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Uncovering the therapeutic potential of adipose tissue derived neural stem cells for Hirschsprung's disease.

揭示脂肪组织来源的神经干细胞治疗先天性巨结肠的潜力。

基本信息

批准号：
10580052
负责人：
ALLAN M GOLDSTEIN
金额：
$ 20.62万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-03-01 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10580052
关键词：
Address Adipose tissue Affect Blood flow Cell Separation Cells Central Nervous System Clinical Clinical Trials Coculture Techniques Colon Colonic Aganglionosis Colorectal Compensation Complex Congenital Megacolon Congenital Neuropathy Constipation Cues Data Deposition Disease Dose Engraftment Enteral Enteric Nervous System Enterocolitis Environment Excision Fatty acid glycerol esters Fecal Incontinence Fluorescence-Activated Cell Sorting Ganglia Gastrointestinal tract structure Goals Harvest Health Human In Vitro Intestinal Secretions Intestines Investigation Knowledge Laboratory culture Life Liquid substance Methods Microsurgery Morbidity - disease rate Movement Mus Nerve Fibers Nerve Tissue Nervous System Neuroglia Neurons Neuropathy Operative Surgical Procedures Patients Physiological Pluripotent Stem Cells Population Postoperative Complications Pre-Clinical Model Procedures Process Property Protocols documentation Rectum Regenerative Medicine Reporting Research Resolution Risk Schwann Cells Scientific Advances and Accomplishments Signal Transduction Smooth Muscle Source Suction Lipectomy System Therapeutic Tissues Transplantation Vasomotor adipose derived stem cell cell motility clinical application experimental study human stem cells human tissue implantation in vivo innovation interest migration minimally invasive mouse model nerve stem cell nervous system disorder neural neurogenesis novel patient safety postnatal postnatal human preservation prevent rectal response single nucleus RNA-sequencing stem cell biomarkers stem cell migration stem cell population stem cell therapy stem cells subcutaneous treatment strategy tumorigenic

项目摘要

PROJECT SUMMARY The overarching objective of our research is to develop a stem cell therapy from subcutaneous fat tissue (SAT) to replace the congenitally absent enteric nervous system (ENS) in Hirschsprung disease (HSCR). Surgical resection of the affected colon is currently the only viable treatment for HSCR. This is a necessary life-saving procedure; albeit, more than 50% of patients still suffer from postoperative complications including constipation, fecal incontinence, and enterocolitis. To overcome these morbidities, investigations into treatments that can preserve the rectum and its functions are warranted. Replacement of the absent ENS via stem cell therapies is touted as the most promising treatment strategy to achieve this goal. Our group has demonstrated the feasibility of stem cell treatments by harvesting neural stem cells (NSCs) from the gut which engraft, migrate and differentiate into neuronal networks when transplanted into mice with HSCR. For clinical application this would require surgical resection of a piece of intestine. To prevent unnecessary resection surgery, other sources of NSCs are of interest. Human fat (adipose) tissue contains a reservoir of stem cells that are readily accessible. These cells have been examined in over 270 clinical trials for numerous diseases that support favourable patient safety profiles. In our preliminary data we have also identified that nerve fiber bundles from murine fat deposits – subcutaneous adipose tissue (SAT) - harbor an endogenous source of NSCs that are unexplored for the treatment of neuropathies. We predict that the SAT could provide a useful source of NSCs to treat colonic aganglionosis in HSCR; however, it remains undetermined if SAT-NSCs can undergo neurogenesis in the aganglionic (absent ENS) environment of the gut and there are currently no methods to purify and expand human SAT-NSCs. In the first aim of this study, we will determine if purified SAT-NSCs from mice are capable of neurogenesis in aganglionic intestine. The ganglionated ENS is supplemented postnatally by NSCs that migrate into the gut from extrinsic nerve fiber bundles and differentiate into enteric neurons in response to environmental cues from the gut. We will address whether SAT-NSCs can also undergo enteric neurogenesis when provided signalling cues from the ganglionic and aganglionic gut in in vitro coculture systems and via microsurgical SAT-NSC implantation in vivo. To determine how to isolate and expand human SAT-NSCs we will address the paucity of knowledge on the origin of these cells. To accomplish this, cells isolated from human SAT nerve fiber bundles will be unbiasedly characterised by single nuclei RNA-Seq before and after stem cell culture procedures. Cells expressing NSC markers will be purified by fluorescence activated cell sorting and their differentiation potential will be assessed in in vitro culture and in ex vivo transplants to the smooth muscle of the gut. The results of these studies will establish procedures to isolate SAT-NSCs and assess their potential to treat the congenital neuropathy in HSCR.

项目概要我们研究的首要目标是开发皮下脂肪组织的干细胞疗法（SAT）取代先天性缺失的先天性巨结肠症（HSCR）肠神经系统（ENS）。手术切除受影响的结肠是目前 HSCR 唯一可行的治疗方法。这是必要的救生程序；尽管如此，超过50%的患者仍然遭受术后并发症，包括便秘、大便失禁和小肠结肠炎。为了克服这些疾病，需要进行调查可以保护直肠及其功能的治疗是必要的。通过以下方式替换缺失的 ENS 干细胞疗法被誉为实现这一目标的最有希望的治疗策略。我们组有证明了通过从肠道采集神经干细胞（NSC）进行干细胞治疗的可行性当移植到具有 HSCR 的小鼠体内时，可移植、迁移并分化为神经元网络。用于临床应用这需要手术切除一段肠道。为了防止不必要的切除手术之外，其他来源的 NSC 也令人感兴趣。人体脂肪组织含有干细胞库易于访问。这些细胞已在 270 多项针对多种疾病的临床试验中得到检验支持良好的患者安全状况。在我们的初步数据中，我们还确定了神经纤维来自小鼠脂肪沉积物的束——皮下脂肪组织（SAT）——含有内源性脂肪尚未探索用于治疗神经病的神经干细胞。我们预测 SAT 可以提供有用的帮助用于治疗 HSCR 中结肠无神经节细胞症的 NSC 来源；然而，SAT-NSC 是否可以在肠道的无神经节（缺乏 ENS）环境中进行神经发生，目前还没有纯化和扩增人类 SAT-NSC 的方法。在本研究的第一个目标中，我们将确定是否纯化来自小鼠的 SAT-NSC 能够在无神经节肠中进行神经发生。神经节化 ENS 是出生后通过从外在神经纤维束迁移到肠道的 NSC 进行补充响应来自肠道的环境信号分化为肠神经元。我们将讨论是否当来自神经节和神经节的信号线索提供时，SAT-NSC 也可以经历肠神经发生。体外共培养系统中的无神经节肠和通过体内显微外科 SAT-NSC 植入。到确定如何分离和扩展人类 SAT-NSC，我们将解决这方面知识的匮乏问题这些细胞的起源。为了实现这一目标，从人类 SAT 神经纤维束中分离出的细胞将被公正地通过干细胞培养程序之前和之后的单核 RNA 测序进行表征。表达 NSC 的细胞标记物将通过荧光激活细胞分选进行纯化，并且它们的分化潜力将是在体外培养和离体移植到肠道平滑肌中进行评估。这些结果研究将建立分离 SAT-NSC 的程序并评估其治疗先天性 HSCR 中的神经病变。