Microphysiological Models to Evaluate the Role of Age-Dependent Fibrinogen Sialylation in Wound Healing

伤口愈合是一个复杂的过程。伤口愈合的第一步是凝血。形成的凝块可以止血，并为细胞重建受损组织提供支持。婴儿和成人的伤口愈合也不同，但这些差异还没有得到很好的理解。该项目将开发新的工具，以更好地了解(1)婴儿和成人之间凝血的差异；(2)凝血的差异如何导致伤口愈合的差异。许多类型的细胞参与愈合，包括血小板和成纤维细胞。血小板凝血中扮演重要的角色,出血停止后成纤维细胞帮助重建受损组织。该项目还将测量成纤维细胞和血小板在伤口愈合过程中的相互作用。更好地了解伤口愈合有望为美国近600万患有凝血功能受损和未愈合伤口的人找到新的治疗靶点。该项目的教育部分包括培训工程生物医学系统的本科生和研究生，包括与临床医生合作。外联活动包括开发实践教育模块和活动，这些模块和活动将为K-12科学竞赛提供创新，并为兽医学院年度开放日和北卡罗来纳科学研讨会构建演示工具包，该活动吸引了该地区的K-12学生。伤口愈合涉及复杂的生化和机械信号的协调，指导多种细胞类型的行为。由于伤口愈合的复杂性，关于损伤愈合的机制仍然未知。受伤后，纤维蛋白凝块形成，作为支持组织修复的浸润细胞的临时支架，止血并促进愈合。纤维蛋白基质缺乏可导致伤口不愈合。虽然已知衰老会影响愈合，但尚不清楚与年龄相关的纤维蛋白特性差异如何影响愈合结果。最近的研究已经确定了成人和新生儿之间纤维蛋白网络特性的广泛差异。例如，与成人纤维蛋白相比，成纤维细胞在新生儿纤维蛋白上的附着和迁移更好，与成人纤维蛋白支架相比，新生儿纤维蛋白支架在小鼠全层损伤模型中促进了更好的体内愈合。虽然翻译后修饰的许多差异可能导致了纤维蛋白原功能的年龄相关差异，但最近的数据强调，新生儿纤维蛋白原中唾液酸（Sia）含量的增加显著导致了凝块结构、力学、聚合和降解动力学的年龄相关差异。这项工作的主要假设是，与成人纤维蛋白原相比，新生儿纤维蛋白原中Sia含量的增加导致纤维蛋白聚合机制、凝块结构和机械性能的改变，从而改善了愈合结果。该项目将研究新生儿和成人纤维蛋白原网络中Sia含量+/- Sia如何影响成人和新生儿成纤维细胞的体外反应。接下来，本项目将研究Sia含量如何影响伤口愈合过程中的细胞串扰。创新的组织工程技术和微流体技术将用于开发微生理系统（MPS）来研究血小板和成纤维细胞在伤口愈合过程中的相互作用。MPS提供了一种方法来生成可操作的，可翻译的数据，通过概述选定的人体组织功能，必要的原始人类细胞类型，显微解剖学，细胞-细胞相互作用和微机械线索。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Wound healing is a complex process. The first step in wound healing is clotting. The clot that forms stops bleeding and provides support for cells to rebuild damaged tissue. Wound healing is also different between babies and adults, but these differences are not well understood. This project will develop new tools to better understand (1) how clotting is different between babies and adults and (2) how differences in clotting lead to differences in wound healing. Many cells types are involved in healing, including platelets and fibroblasts. Platelets play important roles in clotting and fibroblasts help rebuild damaged tissue after bleeding stops. The project will also measure interactions between fibroblasts and platelets in wound healing. Better understanding of wound healing is expected to lead to identification of novel therapeutic targets for the nearly 6 million people in the United States suffering from impaired coagulation and non-healing wounds. The project's education component involves training undergraduate and graduate students in engineering biomedical systems, including working with clinicians. Outreach activities include developing hands-on educational modules and activities that will be innovated for K-12 science fair competitions and constructing a demonstration kit for the Annual College of Veterinary Medicine Open House Day and the North Carolina Science Café, which engages K-12 students in the region. Wound healing involves the complex orchestration of biochemical and mechanical signals directing behavior of multiple cell types. Due to the complex nature of wound healing, much remains unknown about mechanisms underlying impaired healing. After injury, a fibrin clot is formed that stops bleeding and promotes healing by serving as a provisional scaffold for infiltrating cells that support tissue repair. A deficient fibrin matrix can result in non-healing wounds. While it is known that aging influences healing, it is unknown how age related differences in fibrin properties may influence healing outcomes. Recent studies have identified extensive differences in fibrin network properties between adults and neonates. For example, fibroblasts attach and migrate better on neonatal fibrin compared to adult fibrin, and neonatal fibrin scaffolds promote better healing in vivo in a murine full thickness injury model compared to adult fibrin scaffolds. While many variances in post-translational modifications likely contribute to age-related differences in fibrinogen function, recent data highlights that increased sialic acid (Sia) content in neonatal fibrinogen contributes significantly to age-related differences in clot structure, mechanics, and polymerization and degradation dynamics. The overarching hypothesis of this work is that increased Sia content in neonatal fibrinogen results in altered fibrin polymerization mechanisms, clot structure, and mechanical properties compared to adult fibrinogen, which drives improved healing outcomes. This project will investigate how Sia content in neonatal and adult fibrinogen networks +/- Sia influence adult and neonatal fibroblast responses in vitro. Next, this project will investigate how Sia content influences cellular crosstalk in wound healing. Innovative tissue engineering techniques and microfluidics will be employed to develop a microphysiological system (MPS) to study the interactions of platelets and fibroblasts during wound healing. MPS provide a means to generate actionable, translatable data by recapitulating select human tissue functions with the requisite primary human cell types, microanatomy, cell-cell interactions and micromechanical cues.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.