Engineering a Human Microphysiological System for the Characterization of Islet-Immune Interactions

设计人体微生理系统来表征胰岛免疫相互作用

• 批准号: 10665727
• 负责人: Ashutosh Agarwal
• 金额: $ 99.45万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10665727
• 关键词: 3-Dimensional; Acceleration; Address; Antigens; Beta Cell; Biological; Biological Assay; Biological Models; CD8-Positive T-Lymphocytes; Cell Culture Techniques; Cell Line; Cell physiology; Cell surface; Cells; Cellular Stress; Circulation; Clinical; Clinical Trials; Communities; Complex; Cytotoxic T-Lymphocytes; Defect; Dendritic Cells; Development; Devices; Disease; Drug Targeting; Elements; Encapsulated; Endocrine; Endothelial Cells; Endothelium; Engineering; Extracellular Matrix; Extravasation; Female; Foundations; Frequencies; Functional disorder; G6PC2 gene; Gene Transfer; Generations; Genes; Genetic; Genetic Risk; Genotype; Homing; Human; Hydrogels; Immune; In Situ; In Vitro; Insulin; Insulin-Dependent Diabetes Mellitus; Integrins; Interruption; Intervention; Islet Cell; Islets of Langerhans; Knowledge; Lymphatic; Macrophage; Measurement; Mediating; Methods; Modeling; Output; Pancreas; Pathogenesis; Pathologic; Pharmaceutical Preparations; Phase; Phenotype; Proliferating; Protocols documentation; Regulatory T-Lymphocyte; Research Personnel; Resources; Sampling; Source; Structure of beta Cell of islet; System; T-Lymphocyte; Technology; Testing; Therapeutic Intervention; Time; Tissues; Variant; Visualization; acquired factor; antigen test; antigen-specific T cells; cell killing; cell motility; cell type; clinical material; cytotoxic CD8 T cells; diabetes pathogenesis; diabetes risk; engineering design; genetic variant; genome editing; genome wide association study; graft vs host disease; high resolution imaging; humanized mouse; immune modulating agents; immunomodulatory therapies; immunoregulation; in vivo evaluation; induced pluripotent stem cell; innovation; islet; male; microphysiology system; monolayer; mouse model; novel; prevent; repository; risk mitigation; risk variant; screening; sensor; technology platform; trafficking

Summary Three dimensional (3D) microphysiological systems (MPS) represent a powerful intermediate model system employing human cells and tissues capable of bridging in vitro studies and clinical trials. We propose to create an integrated MPS platform to more accurately model the complex cellular interactions involved in human type 1 diabetes (T1D) pathogenesis. We previously generated an MPS containing novel extracellular matrix hydrogels that support sustained islet function and T cell migration along the islet cell surface in 3D (CHIB), and in first-of-their-kind studies, we demonstrated antigen-specific IGRP-reactive human CD8 T cells resulted in targeted β-cell killing (CMAI). Here, we propose an interdisciplinary effort to integrate and expand the MPS platform (referred to as the islet-immune Chip (iiChip)), as well as the cell-based technologies facilitating testing of antigen-specific T cells, isogenic cellular systems capable of deriving multiple cellular lineages, and genome editing technologies for use by the broader HIRN community. Specifically, we will utilize islets or islet- like spheroids, endothelial cell monolayers, and innate and adaptive immune cells, including dendritic cells (DCs), macrophages, CD4+ conventional T cells (Tconv), CD8+ cytotoxic T cells (CTLs), and regulatory T cells (Tregs), to model the spatial configuration and complex cellular interactions involved in human T1D pathogenesis. We hypothesize that this optimized 3D iiChip will facilitate in situ interrogation of Ag- specific and genotype-phenotype interactions that are essential in T1D pathogenesis as well as the mechanistic effects of immunomodulatory therapies with spatial and temporal control. Experimental deliverables will include the ability to assess islet:immune interactions utilizing real-time high-resolution imaging and quantitation of cellular interactions, trafficking, extravasation, and β-cell function/survival. Key features of the iiChip will involve the integration of in-line sensors and bioreporters, spatial and temporal control of inputs for defined stimulation, and integration of matrices with the capacity for fluidic and cellular recirculation, measurement of soluble and cellular readouts in long-term cell culture. In addition, gene edited induced pluripotent stem cells (iPSC) from male and female donors with T1D-risk associated HLA will be available for the generation of immune, endothelial, and endocrine cells that are essential for building an isogenic “disease-on-a-chip” model. When loaded with primary human cells or isogenic iPSC-derived materials (i.e., endothelial, immune, and β-cells), this iiCHIP will enable dynamic interrogation of genotype-phenotype interactions, antigen-specific β-cell killing, and effects of immunomodulatory therapies within a fluidic 3D microenvironment. The iiChip will enable mechanistic studies capable of expediting clinical interventions aimed at inhibition of immune-mediated β-cell destruction, enhancing immune regulation, and testing of β-cell restorative therapies.

