Readily Available Stem Cell-Based Vascular Grafts for Emergent Surgical Care

用于紧急手术护理的现成干细胞血管移植物

• 批准号: 10630420
• 负责人: Yibing Qyang
• 金额: $ 6.38万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10630420
• 关键词: Address; Age; Autologous; Behavior; Bioreactors; Caliber; Caring; Cell Line; Cell physiology; Cells; Data; Development; Disease; Due Process; Endothelial Cells; Endothelium; Genes; Goals; Human; Immunocompromised Host; Implant; Modeling; Operative Surgical Procedures; Patients; Polyglycolic Acid; Procedures; Radioisotopes; Rattus; Research; Research Personnel; SLC5A5 gene; Smooth Muscle Myocytes; Technology Assessment; Thrombosis; Time; Training; Validation; Vascular Graft; Vascular Smooth Muscle; Visualization; Work; X-Ray Computed Tomography; base; endothelial stem cell; health assessment; implantation; in vivo; induced pluripotent stem cell; injury and repair; innovative technologies; parent grant; prevent; repaired; shear stress; single photon emission computed tomography; solute; stem cell derived tissues; stem cells; technology development; time use; tissue stem cells; transcription activator-like effector nucleases; uptake; vascular injury; vascular tissue engineering

Project Summary Tissue engineered vascular grafts (TEVGs) produced through seeding human induced pluripotent stem cell (hiPSC)-derived vascular smooth muscle cells (VSMCs) onto polyglycolic acid (PGA) meshes have been developed to repair patients’ vascular injuries. These hiPSC- TEVGs can be made further applicable through decellularization, allowing them to be stored for long periods and used off the shelf. For large diameter vessels (>6mm inner diameter), these decellularized TEVGs can be implanted directly, with host cells sufficiently able to recellularized the graft over time. However, small diameter TEVGs (2-4mm) are prone to thrombosis if implanted directly, requiring a layer of endothelial cells (ECs) to be coated in the lumen before these procedures. Significant research has been done to create hiPSC-ECs sufficient for endothelialization of small diameter TEVGs, as many patients have autologous ECs unable to be used for this process due to age or disease. For this reason, it is vital for the development of this technology for researchers to know how these coated hiPSC-ECs behave in vivo non- invasively. In this supplemental proposal, this need will be addressed by producing a stable hiPSC line expressing human sodium iodide symporter (hNIS), a solute carrier that confers the ability to uptake radioisotopes into the cell. These hNIS-hiPSCs will be used to derive ECs that can be tracked in vivo non-invasively using SPECT/CT imaging, providing key data to the behavior and function of these cells after implantation. This will be accomplished through the use of transcription activator-like effector nuclease (TALEN) gene editing to insert a constitutively active hNIS cassette into the AAVS1 “safe harbor” locus in stable hiPSCs. hiPSC- TEVGs will then be decellularized and coated with hNIS-ECs derived from the hNIS-hiPSCs, and these grafts will be trained for implantation by undergoing shear stress in a flow bioreactor, to enhance hNIS-EC maturity. These matured hNIS-EC TEVGs will then be used as a aortic interposition grafts in an immunocompromised rat model, which allow for the validation of the functionality of these hNIS-ECs to allow in vivo visualization over time using SPECT/CT imaging. This innovative technology will allow for non-invasive assessment of the health and extent of implanted hiPSC-EC coating in decellularized grafts, providing vital information into the ability of the endothelium to prevent thrombosis in patients who require vascular injury repair.

项目摘要 组织工程化血管移植物（TEVG）通过接种人诱导产生 多能干细胞（hiPSC）衍生的血管平滑肌细胞（VSMC）在聚乙醇酸 已经开发了酸性（PGA）网状物来修复患者的血管损伤。这些hiPSC- TEVG可以通过脱细胞化进一步适用，允许它们储存用于 长时间使用，并使用现成的。对于大直径血管（内径> 6 mm），这些 脱细胞的TEVG可以直接植入，宿主细胞能够充分地再细胞化， 随着时间的推移，然而，小直径TEVG（2- 4 mm）易于血栓形成，如果 直接植入，需要在管腔中涂覆一层内皮细胞（EC）， 这些程序。已经进行了大量的研究以产生足以用于以下目的的hiPSC-EC： 小直径TEVG的内皮化，因为许多患者的自体EC不能 由于年龄或疾病而用于此过程。因此，它对发展至关重要。 这项技术可以让研究人员了解这些包被的hiPSC-EC在体内的行为， 侵入性地在本补充建议中，将通过建立稳定的 表达人钠碘同向转运体（hNIS）的hiPSC系，hNIS是一种溶质载体， 放射性同位素进入细胞的能力。这些hNIS-hiPSC将用于衍生EC， 可以使用SPECT/CT成像在体内非侵入性地进行跟踪， 植入后这些细胞的行为和功能。这将通过以下方式实现： 使用转录激活因子样效应物核酸酶（TALEN）基因编辑来插入 在稳定的hiPSC中，将组成型活性hNIS盒插入AAVS 1“安全港”基因座。hiPSC- 然后将TEVG脱细胞并用衍生自hNIS-hiPSC的hNIS-EC包被， 并且这些移植物将通过在流动生物反应器中经受剪切应力而被训练用于植入， 以增强hNIS-EC成熟度。然后，这些成熟的hNIS-EC TEVG将被用作主动脉瘤。 在免疫功能低下的大鼠模型中插入移植物，这允许验证 这些hNIS-EC允许使用SPECT/CT随时间体内可视化的功能 显像这项创新技术将允许对健康状况进行非侵入性评估， 在脱细胞移植物中植入hiPSC-EC涂层的程度，为研究hiPSC-EC涂层的生物学特性提供了重要信息。 在需要血管损伤修复的患者中，内皮防止血栓形成的能力。