Regulating the Quality and Potency of Stem Cells with Biophysical Cues from Dynamic Nanofibrous Hydrogels for Therapeutic Purposes

利用动态纳米纤维水凝胶的生物物理线索调节干细胞的质量和效力用于治疗目的

• 批准号: 10724060
• 负责人: David Castilla-Casadiego
• 金额: $ 12.48万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10724060
• 关键词: 3-Dimensional; Address; Adhesives; Adipocytes; Affect; Allogenic; Behavior; Biocompatible Materials; Biophysics; Bioreactors; Cell Count; Cell Culture System; Cell Proliferation; Cell Survival; Cell physiology; Cell surface; Cells; Characteristics; Chemicals; Chondroblast; Clinical effectiveness; Coculture Techniques; Collagen; Collagen Type I; Cues; Diameter; Disease; Electrospinning; Encapsulated; Engineering; Ensure; Extracellular Matrix; Fiber; Future; Goals; Human; Human body; Hyaluronic Acid; Hydrazones; Hydrogels; In Vitro; Individual; Integrin Binding; Length; Link; Lymphocyte Suppression; Measures; Mechanics; Memory; Mentors; Mesenchymal Stem Cells; Methods; Molecular; Morphology; Nanotopography; Osteoblasts; Phase; Phenotype; Play; Production; Proliferating; Property; Proteins; Relaxation; Reporting; Reproducibility; Research; Research Proposals; Role; Rosales; Secretory Cell; Signal Pathway; Source; Stress; Structure; Supervision; Surface; System; Testing; Therapeutic; Tissues; Transcriptional Activation; angiogenesis; biophysical properties; crosslink; cytokine; density; design; immunoregulation; improved; in vivo; insight; manufacture; manufacturing scale-up; mechanical properties; mechanical signal; nanofiber; patient population; scaffold; sensor; stem cell expansion; stem cells; three-dimensional modeling; transcription factor; viscoelasticity

Project Summary/Abstract Human mesenchymal stem cells (hMSCs) are considered a source for allogeneic therapies to treat diverse diseases. Due to the exponential increase in demand, there is a need for new strategies to produce potent hMSCs to serve diverse patient populations. Currently, conventional planar culture and bioreactors are used as scale-up manufacturing methods. However, these are not specifically tailored for hMSCs expansion. They may alter the cell phenotype and secretome, affecting clinical effectiveness. Further studies to understand the role of substrate mechanics on hMSC expansion are required to achieve reproducible production. Numerous scaffolding alternatives replicate several characteristics of the native extracellular matrix (ECM). However, its dynamic mechanics, which plays a fundamental role in regulating crucial cellular processes, has not been amply studied yet. Furthermore, most in-vitro substrates are static and supraphysiologically stiff. Static substrates have offered a substantial benefit for generating high cell numbers; however, hMSCs have been shown to retain mechanical information, limiting therapeutic capabilities. To address this problem, this proposed research seeks to investigate the role of dynamic cell-matrix interactions and nano-topographical cues on the immunomodulatory potential of hMSCs using a composite of electrospun-fibers encapsulated in a dynamic hydrogel, with the hypothesis that this composite biomaterial will promote high hMSCs production with relevant therapeutic value, while eliminating the limitations reported for the conventional cell culture systems. The K99 period will focus on engineering and characterizing the dynamic nanofibrous hydrogel composites to propel me toward establishing the mechanisms by which they modulate cell quality and potency attributes with relevant therapeutic value (during the R00 phase). In Aim 1, we will develop the dynamic nanofibrous system using a hyaluronic acid hydrogel network crosslinked via dynamic covalent hydrazone bonds that capture the viscoelasticity of ECM in tissues. Four variables, including the encapsulation of the electrospun collagen nanofibers at various densities, fiber diameter, fiber length, and the stress relaxation timescale of the hydrogel will be characterized in this aim to promote hMSC viability and proliferation. In Aim 2, hMSCs cell quality and potency will be assessed by measuring the effect of hydrogel parameters on cellular secretory activity. Immunomodulatory properties will be evaluated by quantifying lymphocyte suppression in co-culture, as well as expression of hMSC surface markers. The capacity of the hMSCs to differentiate will also be assessed. In aim 3, the mechanism linking the biophysical parameters of the nanofibrous hydrogel to hMSC secretory activity will be probed by examining cell adhesive proteins and the activation of transcription factors or sensors of mechanical cues. In sum, the proposed research will lead to new insights to produce hMSCs with high therapeutic value, which will enable new culture substrates that achieve control in reproducibility and cell quality to serve diverse patient populations.

项目概要/摘要 人类间充质干细胞 (hMSC) 被认为是同种异体疗法的来源 多种疾病。由于需求呈指数级增长，需要新的生产策略 有效的 hMSC 可以服务于不同的患者群体。目前，传统的平面培养和生物反应器 用作放大生产方法。然而，这些并不是专门为 hMSC 扩增而设计的。 它们可能会改变细胞表型和分泌组，影响临床效果。进一步研究以了解 基质力学对 hMSC 扩增的作用是实现可重复生产所必需的。很多的 支架替代品复制了天然细胞外基质（ECM）的几个特征。然而，其 动态力学在调节关键细胞过程中发挥着基础作用，但尚未得到充分的研究 还研究过。此外，大多数体外基质是静态的且超生理刚性的。静态基板有 为产生大量细胞提供了巨大的好处；然而，hMSCs 已被证明可以保留 机械信息，限制治疗能力。为了解决这个问题，本项研究旨在 研究动态细胞-基质相互作用和纳米拓扑线索对免疫调节的作用 使用封装在动态水凝胶中的电纺纤维复合材料的 hMSC 的潜力， 假设这种复合生物材料将促进 hMSCs 的高产量并具有相关的治疗价值， 同时消除了传统细胞培养系统的局限性。 K99时期将重点关注 设计和表征动态纳米纤维水凝胶复合材料，推动我建立 它们调节具有相关治疗价值的细胞质量和效力属性的机制 （在 R00 阶段）。在目标 1 中，我们将使用透明质酸开发动态纳米纤维系统 水凝胶网络通过动态共价腙键交联，捕获 ECM 的粘弹性 组织。四个变量，包括不同密度的电纺胶原纳米纤维的封装， 该目标将表征水凝胶的纤维直径、纤维长度和应力松弛时间尺度 促进 hMSC 活力和增殖。在目标 2 中，hMSC 细胞质量和效力将通过以下方式评估： 测量水凝胶参数对细胞分泌活性的影响。免疫调节特性将是 通过量化共培养中的淋巴细胞抑制以及 hMSC 表面标志物的表达来评估。 hMSC 的分化能力也将被评估。在目标 3 中，连接生物物理的机制 纳米纤维水凝胶对 hMSC 分泌活性的参数将通过检查细胞粘附力来探测 蛋白质和转录因子或机械线索传感器的激活。总之，拟议的研究 将带来生产具有高治疗价值的 hMSC 的新见解，从而使新的培养基质成为可能 实现再现性和细胞质量的控制，为不同的患者群体提供服务。