A Human Organoid Model of Polycystic Kidney Disease

多囊肾病的人体类器官模型

基本信息

批准号：
10190922
负责人：
Benjamin Solomon Freedman
金额：
$ 35.3万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-15 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10190922
关键词：
Actomyosin Adhesions Affect Animal Model Animals Architecture Biochemical Cell Adhesion Cell Line Cell membrane Cell model Cell surface Cells Cilia Complement Complex Cyst Cystic Fibrosis Cystic Fibrosis Transmembrane Conductance Regulator Cytoskeleton Data Defect Disease Embryo Endoplasmic Reticulum Epithelial Cells Excision Experimental Models Extracellular Matrix Fibrosis Genes Genetic Genetic Diseases Genetic Transcription Goals Hereditary Disease Heterozygote Human Inherited Integral Membrane Protein Intervention Kidney Kidney Diseases Kidney Failure Laboratories Life Liquid substance Lysosomes Membrane Modeling Molecular Mus Mutation Myosin ATPase Myosin Type II Natural regeneration Nature Nephrons Organ Organoids PKD2 protein Pathway interactions Patients Pattern Pharmaceutical Preparations Pharmacology Phenocopy Phenotype Polycystic Kidney Diseases Process Proteins Publishing Receptor Signaling Renal tubule structure Research Role Safety Site Source Structure Surface System Techniques Testing Therapeutic Therapeutic Intervention Time Tissue Sample Tissues Tube Tubular formation Work base cell immortalization cohort experimental study human model human pluripotent stem cell human tissue innovation loss of function mutation mouse model multicatalytic endopeptidase complex mutant non-muscle myosin novel polycystic kidney disease 1 protein prenatal prevent receptor stem cell model stoichiometry success targeted treatment therapeutic candidate trafficking

项目摘要

A HUMAN ORGANOID MODEL OF POLYCYSTIC KIDNEY DISEASE ! PROJECT SUMMARY Polycystic kidney disease (PKD) is the world's most common life-threatening genetic disease and fourth-leading cause of kidney failure, affecting approximately 12 million people worldwide. In PKD, the normal tubular architecture of the kidneys and other organs is gradually replaced by cysts and fibrosis. There is no cure for PKD, and candidate therapeutics are of uncertain efficacy and safety. PKD is commonly inherited as a germline heterozygous loss-of-function mutation in PKD1 or PKD2, encoding polycystin-1 (PC1) and polycystin-2 (PC2), respectively. These large, transmembrane proteins form a channel-receptor complex at the primary cilium and other sites. It is not yet known how mutations result in cyst formation from tubular epithelial cells. A major barrier to deciphering PKD mechanistically is the lack of experimentally accessible models that faithfully recapitulate PKD- specific cystogenesis from tubules. Primary and immortalized cell lines are heterogenous, de- differentiated, non-human, or represent only later stages of disease, while animal models differ substantially from humans and are challenging to decipher mechanistically. To overcome this gap, our laboratory is pioneering the use of human pluripotent stem cells (hPSC) for modeling PKD. hPSC represent a very early embryonic state and provide a renewable source of patient-matched human cells for analysis and regeneration. We have established techniques to differentiate hPSC into human kidney organoids, which are complex, multicellular structures with patterned segments that resemble nephrons. We have further compared gene-edited and patient-derived organoids to mouse and human tissue samples with disease to complement and validate this new system. In hPSC with PKD mutations, we have identified several disease-relevant phenotypes, including cystogenesis from kidney organoid tubules. The goal of this proposal is to elucidate the mechanistic determinants of human PKD cystogenesis in kidney organoids. Based on our preliminary data, we hypothesize that balanced expression of PC1 and PC2 regulates physical adhesion between the kidney tubule and its microenvironment. In Aim 1, we will establish a more faithful experimental model of human PKD by quantifying cystogenesis in heterozygous kidney organoids. In Aim 2, we will investigate how polycystin protein levels are controlled by clarifying the mechanisms underlying PC1 loss in PKD2-/- cells. Finally, Aim 3 will explore a novel hypothesized molecular pathway for PKD by identifying defects in cell adhesion during cyst initiation. Key findings will be validated in primary PKD tissues and non-organoid cells. Collectively, these studies will unveil critical molecular pathways that underlie the enigmatic process of cyst formation, revealing new potential targets for therapeutic intervention.

多囊性肾脏疾病的人体器官模型呢项目摘要多囊性肾脏疾病（PKD）是世界上最常见的威胁生命的遗传疾病和肾衰竭的第四个领先原因，全世界约有1200万人。在PKD中，肾脏和其他器官的正常管状结构逐渐被囊肿和纤维化。 PKD无法治愈，候选治疗具有不确定的功效和安全性。 PKD通常被遗传为PKD1或PKD2中的种系杂合功能丧失突变分别编码Polycystin-1（PC1）和Polycystin-2（PC2）。这些大的跨膜蛋白质在原发性纤毛和其他部位形成通道受体复合物。尚不清楚突变如何导致肾小管上皮细胞形成囊肿。解密PKD的主要障碍机械上是缺乏实验可访问的模型，这些模型忠实地概括了PKD- 小管的特异性囊肿发生。原发性和永生的细胞系是异质的，差异化，非人类或仅代表疾病的后期阶段，而动物模型则不同基本上是来自人类的，具有挑战性的机械师。为了克服这一差距，我们的实验室正在开创使用人多能干细胞（HPSC）进行PKD建模的。 HPSC代表非常早期的胚胎状态，并提供可再生的患者匹配来源人类细胞进行分析和再生。我们已经建立了将HPS分为区分的技术人类肾脏类器官，具有复杂的多细胞结构，具有图案段类似于肾单位。我们进一步比较了基因编辑和患者衍生的器官与小鼠和疾病的人体组织样本以补充和验证该新系统。在HPSC中 PKD突变，我们已经确定了几种与疾病的表型，包括来自肾脏类小管。该提议的目的是阐明肾脏器官中的人类PKD囊肿发生。根据我们的初步数据，我们假设 PC1和PC2的平衡表达调节肾小管及其肾小管之间的身体粘附微环境。在AIM 1中，我们将建立一个更忠实的人类PKD实验模型量化杂合肾脏器官中的囊肿性。在AIM 2中，我们将调查如何通过阐明PKD2 - / - 中PC1损失的机制来控制多囊蛋白蛋白水平细胞。最后，AIM 3将通过识别新型假设的PKD分子途径囊肿开始期间细胞粘附的缺陷。主要发现将在原发性PKD组织中进行验证和非甲状腺细胞。总的来说，这些研究将揭示基础的关键分子途径囊肿形成的神秘过程，揭示了治疗干预的新潜在靶标。