Identifying pathways required for integration of kidney organoid and host epithelia

确定肾类器官和宿主上皮细胞整合所需的途径

• 批准号: 10248562
• 负责人: IAIN A. DRUMMOND
• 金额: $ 62.78万
• 依托单位: MOUNT DESERT ISLAND BIOLOGICAL LAB
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10248562
• 关键词: 3-Dimensional; Adult; Basement membrane; Bioinformatics; Biological Assay; Blood; Cause of Death; Cell Aggregation; Cell Culture System; Cell Culture Techniques; Cell fusion; Cells; Coupled; Data; Development; Dialysis procedure; Distal; End stage renal failure; Engineering; Engraftment; Environment; Epithelial; Epithelial Cells; Event; Excision; Extracellular Matrix; Fibroblast Growth Factor; Filtration; Fishes; Genetic; Goals; Human; Immune; Implant; In Situ Hybridization; In Vitro; Injury; Invaded; Investigation; Kidney; Kidney Diseases; Kidney Failure; Kidney Transplantation; Legal patent; Ligands; Liquid substance; Mammalian Cell; Matrix Metalloproteinases; Mediating; Methodology; Modeling; Molecular; Mus; Mutation; Natural regeneration; Nephrons; Organoids; Output; Pathway interactions; Patients; Pattern; Phenotype; Play; Plumbing; Pluripotent Stem Cells; Process; Renal function; Renal tubule structure; Reporting; Role; Series; Signal Pathway; Signal Transduction; Site; Structure; System; TIMP1 gene; Technology; Testing; Tissue Inhibitor of Metalloproteinases; Tissues; Urine; WNT Signaling Pathway; WNT9A gene; Work; Zebrafish; base; experimental study; genetic analysis; glucose uptake; in vivo; insight; mutant; nephrogenesis; nephron progenitor; polarized cell; prevent; programs; receptor; recruit; regenerative therapy; renal damage; repaired; scale up; specific biomarkers; stem cells; three dimensional cell culture; tool; transcriptomics; urinary

Project Summary Kidney disease is the 9th leading cause of death in the U.S. Because few therapies exist to prevent or slow progression, over 700,000 patients have End Stage Renal Disease. These patients are treated with dialysis or renal transplant, the latter resulting in markedly superior survival. However, kidney donors are limited and there is an important unmet need for strategies that enhance renal repair or generate new nephrons for renal replacement. Pluripotent stem cell derived organoids display key features of differentiated kidney tubules and glomerular structures in vitro, and we have shown that they generate patterned nephrons in vivo displaying kidney functions such as filtration and glucose uptake by the proximal tubule. To develop this technology for renal replacement, stem cell derived tubules must be connected to host tubules for urinary output. Our recent work in the zebrafish demonstrated that FGF signaling acts as a chemotactic signal to recruit and polarize cells at sites of new nephron formation and canonical Wnt signaling is required for invasive cell rearrangement to connect tubule lumens. Additional signaling pathways including non-canonical wnt signaling are also likely to play a role in tubule interconnection. To fully explore the requirements for tubule interconnection we have established a synergistic, three-part discovery platform comprising 1) genetic analysis of in vivo new nephron addition in the regenerating zebrafish adult kidney, 2) in vitro 3D cell culture analysis of mammalian epithelial fusion, and 3) in vivo stem cell-derived kidney organoid engraftment to a host mouse collecting system. We will combine these approaches to analyze multiple steps of the tubule fusion process involving 1) recruitment of nephron progenitor cells to target epithelia, 2) removal of intervening ECM/basement membranes, 3) patterned collective cell invasion of target epithelia, and 4) establishment of a continuous patent new lumen to convey the nephron filtrate. These studies will provide important new insights about an essential but understudied cellular mechanism that will be required for in vivo engraftment of new kidney tissue-based renal regeneration therapies.

项目概要 肾脏疾病是美国第九大死亡原因，因为很少有治疗方法可以预防或减缓肾脏疾病的发生 进展，超过 700,000 名患者患有终末期肾病。这些患者接受透析或 肾移植，后者的生存率明显更高。然而，肾脏捐赠者数量有限，而且 对于增强肾脏修复或生成新肾单位的策略来说，这是一个重要的未满足的需求 替代品。多能干细胞衍生的类器官显示分化肾小管的关键特征 体外的肾小球结构，我们已经证明它们在体内产生图案化的肾单位 肾脏功能，例如近曲小管的过滤和葡萄糖摄取。开发这项技术是为了 肾脏替代时，干细胞衍生的肾小管必须与宿主肾小管连接以产生尿输出。我们最近的 在斑马鱼中的研究表明，FGF 信号作为趋化信号来招募和极化细胞 在新肾单位形成的位点，侵袭性细胞重排需要典型的 Wnt 信号传导 连接小管管腔。其他信号传导途径，包括非经典 Wnt 信号传导也可能 在肾小管互连中发挥作用。为了充分探索小管互连的要求，我们有 建立了一个协同的、三部分的发现平台，包括 1) 体内新肾单位的遗传分析 添加在再生斑马鱼成年肾脏中，2) 哺乳动物上皮细胞的体外 3D 细胞培养分析 融合，3) 体内干细胞衍生的肾脏类器官移植到宿主小鼠收集系统中。我们将 结合这些方法来分析肾小管融合过程的多个步骤，包括 1) 招募 肾单位祖细胞靶向上皮细胞，2) 去除介入的 ECM/基底膜，3) 图案化 目标上皮细胞的集体细胞侵袭，以及4)建立连续的专利新管腔以传递 肾单位滤液。这些研究将为有关重要但尚未充分研究的细胞提供重要的新见解。 基于新肾组织的肾再生体内移植所需的机制 疗法。