产生了相当大的信心。要进一步验证这一方法，需要更多地了解肠道内神经干细胞分化和存活的机制。在这方面，我们最初关注的是负责肠道神经系统正常发育的最具特征性的通路，胶质源性神经营养因子或GDNF及其同源受体RET。我们的初步数据表明，用RET突变体(RET/PTC2)转染的CNS-NSC可以促进NSC的增殖、存活和神经元分化。我们的总体假设是，RET的激活足以改善CNS-NSC移植到肠道后的存活和分化，这可能是替代肠道神经系统功能的一种有效的治疗策略。为了验证这一假设，我们将利用从中枢神经系统分离的神经干细胞(CNS-NSC)和肠道神经干细胞(ENS-NSC)来实现以下特定目的：特定目标1：使用可诱导的、具有结构性活性的RET/PTC2基因结构来确定移植后获得最佳结果所需的RET激活持续时间。具体目标2：确定RET激活是否能在CNS或ENS来源的NSC中产生类似的表型。具体目的3：确定RET激活的CNS和ENS来源的NSC是否能有效地恢复临床相关疾病模型(如先天性巨结肠和炎症性神经病)的肠神经功能。我们的长期目标是开发神经移植作为治疗人类肠道神经系统疾病的一种方法。建立干细胞的最佳来源(成体与胚胎、中枢神经系统与胚胎干细胞等)目前仍然是一个难以实现的目标，世界各地的许多实验室还需要几年的工作。通过建立RET允许的和必要的作用，拟议的研究将为来自各种来源的神经元前体的治疗应用提供关键的基础。公共卫生相关性：胃肠动力障碍很常见，给我们的社会带来了严重的疾病负担。这些疾病是由负责消化道内有序运动的神经和肌肉的功能障碍引起的，并导致严重便秘、肠梗阻、胃排空延迟或吞咽困难引起的恶心和呕吐等症状。其中几种情况是由胃肠神经功能丧失或异常引起的，单靠药物治疗不能令人满意。治疗这些疾病的理想策略是通过移植神经干细胞来恢复神经功能。这项建议的目的是对这一方法进行实验研究，并确定可以优化使用神经干细胞治疗甚至治愈这些疾病的生物学因素。