缓释exosomes的抗菌生物玻璃基仿生支架的构建及其促糖尿病创面修复的研究

The repair of diabetic wounds still faces great challenges in clinic due to infection, poor vascularization and long healing period. At present, wound dressings are mostly inactive, which cannot meet the needs of the patients. We previously found that bioactive glass nanoparticles (BGN) and its composites can induce rapid vascularization and healing of the wound. This project proposes to fabricate a structure-mimic bilayer hierarchical nanofibrous scaffold with anti-bacterial and angiogenic property to facilitate tissue regeneration and diabetic cutaneous wound healing. The biomimetic bilayer nanofibrous scaffold will be composed of collagen modified copper-containing bioactive glass nanoparticles (cCu-BGN), collagen (Col) and polycaprolactone (PCL), and adipose stem cells derived exosomes (Exo), whose upper layer consists of aligned cCu-BGN/Col/PCL (CCP) nanofibers (mimicking the ordered epidermal structure), and sublayer layer consists of random Exo/cCu-BGN/Col/PCL (Exo/CCP) core-shell structured nanofibers (mimicking dermal structure) with sustained releasing ability of Exo. We will investigated the influence of the morphology and size of cCu-BGN and the processing parameters on the morphology, ion and exosomes releasing, degradation, antibacterial and biological activity of the biomimetic bilayer nanofibrous scaffold. Meanwhile, the behaviors of fibroblasts and endothelial cells induced by the structure, released ions and exosomes of the biomimetic bilayer nanofibrous scaffold, including the proliferation, migration and differentiation as well as the function of vital HIF-1α/VEGF signaling pathway will be studied. Furthermore, a diabetic cutaneous full-thickness wound model will be established to explore the stimulated effect of the biomimetic bilayer nanofibrous scaffold on diabetic wound healing. In summary, this project will provide brand new ideas and theoretical bases for the elaborate construction and clinical application of novel diabetic wound repair biomaterials and their related products.

糖尿病创面易感染、血管化差、愈合周期长，是临床常见又难修复的疾病。目前所用敷料缺乏活性，不能满足患者需求。我们前期发现纳米生物活性玻璃（BGN）及其复合材料可加快创面愈合。本项目拟构建一种模拟皮肤双层结构、抗菌并诱导组织再生的仿生双层分级结构纳米纤维支架用于加快糖尿病创面修复，该支架由胶原修饰含铜BGN（cCu-BGN）、胶原（Col）、聚己内酯（PCL）及外泌体（Exo）组成，上层为取向排列的cCu-BGN/Col/PCL（CCP）纤维、下层为缓释Exo的核壳结构Exo/CCP纤维。研究cCu-BGN对支架形貌、离子及外泌体释放、抗菌性能的影响；研究纤维取向、离子和外泌体通过物理、化学和生物介导细胞行为的效应关系及HIF-1α/VEGF通路的作用，揭示细胞应答机制；构建糖尿病全层皮肤缺损模型，阐明仿生双层支架诱导糖尿病创面修复的作用，为新型创面修复材料的精细构建及临床应用提供重要依据。

本项目针对目前糖尿病创面修复存在的易感染、血管化困难和愈合周期长而临床常用的敷料缺乏组织再生诱导性导致修复效果不理想等问题，构建具有抗菌、快速血管化并诱导组织再生性能的多功能支架以促进糖尿病创面愈合。本项目采用溶胶-凝胶技术结合有机模板来制备分布均匀、结构均一的含铜生物活性玻璃，在此基础上，制备了一种具有良好的抗菌活性和血管再生能力的可注射自修复的含铜生物活性玻璃复合支架，可促进内皮细胞的增殖和体外成血管能力，且释放的Cu2+通过激活HIF-1α/VEGF/VEGFR2信号级联上调了VEGF的表达，促进创面血管生成，加速胶原的沉积和重塑，有效促进了糖尿病创面的愈合。进一步，成功构建了一种外泌体的缓释支架将所提取的脂肪干细胞外泌体包裹持续缓慢释放，与单纯外泌体比，能更有效地促进糖尿病创面修复的快速及更完整修复。在上述基础上，通过静电纺丝技术，将含铜纳米生物活性玻璃与丝素蛋白等制备成不同结构的复合支架，体外实验证实所构建的复合支架能促进内皮细胞的增殖、迁移和体外成血管，通过构建糖尿病全层皮肤缺损模型证实所制备的含铜生物活性玻璃有序复合支架可加快糖尿病创面的更快和更完整的修复。综上所述，本项目研究证实了含铜生物活性玻璃和含外泌体复合支架具有优异的理化性能及良好的抗菌、血管再生和促组织修复等多功能特性，使其在糖尿病创面修复领域具有广阔的应用前景。本项目为高活性创面修复材料的研发和糖尿病创面的治疗提供了新思路。