F31 Mulero Russe

项目摘要 炎症性肠病（IBD）目前影响超过160万美国人。IBD的特征是破坏 肠上皮细胞和高度炎症的环境。现有的治疗方法针对炎症 然而，通过生物制剂，较少的努力集中在上皮愈合上， 适用于修复肠上皮的疗法。人类肠道类器官（HIO）是三维的（3D） 多细胞结构，来源于成体肠干细胞或人多能干细胞（hPSC）， 它重现了人类肠道组织结构。HIO是一种有前途的肠上皮细胞来源 修复、疾病建模和药物筛选。以前的工作已经证明，HIO移植到受伤的 然而，由于缺乏适当的递送，这些方法受到显著限制 车辆驱动HIO植入。从hPSC产生HIO是多阶段定向分化过程 包括三个阶段：（I）分化成定形内胚层单层，（II）后肠和原管 分化成自由漂浮的、自组织的3D聚集体（人肠球状体，HIS），和（III） 在3D细胞外基质内将肠特化为HIO。这种体外培养过程跨越了2D生长 从基底（阶段I和II）到3D矩阵（阶段III）。第三阶段需要在Matrigel中培养，Matrigel是一种鼠肿瘤- 衍生的基底膜提取物成分不明确、批次间变异性和临床应用有限 翻译潜力HIO技术的另一个障碍是HIS差异化的低产量和一致性 在HIO中。本项目的目标是（1）设计一个具有独立控制的合成水凝胶平台 引导hPSC在体外分化为HIO的整个过程的生物化学和生物物理学线索，以及（2） 将合成涂层中的HIO递送至体内肠损伤。核心假设是， 具有适当的生物物理和生物化学线索的合成基质将支持HIO自组织，生长， 和分化过程，并增强HIO植入和肠伤口愈合。目标1：工程师a 促进从hPSC产生球状体的2D合成基质。目标2：评估HIO的成熟度， 合成龛内生成的球状体。目标3：设计一种临床上可翻译的治疗方法 用于HIOs的材料到受伤的肠组织。这项研究的结果将增加临床相关性， 生成HIO，并将为差异化和交付提供可扩展和可翻译的合成材料 的HIO。

PROJECT SUMMARY Inflammatory Bowel Diseases (IBD) currently affect >1.6 million Americans. IBD is characterized by disruption to the intestinal epithelium and a high inflammatory environment. Available treatments target the inflammation through biological agents, however, fewer efforts have focused on epithelium healing and there are no broadly applicable therapies to repair intestinal epithelium. Human intestinal organoids (HIOs) are three-dimensional (3D) multicellular structures, derived from either adult intestinal stem cells or human pluripotent stem cells (hPSCs), that recapitulate human intestinal tissue architecture. HIOs are a promising cell source for intestinal epithelium repair, disease modeling, and drug screening. Previous work has demonstrated that HIOs engraft to the injured intestinal wall in vivo, however, these approaches are significantly limited by the lack of an appropriate delivery vehicle to drive HIO engraftment. HIO generation from hPSCs is multi-stage directed differentiation process comprising three stages: (I) differentiation into a definitive endoderm monolayer, (II) hindgut and primitive tube differentiation into free-floating, self-organized 3D aggregates (human intestinal spheroids, HIS), and (III) intestinal specification into HIOs within a 3D extracellular matrix. This in vitro culture process spans a 2D growth substrate (stage I and II) to a 3D matrix (stage III). Stage III requires culture within Matrigel, a murine tumor- derived basement membrane extract with ill-defined composition, lot-to-lot variability, and limited clinical translation potential. Another roadblock to HIO technologies is the low yield and consistency of HIS differentiation in HIOs. The objectives of this project are to (1) engineer a synthetic hydrogel platform with independent control of the biochemical and biophysical cues guiding the entire in vitro differentiation of hPSCs into HIOs, and (2) deliver HIOs in a synthetic coating to intestinal injuries in vivo. The central hypothesis is that an engineered synthetic matrix with appropriate biophysical and biochemical cues will support the HIO self-organization, growth, and differentiation process and enhance HIO engraftment and healing of intestinal wounds. Aim 1: Engineer a 2D synthetic matrix promoting spheroid generation from hPSCs. Aim 2: Evaluate the maturation of HIOs from the generated spheroids within synthetic niches. Aim 3: Engineer a clinically translatable therapeutic delivery material for HIOs to injured intestinal tissue. The results of this study will increase the clinical relevance the generated HIOs and will provide a scalable and translatable synthetic material for the differentiation and delivery of HIOs.