微观力学刚度对脂肪干细胞促进皮瓣血管化的调控机制研究

Skin flap necrosis is always clinical problematic in wound repairing. Adipose derived stem cells (ADSCs) local therapy is one of the most promising approach for improving injured tissue vascularization on clinic. However, an ischemic microenvironment results in ADSCs apoptosis, which weakens the therapeutic effect of promoting vascularization. Studies have shown that the microenvironment created by the medical hydrogel for ADSCs in vivo is beneficial to the survival of ADSCs, and the mechanical stiffness of the matrix in the hydrogel microenvironment plays an important role in the ADSCs promoting vascularization. However, the mechanism of how the mechanical parameters and chemical signals in microenvironment regulate of ADSCs promoting vascularization is not clear. Therefore, the aim of this project is to construct a biodegradable medical hydrogel with adjustable mechanical stiffness, which can envelop ADSCs and improve the survival rate of ADSCs in vivo. Then different substrate mechanical stiffness of hydrogels regulating the effect of vascular endothelial growth factor (VEGF) on the role of ADSCs promoting vascularization will be systematically investigated. Thus, the hydrogel with the best mechanical stiffness for ADSCs promoting vascularization will be obtained. Furthermore, molecular mechanisms of mechanical stiffness regulating the effect of VEGF on the role of ADSCs promoting vascularization will be explored by “matrix stiffness -integrin-VEGFR-cell function”. Finally, the effect of the different matrix mechanical stiffness of the hydrogel to improve the ADSCs promoting vascularization will be evaluated by ischemic skin flap model.

皮瓣缺血坏死是困扰创面修复的一个临床难题。脂肪干细胞（ADSCs）移植是促进损伤组织血管化最具前景的方法之一。但是，缺血微环境会导致移植ADSCs凋亡，削弱了其促血管化作用。研究表明医用水凝胶创建的ADSCs体内移植微环境有利于其存活，其中水凝胶微环境中基质的力学刚度对ADSCs促血管化作用有重要影响。然而，微环境中基质力学参数和化学信号对力学刚度调控ADSCs促血管化的作用机制尚不明确。因此，本项目拟构建力学刚度可调的可降解医用水凝胶，并实现对干细胞装载和提高体内存活率，系统研究水凝胶基质的不同力学刚度调控血管内皮生长因子（VEGF）对ADSCs促血管化的影响，得到促血管化力学刚度最佳的水凝胶体系，并从“基质刚度-整合素-VEGFR-细胞功能”出发，探究基质刚度对VEGF作用ADSCs促血管化功能调控的机制。最后通过缺血皮瓣模型，对不同力学刚度水凝胶提高ADSCs促血管化疗效进行评价。

皮瓣移植是整复外科常用的修复手段，但缺血坏死是临床上面临的主要问题。脂肪干细胞（ADSCs）因为其具有促血管化功能，近年来被广泛应用于细胞治疗、组织工程和再生医学。但是，缺血微环境会导致移植ADSCs凋亡，削弱了其促血管化作用。研究表明医用水凝胶创建的ADSCs体内移植微环境有利于其存活，其中水凝胶微环境中基质的力学刚度对ADSCs促血管化作用有重要影响。然而，微环境中基质力学参数和化学信号对力学刚度调控ADSCs促血管化的作用机制尚不明确。本项目首先比较了微环境的基质刚度对ADSCs功能的影响，发现低刚度(3.24±0.18Pa)的细胞外基质可以促进ADCSs的促血管化功能，而这一调控过程由整合素参与介导，通过“细胞外基质刚度-整合素-Src-VEGFR-2-ERK-VEGF”通路调控ADSCs促血管化功能。其次通过在明胶高分子中引入长链的、可光交联的聚乙二醇(PEG）丁二酰亚胺羧基甲酯（ACLT-PEG2000-SCM），可以合成软弹性的聚乙二醇明胶（PEG-Gel），从而使材料交联后形成疏松的网络结构，提升其柔软度，合成超柔软水凝胶。超柔软水凝胶可以为通过力学微环境，通过“细胞外基质力学刚度-整合素-VEGFR”调节ADSCs的促血管化过程。最后，本项目通过动物体内实验进一步验证ADSC于力学刚度为3.24±0.18Pa的PEG-Gel水凝胶促进血管形成和组织修复的能力。本研究为ADSCs体内植入提供适宜生长、迁移且具有提高其促血管化功能的微环境，进而为ADSCs在皮瓣缺血修复过程中的应用提供新的策略。