尿源干细胞外泌体借助适配体靶向BMSCs促进成骨的作用功效与分子机制研究

The lack of clinical ideal therapies has long been a major problem of osteoporosis. Stem cells-derived exosomes exhibit similar pro-regenerative properties as stem cells, and the direct treatment with exosomes may avoid the potential risks associated with stem cell-based therapy, which may be a good strategy for promoting osteogenesis and treating osteoporosis. Thus, obtaining exosomes that can target bone tissues and efficiently promote osteogenesis has become a key scientific problem that need to be solved urgently. We previously found that exosomes secreted by human urine-derived stem cells (USC-Exo) were enriched in a class of pro-osteogenic proteins, and could accelerate osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), as well as prevent osteopenia in the ovariectomized mice. In this study, we will perform a series of experiments to validate the key pro-osteogenic proteins in USC-Exo and their downstream effectors. Next, we will augment the bone regenerative property of USC-Exo by enhancing the expression of the key pro-osteogenic proteins. Meanwhile, based on our previous study that has discovered a BMSCs-targeting Aptamer, we will utilize this Aptamer to decorate USC-Exo and thereby give USC-Exo the ability to target bone. Then, we will evaluate the pro-osteogenic effects of this USC-Exo-Aptamer system on BMSCs and osteoporotic mice. Our project aims to explore a new mode that USC-Exo enhance osteogenesis by targeting BMSCs via the guide of Aptamer and to elucidate the mechanisms involved, then to provide new ideas for the prevention and treatment of osteoporosis.

骨质疏松症（OP）一直缺乏理想的治疗手段。干细胞外泌体具有干细胞样促组织再生功能，却避免了干细胞直接移植的潜在风险，可能是促进骨再生及治疗OP的良好选择。获得靶向骨组织并高效促进骨再生的外泌体就成为亟待解决的关键科学问题。我们前期发现人尿源干细胞外泌体（USC-Exo）富集了一群促成骨分化相关蛋白，可促进BMSCs成骨分化，防止OP小鼠骨丢失。本项目拟进一步筛选验证USC-Exo中促进骨再生的关键蛋白，探明其下游效应分子；过表达上述关键蛋白强化USC-Exo的促成骨功能，同时，采用我们已筛选到的靶向BMSCs的核酸适配体（Aptamer）修饰USC-Exo以赋予其骨靶向性，在BMSCs和OP动物模型验证这种富含关键蛋白的USC-Exo-Aptamer复合体的促骨再生功效。该项目旨在探索USC-Exo借助适配体靶向BMSCs促进骨再生的新模式并揭示其分子机制，为临床OP防治研究提供新思路。

骨质疏松症（OP）导致骨脆性增加而易发生骨折，已成为危害人类健康的主要疾病之一。我们前期研究发现来源于1名健康成年人的尿源干细胞分泌的外泌体（USC-Exos）富集了多种功能性蛋白，可促进BMSCs成骨分化，防止去卵巢小鼠（绝经后OP模型）骨量丢失。本项目在此基础上，进一步探究了USC-Exos对多种OP小鼠模型的骨保护作用和促进OP性骨折修复的功效，在绝经后OP小鼠模型上评估了不同年龄阶段供者（儿童、成年人和老年人）来源USC-Exos的促骨再生功效，并明确介导USC-Exos发挥骨保护作用的关键蛋白分子。结果显示，USC-Exos不仅可促进成骨分化，还能显著抑制破骨细胞形成和增强内皮细胞血管新生活性，减轻老年性OP、绝经后OP和废用性OP小鼠模型骨量丢失，加速OP小鼠骨折修复，增强三种OP小鼠模型的骨强度，且其抗OP效能并不因USCs的供者年龄、性别以及健康状态（患OP或非OP）受到显著影响。USC-Exos中显著富集了胶原三螺旋重复蛋白1（CTHRC1）、骨保护素（OPG）和人恶性脑肿瘤缺失蛋白1（DMBT1），敲低上述蛋白分子后，USC-Exos的促成骨分化、抑破骨分化或（和）促血管新生功能显著减弱，对抗骨量丢失或（和）促进骨折修复的作用明显降低。本项目证实USC-Exos可通过转运功能性蛋白减轻OP和促进OP性骨折愈合，为临床OP及其骨折的防治开辟了一条新途径。