概括 三维（3D）微生物生理系统（MP）代表强大的中间模型系统 采用能够桥接体外研究和临床试验的人类细胞和组织。我们建议创建 一个集成的MPS平台，以更准确地模拟人类类型中涉及的复杂细胞相互作用 1个糖尿病（T1D）发病机理。我们以前生成了一个包含新型细胞外基质的MPS 支持持续的胰岛功能和T细胞沿3D（CHIB）的胰岛细胞表面迁移的水凝胶， 在第一届研究中，我们证明了抗原特异性的IGRP反应性人CD8 T细胞导致 在靶向的β细胞杀伤中（CMAI）。在这里，我们提出了跨学科的努力，以整合和扩展国会议员 平台（称为胰岛免疫芯片（IICHIP））以及基于细胞的技术促进 测试抗原特异性T细胞，能够得出多个细胞谱系的同基因细胞系统， 基因组编辑技术，用于广泛的HIRN社区。具体而言，我们将利用胰岛或胰岛 像球体，内皮细胞单层以及先天和适应性免疫小球一样，包括树突状细胞 （DC），巨噬细胞，CD4+常规T细胞（TCONV），CD8+细胞毒性T细胞（CTL）和调节性T细胞 （Tregs），建模人类T1D涉及的空间构型和复杂的细胞相互作用 发病。我们假设这种优化的3D IICHIP将有助于对Ag-的原位审问 特异性和基因型 - 表型相互作用在T1D发病机理以及 具有空间和临时控制的免疫调节疗法的机械作用。实验 可交付成果将包括评估胰岛的能力：使用实时高分辨率的免疫相互作用 细胞相互作用，运输，渗出和β细胞功能/存活的成像和定量。钥匙 IICHIP的特征将涉及在线传感器和生物堆积物，空间和临时控制的集成 用于定义刺激的输入，以及具有流体和细胞能力的物品的整合 长期细胞培养中固体和细胞读数的再循环，测量。另外，基因编辑 来自具有T1D风险相关HLA的男性和女性供体的诱导多能干细胞（IPSC）将是 可用于生成免疫，内皮和内分泌细胞，这些细胞对于建立一个必不可少的 等源性的“芯片疾病”模型。当装有原代人细胞或同源IPSC衍生的材料时 （即内皮，免疫和β细胞），该IICHIP将对基因型 - 表型进行动态询问 相互作用，抗原特异性β细胞杀伤以及流体3D中免疫调节疗法的影响 微环境。 IICHIP将实现能够加快针对临床干预措施的机械研究 抑制免疫介导的β细胞破坏，增强免疫调节和测试β细胞 恢复疗法。

项目成果

Spermatogonial Stem Cells and In Vitro Spermatogenesis: How Far Are We from a Human Testis on a Chip?

精原干细胞和体外精子发生：我们离芯片上的人类睾丸还有多远？

• DOI: 10.1016/j.euf.2022.11.006
• 发表时间: 2023
• 期刊: European urology focus
• 影响因子: 5.4
• 作者: Ramsoomair,ChristianK;Alver,CharlesG;Flannigan,Ryan;Ramasamy,Ranjith;Agarwal,Ashutosh
• 通讯作者: Agarwal,Ashutosh