缓释外泌体的种植体表面改性及其靶向抑制破骨细胞分化增强骨结合的研究

In recent years, titanium implant has become one of the major methods of dental prosthesis restoration. Bone implant contact (BIC) is essential for long-term suc¬cessful implant osseointegration and, therefore, bone quality and quantity around the implant are particularly important. Osteoporosis is a bone metabolism-related disease characterized by bone loss and bone microstructural destruction, which in turn leads to increased bone fragility and fracture. Furthermore, osteoporosis is the main factor that restricts the successful repair of implant in elderly patients. The dynamic balance of osteoclasts and osteoclasts are the main causes of osteoporosis. It is well established that excessive activation of osteoclasts is essential for osteoporosis. Surprisingly, the preosteoclast has a pro-osteogenesis role. Therapeutically blocking osteoclast differentiation around the implant holds great promise for osteointegration enhancement. Thus, revealing the main mechanism governing the excessive activation of osteoclasts and developing therapeutic strategies hold the promise for osteointegration enhancement. Previously, we found that monocytes from either OVX rat or aged rat expressed much higher level of Cathepsin K (CTSK), which in turn promotes osteoclast differentiation. On the other hand, we found that exosomes are an ideal vehicle for delivering siRNA into monocytes safely and efficiently, with many advantages when compared with other nanoparticles. In light of our previous work on implant surface modification and gene drug delivery, we will embed CTSK-siRNA exosomes onto the implant surface by the layer-by-layer (LbL) assembly technique. The surface characteristics and control release of CTSK-siRNA will be analyzed by SEM and endocytosis assay. Furthermore, the effects on the osseointegration ability of our novel implant design in osteoporotic condition will be explored by both in vitro and in vivo experiments, through analyzing the effects on angiogenesis, osteogenesis and others. On this basis, the specific mechanism of bone regeneration was clarified by molecular biology, by focusing on the downstream effects of CTSK on the paracrine factors. The proposal here would be promising in developing a novel strategy to improve the osseointegration in the context of osteoporosis.

骨质疏松时种植体骨结合能力降低，是相关患者种植修复效果差的主要原因。靶向增强种植体骨结合能力，对改善种植治疗效果有重要意义。既往研究发现，破骨细胞过度活化是骨质疏松的重要原因，与破骨细胞功能相反，其前体细胞却促进成骨分化；靶向抑制种植体局部破骨细胞分化有望促进种植体骨结合。课题组前期发现，雌激素缺乏和衰老个体来源破骨前体细胞中CTSK表达增高是其破骨细胞分化能力增强的共同机制；另一方面，外泌体能够向单核细胞安全高效递送核酸药物。据此，我们推测在种植体表面构建能够缓释携带siCTSK的外泌体涂层，有望高效抑制破骨细胞分化，促进骨结合。本项目拟结合前期种植体表面改性的研究基础，通过层层自组装技术将外泌体负载于钛表面；通过材料学、动物和细胞实验系统证明阐明该钛材表面修饰策略促进骨结合的作用及具体机制，重点关注CTSK引起破骨前体细胞旁分泌关键因子的改变及其功能；期望为改善种植体骨结合提供新策略。

骨质疏松时种植体骨结合能力降低，是相关患者种植修复效果差的主要原因。靶向增强种植体骨结合能力，对改善种植治疗效果有重要意义。破骨细胞过度活化是骨质疏松的重要原因，与破骨细胞功能相反，其前体细胞却促进成骨分化；靶向抑制种植体局部破骨细胞分化有望促进种植体骨结合。本项目结合前期种植体表面改性的研究基础，通过层层自组装技术将外泌体负载于钛表面；通过材料学、动物和细胞实验系统证明阐明该钛材表面修饰策略促进骨结合的作用及具体机制，重点关注CTSK引起破骨前体细胞旁分泌关键因子的改变及其功能。主要取得了以下几方面的结果：1）建立了高生物相容性多层级纳米涂层改构种植体的策略；2）建立了缓释负载siCTSK纳米颗粒的种植体，阐明其通过重塑微环境促进骨再生的功能和具体机制；3）创建负载siCTSK、IL10、BMP2、VEGF等的外泌体，并成功将其用于钛基表面改性；证实其促进种植体周围血管新生和成骨的功能；4）创建了OX40L和CD62L修饰的外泌体，为调节免疫功能的外泌体工程化提供了依据和策略，为增强外泌体的组织相容性研究奠定了基础；5）建立靶向IL1等多个炎性基因的基因编辑和递送系统，为外泌体调控炎症及种植体改性提供了新选择。上述研究在Biomaterials、Molecular Therapy-Nucleic Acids期刊发表SCI论文2篇，授权国家发明专利3项，并在学术会议上做报告1次。