Tailoring of cellular mechanical microenvironments to rescue age-related impairments in bone regeneration

衰老损害骨细胞对机械负荷的反应性和骨的成功 受伤后的愈合。越来越多的证据表明，这两种现象是相关的： 存在于细胞微环境中的机械信号（“机械微环境”）调节 骨愈合的许多方面，包括骨髓的募集和成骨细胞分化 基质细胞（MSCs）。重要的是，多组研究人员的研究表明， 细胞保持机械敏感性，但需要更高的刺激强度。因此，这一重点 该项目是在骨愈合过程中直接操纵机械微环境，通过 定制增材制造（AM）、骨诱导、生物陶瓷 支架，以确定细胞对局部机械线索的反应如何随年龄而变化。的 这项工作假设是通过先进的AM来剪裁细胞机械微环境， 支架设计可以挽救骨再生中与年龄相关的损伤。这项工作建立在 初步数据表明，使用创新的AM方法打印机械坚固的陶瓷 具有高孔隙率（>80%）和大到足以 促进体内血管形成和骨生成。在目标1中，我们将比较 年轻（12周龄）vs.成熟（77周龄）vs.老龄（104周龄）鼠MSC与 体外机械加载支架内的机械微环境。当地的机械线索 将使用有限元建模和机械测试来确定这些支架中存在的量，并且 使用新的技术组合评估细胞反应，包括空间评估 在单细胞水平上的成骨进展。目标2也将利用这些分析工具， 并将比较骨再生反应与内定制的机械微环境， 植入年轻和老年小鼠体内的支架。这两个目标共同解决了一个高度临床化的领域， 需要并将填补有关骨机械年龄相关变化的知识方面的关键空白， 治疗期间的反应。这项工作的成果将为新一代奠定基础 骨修复技术，可以适应细胞行为的变化， 细胞机械敏感性，以促进成骨分化、骨组织形成，并最终， 损伤后骨功能的恢复。

Aging impairs both the responsiveness of bone cells to mechanical loading and the success of bone healing following injury. Growing evidence indicates that these two phenomena are related: local mechanical cues present in the cellular microenvironment (“mechanical microenvironment”) regulate numerous aspects of bone healing, including recruitment and osteoblastic differentiation of marrow stromal cells (MSCs). Importantly, studies by multiple groups of investigators have indicated that aged cells remain mechanosensitive, but require higher stimulus magnitudes. Therefore, the focus of this project is on directly manipulating the mechanical microenvironment during bone healing, via customization of the architecture of additively manufactured (AM), osteoinductive, bioceramic scaffolds, in order to identify how the cellular responses to local mechanical cues differ with age. The hypothesis of this work is that tailoring of cellular mechanical microenvironments through advanced AM scaffold design can rescue age-related impairments in bone regeneration. This work builds on preliminary data demonstrating use of an innovative AM method to print mechanically robust ceramic scaffolds of exceptionally tunable architectures with high porosity (>80%) and pore sizes large enough to facilitate vascularization and osteogenesis in vivo. In Aim 1, we will compare the osteogenic responses of young (12-week-old) vs. mature (77-week-old) vs. aged (104-week-old) murine MSCs to the mechanical microenvironment within mechanically loaded scaffolds in vitro. The local mechanical cues present in these scaffolds will be determined using finite element modeling and mechanical testing, and the cellular responses evaluated using a novel combination of techniques including spatial assessment of the progression of osteogenesis at the single-cell level. Aim 2 will leverage these analytical tools as well, and will compare the bone regeneration responses to tailored mechanical microenvironments within scaffolds implanted in young and aged mice. Together, these two aims address an area of high clinical need and will fill critical gaps in knowledge regarding age-related changes in bone mechano- responsiveness during healing. The outcomes of this work will lay the foundation for a new generation of bone repair technologies that can accommodate alterations in cell behavior with aging and harness cell mechano-sensitivity to promote osteogenic differentiation, bone tissue formation, and ultimately, restoration of bone function following injury.