Development of a Collagen-based 3D Bioprinted Microfluidic Platform for Vascular Tissue Engineering and Disease Modeling

项目摘要/摘要 高血压血管疾病是导致发病率和死亡率的主要全球风险因素，影响超过116人。 百万人。其特点是细胞外基质(ECM)成分、生物力学 性质，以及导致血压升高和血管僵硬的分子信号异常。广泛性 工作的重点是开发改进的药物治疗方法、组织工程血管和 使工程化的体积组织血管化，但往往忽视了细胞信号之间的相互依赖 以及导致疾病的生物力学力量。认识细胞外基质生物力学与生物力学的关系 受体信号和开发操纵它的工具对于创造健康、成熟的血管是必不可少的。 组织，同时避免病变。在这里，我建议合并空间定义ECM 以血管为灵感的3D生物打印组织支架的组成和细胞排列将产生 工程化的小动脉，从生理上控制血管张力，并促进对ECM如何 成分和受体运输的改变会损害血管的反应性，并促进高血压的表型。我 将利用两个新的平台：Fresh 3D生物打印直接从ECM制造可灌流的血管 蛋白质，以及基于荧光的纳米机械生物传感器(NMBS)，用于在体内定位组织应变和 目标1：开发基于胶原蛋白的三维生物印迹血管微流控材料 集成可控流体流动和内皮化的平台复制血管内皮细胞生物力学和 内皮屏障功能；目标2：使用FRESH直接构建ECM结构和细胞组织 逐层打印以概括阻力动脉、血管平滑肌细胞和内皮细胞 细胞功能；以及目标3：研究细胞外基质的病理变化如何改变受体的运输和损伤 使用新型生物墨水复制健康和高血压血管ECM组合物的血管反应性 材料特性。这项建议旨在加强我们对生物力学与生物力学之间相互作用的认识。 在血管发育和高血压疾病进展过程中强制生化信号。这个 微流体(K99)和工程血管组织(R00)将在药物筛选和 疾病模型。职业发展计划，在共同导师范伯格博士和克莱曼博士的指导下， 和我的顾问委员会，将提供微流体、生物力学分析和血管方面的高级培训 生物学/疾病建模。指导阶段利用一个跨学科的指导团队，并 卡内基梅隆大学、卡内基梅隆大学 匹兹堡和血管医学研究所。这款K99/R00，结合我之前在3D方面的专业知识 生物打印、先进的荧光显微镜、定量图像分析和细胞/分子生物学将 促进我过渡到专注于ECM组成和结构如何改变受体的独立职业生涯 驱动组织成熟和疾病进展的信号。

Project Summary/Abstract Hypertensive vascular disease is a leading global risk factor for morbidity and mortality, affecting over 116 million people. It is characterized by alterations to extracellular matrix (ECM) composition, biomechanical properties, and aberrant molecular signaling leading to increased blood pressure and vessel stiffening. Extensive work has focused on developing improved pharmacological treatments, tissue engineered blood vessels, and vascularizing engineered volumetric tissue, but often overlook the interdependence between cellular signaling and biomechanical forces leading to disease. Understanding the relationship between ECM biomechanics and receptor signaling, and developing tools to manipulate it, are essential for creating a healthy, mature vascular tissue while avoiding pathological changes. Here I propose that incorporation of spatially defined ECM composition and cellular alignment into a vascular-inspired 3D bioprinted tissue scaffold will produce an engineered small artery that physiologically controls vascular tone and facilitates investigation into how ECM composition and altered receptor trafficking impair vascular reactivity and promote a hypertensive phenotype. I will utilize two novel platforms: FRESH 3D bioprinting to directly fabricate perfusable vasculature from ECM proteins, and a fluorescence-based nanomechanical biosensor (NMBS) for mapping in vivo tissue strain and vascular smooth muscle contractility to Aim 1: Develop a collagen-based 3D bioprinted vascular microfluidic platform integrating controlled fluid flow and endothelialization to replicate vascular ECM biomechanics and endothelial barrier function; Aim 2: Directly pattern ECM structure and cellular organization using FRESH printing in a layer-by-layer manner to recapitulate resistance artery vascular smooth muscle cell and endothelial cell function; and Aim 3:Investigate how pathologic changes in the ECM alters receptor trafficking and impairs vascular reactivity using novel bioinks to replicate a healthy and hypertensive vessel ECM composition and material properties. This proposal seeks to enhance our knowledge of the interplay between biomechanical forces biochemical signaling during vascular development and hypertensive disease progression. The microfluidics (K99) and engineered vascular tissues (R00) created will have wide utility for drug screening and disease modeling. The career development plan, under the guidance of co-mentors Drs. Feinberg and Kleyman, and my advisory committee, will provide advanced training in microfluidics, biomechanical analysis, and vascular biology/disease modeling. The mentored phase capitalizes on an inter-disciplinary mentoring team and substantial research and professional development resources at Carnegie Mellon University, the University of Pittsburgh, and the Vascular Medicine Institute. This K99/R00, combined with my prior expertise in 3D bioprinting, advanced fluorescence microscopy, quantitative image analysis, and cellular/molecular biology, will facilitate my transition to an independent career focused on how ECM composition and structure alter receptor signaling to drive tissue maturation and disease progression.