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.

项目总结/摘要 人类间充质干细胞（hMSC）被认为是同种异体疗法的来源， 各种疾病。由于需求呈指数级增长，需要新的战略来生产 强大的hMSC服务于不同的患者群体。目前，常规的平面培养和生物反应器是 用作放大制造方法。然而，这些并不是专门为hMSC扩增而定制的。 它们可能改变细胞表型和分泌组，影响临床疗效。进一步研究，了解 基质力学对hMSC扩增作用是实现可再现生产所必需的。许多 支架替代物复制了天然细胞外基质（ECM）的几个特征。但其 在调节关键细胞过程中起着基本作用的动力学机制， 研究过了。此外，大多数体外基质是静态的和超生理刚性的。静态基质具有 提供了产生高细胞数量的实质性益处;然而，hMSC已被证明保留了 机械信息，限制治疗能力。为了解决这个问题，这项研究旨在 研究动态细胞-基质相互作用和纳米形貌线索对免疫调节的作用， 使用包封在动态水凝胶中的电纺纤维的复合物， 假设这种复合生物材料将促进具有相关治疗价值的hMSCs的高产量， 同时消除了常规细胞培养系统所报道的局限性。K99期间将重点关注 设计和表征动态纳米纤维水凝胶复合材料，以推动我建立 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R 00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 通过动态共价腙键交联的水凝胶网络， 组织中四个变量，包括以各种密度包封电纺胶原纳米纤维， 纤维直径、纤维长度和水凝胶的应力松弛时间尺度将在该目的中表征 以促进hMSC活力和增殖。在目标2中，将通过以下方法评估hMSC细胞质量和效力： 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。 还将评估hMSC分化的能力。在目标3中，将生物物理和生物技术 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来探测 蛋白质和转录因子或机械信号传感器的激活。总之，拟议的研究 将导致新的见解，以产生具有高治疗价值的hMSCs，这将使新的培养基质 其实现了对再现性和细胞质量的控制以服务于不同的患者群体。