力介导基质金属蛋白酶降解细胞外基质的物理机制研究

Understanding how cells respond and adapt to the 3D microenvironment provide by biomaterials is the key for tissue regeneration. Despite extensive studies show that the mechanical properties of biomaterials can effectively regulate many cellular processes, including cell migration, proliferation and differentiation, little is known about how cells actively respond and modulate the mechanical properties of the microenvironment to maintain tissue-level structural integrity and functionality. To regulate the mechanical properties of extracellular matrix (ECM), cells secrete various matrix metalloproteinases (MMPs) which can hydrolyze special protein substrates and cause the degradation of ECM. This process would be influenced by both the mechanical stress of the ECM network and the forces directly exerted by resident cells inevitably. Here we employ atomic force microscopy (AFM) and magnetic tweezers-based single-molecule force spectroscopy to investigate how the hydrolysis of protein substrates by MMPs is regulated by forces under different physiological and pathological environments. Our studies aim to provide a quantitative physical picture underlying the dynamic process of matrix hydrolysis in different mechano-chemical environments. Furthermore, the findings at the single molecule level will be corroborated by directly studying the effects of external force on the degradation of model hydrogels containing MMP hydrolysable cross-linkers. Our study would lay a foundation to understand many physiological or pathological processes involving mechanical homeostasis, such as cell proliferation, tissue regeneration and wound healing. Meanwhile, our findings can inspire the design of novel hydrogels with tailored mechanical properties for many biomedical applications, such as drug delivery and tissue engineering.

生物材料与组织细胞相互作用形成的三维微环境是连接生物材料与组织再生的纽带。虽然目前材料的力学特性对细胞的迁移、增殖和分化等不同生理功能的影响已被广泛研究，但对细胞如何主动调节三维微环境的力学特性从而实现组织再生方面的研究还缺乏完备的技术和完善的思路。细胞可分泌多种基质金属蛋白酶，通过降解特定底物来改变微环境，而此过程必然会受到基质本身的力学特性和细胞对基质所施加的作用力的影响。在本项目中，我们将发展基于原子力显微镜和磁镊的单分子测量方法，深入研究在不同生理、病理环境下，基质金属蛋白酶对底物水解受力调控的物理机制，定量刻画基质水解在不同化学、力学微环境下的动力学过程，并通过仿生水凝胶材料的制备及宏观力学测量，验证单分子实验结论。我们的研究将为进一步解释细胞增殖、组织重构、伤口愈合等生理或病理过程打下基础，同时也为设计和制备具有特殊力学性能水凝胶材料及其在载药、组织工程等生物医学应用提供支持。

在细胞微环境中，细胞能感受力信号并将其转变为生物化学信号，引起胞内力学信号的响应过程，从而实现细胞迁移、增殖、分化等不同的生理功能，研究力在不同生理或病理环境下对细胞行为的调控，对细胞培养、组织工程和器官再生具有重要意义，也是目前生物物理研究的重点问题。在本项目中，我们围绕力信号调控分子动力学过程的物理机制开展研究，在基金的支持下 共发表论文8篇，并有数篇在投，申请发明专利2项，培养博士研究生2名，硕士研究生2名。主要成果包括：1）在单分子力谱技术方法上进行创新，设计了可进行高精度单分子测量的分子结构；同时设计了基于组氨酸磷酸化的新型高稳定的共价结合方法，有效地提高了单分子力谱实验的效率。2）通过基于原子力显微镜的单分子力谱技术，在单分子层面对金属蛋白酶降解细胞外基质、含酯键蛋白折叠与解折叠、含阳离子-π相互作用等体系的分子机制和物理机理进行探究，为深入理解细胞行为提供了重要参考。3）在生物材料方面，提出了一套基于蛋白质折叠与解折叠动力学特性的生物材料设计方案，并设计出抗疲劳性质的水凝胶材料。通过引入酶降解机制，设计制备出可受力信号调控的可降解水凝胶材料。这些研究揭示了生物材料的宏观力学特性与微观的分子间相互作用/分子动力学特性之间的关系，进一步深化了人们对生物材料力学构造的认识，为设计特殊力学性能的生物材料提供了思路。