A microphysiological system of tendon inflammation and fibrosis for drug screening and efficacy testing

用于药物筛选和疗效测试的肌腱炎症和纤维化的微生理系统

• 批准号: 10674534
• 负责人: Hani A Awad
• 金额: $ 72.19万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-15 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10674534
• 关键词: 3-Dimensional; Acceleration; Acute; Adhesions; Antibodies; Biological; Biological Assay; Biological Markers; Biological Models; Blood Tests; Blood Vessels; Blood flow; Cell Cycle Regulation; Cells; Chronic; Cicatrix; Circulation; Clinical; Clinical Trials; Collagen; Complex; Confocal Microscopy; Connective and Soft Tissue; Detection; Devices; Disease; Disease model; Dose; Drug Screening; Endothelial Cells; Endothelium; Engineering; Enzyme-Linked Immunosorbent Assay; Evaluation; Extracellular Matrix; Extravasation; FDA approved; FRAP1 gene; Fibroblasts; Fibrosis; Flexor; Gamma-H2AX; Gene set enrichment analysis; Goals; Health; Human; Hydrogels; Image; Immunology; Impairment; In Situ; Infiltration; Inflammation; Inflammation Mediators; Inflammatory; Intervention Trial; Label; Libraries; Link; Macrophage; Measurement; Measures; Mechanics; Mediating; Membrane; Microfabrication; Microfluidic Microchips; Microfluidics; Microscopic; Microscopy; Molecular; Mus; Musculoskeletal; Myofibroblast; Nanoporous; Operative Surgical Procedures; Optics; Outcome; PI3K/AKT; Paracrine Communication; Pathology; Patients; Periodicity; Permeability; Pharmaceutical Preparations; Phase; Phenotype; Porosity; Procedures; Process; Protein Secretion; Publishing; Replacement Arthroplasty; Role; SDZ RAD; Safety; Sampling; Sensitivity and Specificity; Serum; Side; Signal Transduction; Silicon; Sirolimus; Spinal Fusion; System; Tendinopathy; Tendon Injuries; Tendon structure; Testing; Therapeutic; Time; Tissue Engineering; Tissues; Transforming Growth Factor beta; Translating; Vascular Endothelial Cell; biological development; biomarker selection; cell motility; clinically relevant; design; drug candidate; drug discovery; drug efficacy; druggable target; efficacy evaluation; efficacy testing; experience; healing; high throughput screening; human tissue; improved; induced pluripotent stem cell; induced pluripotent stem cell technology; inhibitor; injured; innovation; intercellular communication; joint function; mTOR Inhibitor; manufacture; meter; microphysiology system; microscopic imaging; mouse model; nano; nanofabrication; next generation sequencing; organ on a chip; photonics; preclinical trial; primary outcome; programs; regenerative therapy; repaired; secondary outcome; senescence; sensor; sensor technology; virtual clinical trial

