Cellular and molecular characterization of the digestive tract sub-epithelium

消化道上皮下层的细胞和分子特征

• 批准号: 9764595
• 负责人: Ramesh A Shivdasani
• 金额: $ 37.31万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9764595
• 关键词: Address; Adult; Agonist; Anatomy; Basic Science; Biology; Bone Morphogenetic Proteins; CD34 gene; CD81 gene; Cell Line; Cell Maturation; Cells; Clinical; Colorectal Cancer; Confocal Microscopy; Data; Development; Diffuse; Digestive Physiology; Disease; Epidermal Growth Factor; Epithelial; Epithelial Cells; Epithelium; FOXL1 gene; Fibroblasts; Gastric mucosa; Gastrointestinal tract structure; Human; In Vitro; Infection; Inflammatory Bowel Diseases; Intestines; Knockout Mice; Knowledge; Lamina Propria; Ligands; Logic; Malabsorption Syndromes; Malignant Neoplasms; Mesenchymal; Mesenchyme; Microanatomy; Mitotic; Molecular; Molecular Profiling; Mus; Myofibroblast; Organ; Organoids; PDGFA gene; PDGFRA gene; Pathologic; Physiological; Platelet-Derived Growth Factor alpha Receptor; Population; Population Heterogeneity; Process; RNA analysis; Recombinants; Research; Research Project Grants; Resolution; Signal Transduction; Small Intestines; Smooth Muscle Actin Staining Method; Source; Stem cells; Stomach; Stromal Cells; Supporting Cell; Surface; Testing; Tissues; Ulcer; Villus; WNT Signaling Pathway; base; cell injury; cell type; enteric infection; experimental study; healing; improved; inhibitor/antagonist; innovation; insight; intestinal epithelium; novel; progenitor; response; self-renewal; single-cell RNA sequencing; stem; stem cell differentiation

Project Description Epithelial development, integrity, and turnover in the digestive tract depend on signals from adjacent mesenchymal cells, and defective signaling may underlie aspects of disorders such as Inflammatory Bowel Disease, pathologic response to infection, malabsorption, and cancer. Crucial mesenchymal signals include Wnt factors and bone morphogenetic protein (BMP) agonists and antagonists, and recent research implicates various (CD34+, Foxl1+, PDGFRA+, Gli1+) cell types as possible sources. However, the precise identities, overlap, and requirements of these populations remain unclear. Integrated consideration of high- resolution microscopic anatomy, molecular profiling of diverse marker-defined cells, and single-cell RNA analysis has led us to identify 3 distinct mesenchymal cell types in the mouse small intestine. Cells expressing high platelet-derived growth factor receptor A (PDGFRAhi) lie closest to the epithelium and correspond to recently described Foxl1+ telocytes. These cells express abundant BMPs and congregate particularly at the crypt-villus junction, whereas two PDGFRAlo cell populations are anatomically and molecularly distinct: those located near the crypt stem-cell zone that uniquely co-express PDGFRA, tetraspanin CD81, and the BMP antagonist Gremlin1, and cells in the villus lamina propria that lack surface CD81. These data collectively suggest the novel hypothesis that crucial crypt-villus BMP gradients reflect dense sources of agonists at the villus base and antagonists at the crypt base. Specific Aim 1 will test this hypothesis and further resolve the distinctive features and functions of mesenchymal PDGFRAhi and PDGFRAlo cells in the mouse small intestine. Building on preliminary data that unfractionated PDGFRAlo cells support expansion of ISCs in vitro, we will investigate the possibly opposing functions of pericryptal CD81+ and villus CD81- subpopulations. In response to ISC damage, the mesenchyme triggers crypt progenitors to dedifferentiate, likely through the same soluble factors that maintain uninjured ISC. Aim 2 proposes a systematic, unbiased approach to identify the specific cellular sources of key restitutional signals. We will also test the hypothesis that mesenchymal PDGFRAhi and PDGFRAlo cells depend on the intestinal epithelium to provide varying concentrations of the ligand PDGFA for their survival, proliferation, and distinct physiologic functions. ISC and crypt progenitors have counterparts in the stomach epithelium, which turns over more slowly, depends less on Wnt signaling, and does not extend villi. Mesenchymal PDGFRA+ cells are equally abundant in both organs, with likely important similarities and differences in organization and functions. To better understand gastric stem/progenitor cell control and the underlying signaling logic in both organs, Aim 3 will characterize stomach mesenchymal PDGFRA+ cells in the same depth that we have achieved in the intestine. Together, these studies apply innovative experimental approaches to examine specific hypotheses that address fundamental questions in digestive tract biology.

项目说明 消化道上皮的发育、完整性和周转依赖于来自邻近区域的信号 间质细胞和信号缺陷可能是炎症性肠病等疾病的基础 疾病、对感染的病理反应、吸收不良和癌症。重要的间充质信号 包括WNT因子和骨形态发生蛋白(BMP)激动剂和拮抗剂，以及最近研究 提示多种细胞类型(CD34+、Foxl1+、PDGFRA+、Gli1+)可能的来源。然而，准确的 这些群体的身份、重叠和要求仍然不清楚。综合考虑高- 分辨率显微解剖学、不同标记定义的细胞的分子图谱和单细胞RNA 通过分析，我们确定了小鼠小肠中三种不同的间充质细胞类型。单元格 高表达的血小板衍生生长因子受体A(PDGFRAhi)位于离上皮最近的位置，并且 与最近描述的Foxl1+端粒细胞相对应。这些细胞表达丰富的骨形态发生蛋白并聚集 特别是在隐窝-绒毛交界处，而两个PDGFRAlo细胞群在解剖学上和 分子上不同的：位于隐窝干细胞区附近的那些独特地共表达PDGFRA的细胞， Tetraspanin CD81和BMP拮抗剂Gremlin1，以及绒毛固有层中缺乏表面的细胞 CD81。这些数据共同提出了一个新的假设，即关键的隐窝绒毛BMP梯度反映了 绒毛底部有密集的激动剂来源，而地穴底部有大量拮抗剂。《特定目标1》将测试这一点 假说并进一步解析间充质PDGFRAhi和 小鼠小肠中的PDGFRAlo细胞。建立在初步数据的基础上，PDGFRAlo 细胞支持ISCs的体外扩增，我们将研究密码周围可能存在的相反功能 CD81+和绒毛CD81-亚群。作为对ISC损伤的反应，间充质触发隐窝 祖细胞可能通过维持未受损伤的ISC的相同的可溶性因子进行去分化。目标2 提出一种系统、不偏不倚的方法来确定关键修复的特定细胞来源 信号。我们还将测试间充质PDGFRAhi和PDGFRAlo细胞依赖于 为肠上皮细胞提供不同浓度的配体PDGFA，以供其生存、增殖、 和独特的生理功能。间质干细胞和隐窝祖细胞在胃上皮细胞中有对应的细胞， 它的翻转速度更慢，对Wnt信号的依赖较少，并且不会延伸绒毛。间充质 PDGFRA+细胞在两个器官中同样丰富，可能有重要的相似性和差异性 组织机构和职能。为了更好地了解胃干/祖细胞控制和潜在的 两个器官中的信号逻辑，Aim 3将在相同的器官中表征胃间充质PDGFRA+细胞 我们在肠道中达到的深度。总之，这些研究应用了创新的实验 检验解决消化道生物学基本问题的具体假设的方法。