Acute and chronic tendon injuries are among the most common musculoskeletal health problems. Typically, an injured tendon experiences fibrotic scarring that leaves the tissue mechanically compromised and prone to debilitating adhesions that impair joint function. In a fibrotic tendon scar, the cell-cell and paracrine signaling between inflammatory cells, such as macrophages, and tendon fibroblasts activate the latter into fibroproliferative myofibroblasts, ultimately differentiating into a senescent phenotype. Our previous studies using next-generation sequencing and gene set enrichment analysis mechanistically linked fibrosis and senescence in injured mouse tendons with TGF-beta activated mTOR signaling. To further elucidate this pathology, the goal of this proposal is to engineer a microfluidic human tendon-on-chip (hToC) system and use it to more accurately model the biological aspects of the inflammation and fibrosis in injured tendons. In the UG3 phase of this proposal, the chip will be fabricated featuring a multicompartmental design and microfluidic channels to incorporate a fibroblast-seeded collagen hydrogel and simulate vascular blood flow, respectively. Ultrathin, highly permeable, and optically transparent porous silicon membranes (SiM) will separate the hydrogel from circulation and provide a substrate for an endothelial barrier in between. The signaling between the fibroblasts, hydrogel-resident- and circulating-macrophages, and endothelial cells will be enabled through nanoporous SiM (~60 nm), while a microporous SiM (~ 8 µm) will allow extravasation of circulating macrophages and infiltration of the hydrogel under TGF-beta stimulation. To allow for a patient-centric chip, tendon fibroblasts will be used to create the tendon hydrogel and to reprogram donor-matching iPSCs to derive the endothelial cells and macrophages, respectively. An additional innovation will be the integration of label- free photonic sensors into the microfluidic device to allow on-chip sensing, which has been long appreciated as a critical, unmet need for organ-on-chip devices. The UG3 studies will use the chip to validate the role of mTOR in the disease model and identify biologically relevant biomarkers. In the UH3 phase, we will utilize the chip as a pre-clinical trial platform for testing efficacy and safety of FDA-approved mTOR inhibitors as potential disease modifying drugs, and as a drug screening platform to identify and prioritize safer and more potent inhibitors of mTOR and senescence in tendon injury for clinical trials. To successfully complete this innovative project, we have assembled a team of accomplished experts in tendon tissue engineering and surgery, immunology, iPSC technology, GMP cell manufacturing, nano- and micro-fabrication, sensor technology, and high throughput screening. The proposed studies will develop a human microphysiological system to catalyze clinical trials and accelerate drug discovery for acute and chronic tendon injuries.

急性和慢性肌腱损伤是最常见的肌肉骨骼健康问题。通常，一个 受伤的肌腱会形成纤维化疤痕，使组织机械受损并容易发生 削弱关节功能的粘连。在纤维化肌腱疤痕中，细胞间和旁分泌信号传导 炎症细胞（例如巨噬细胞）和肌腱成纤维细胞之间的相互作用激活后者 纤维增殖性肌成纤维细胞，最终分化为衰老表型。我们之前的研究 使用下一代测序和基因集富集分析在机制上关联纤维化和 TGF-β 激活 mTOR 信号传导导致小鼠损伤肌腱衰老。为了进一步阐明这一点 病理学，该提案的目标是设计微流体人体腱芯片（hToC）系统并使用 它可以更准确地模拟受伤肌腱炎症和纤维化的生物学方面。在 该提案的 UG3 阶段，该芯片将采用多室设计和微流控技术制造 通道分别掺入成纤维细胞种子胶原水凝胶和模拟血管血流。 超薄、高渗透性和光学透明的多孔硅膜 (SiM) 将分离 水凝胶从循环中分离出来，并为其间的内皮屏障提供基质。之间的信令 成纤维细胞、水凝胶驻留巨噬细胞和循环巨噬细胞以及内皮细胞将通过 纳米多孔 SiM (~60 nm)，而微孔 SiM (~ 8 µm) 将允许循环液外渗 TGF-β刺激下的巨噬细胞和水凝胶的浸润。为了实现以患者为中心的芯片， 肌腱成纤维细胞将用于创建肌腱水凝胶并重新编程与供体匹配的 iPSC 以衍生 分别为内皮细胞和巨噬细胞。另一项创新将是标签的集成 将光子传感器免费插入微流体装置中以实现片上传感，这长期以来一直被认为是 对片上器官设备的关键、未满足的需求。 UG3研究将使用该芯片来验证 mTOR 在疾病模型中的应用并识别生物学相关的生物标志物。在UH3阶段，我们将利用 芯片作为临床前试验平台，用于测试 FDA 批准的 mTOR 抑制剂的功效和安全性 疾病缓解药物，并作为药物筛选平台来识别和优先考虑更安全、更有效的药物 mTOR 抑制剂和肌腱损伤衰老的临床试验。为顺利完成这一创新 项目中，我们组建了一支由肌腱组织工程和外科领域卓有成效的专家组成的团队， 免疫学、iPSC 技术、GMP 细胞制造、纳米和微米制造、传感器技术以及 高通量筛选。拟议的研究将开发一种人体微生理系统来催化 临床试验并加速治疗急性和慢性肌腱损伤的药物发现。