非病毒载体PEI-LA介导BMP-2基因直接投递治疗种植体周围炎性骨缺损的实验研究

Peri-implantitis could lead to the inreversible supporting bone loss, which severely damage the longterm success of implant. The use of gene therapy to express target genes to accelerate the regeneration of infective bone defect is the research frontier. Previously, we applied the viral vector to deliver BMP-2 gene via ex vivo pathway to successfully correct canine peri-implantitis. However, this approach suffers from biosecurity problems and requiring complex steps, which suggests that the key of advancing gene therapy is to upgrade the safety and effectiveness of vector design. We have proven the ex vivo BMP-2 gene delivery effect of our new non-viral vector PEI-LA in the critical cranial bone defect model on rats, which suggested the possibility of in vivo delivery of target genes by PEI-LA. Also, we noticed the decreased differentiation of Th17 in mouse caused by the injection of vector/gene complex. And, this cell has been recognized playing a central role in osteoclastogenesis. However, the relevant mechanism is still unclear. Thus this study will focus on the gene therapy of peri-implantitis to realize the supporting bone formation, and to judge the efficiency of gene transfection under the inflammation environment. Meanwhile, we will focus on the interaction between immunology and bone formation, to explore the effect of PEI-LA vector and vector/gene complex on the differentiation of CD4+T, to elucidate the influence elements of gene therapy applying PEI-LA on the reconstruction of inflammation caused bone defect, and to lay the foundation for peri-implantitis correction by gene therapy.

种植体周围炎可导致支持骨的不可逆性丧失，严重威胁种植体的长期成功。应用基因治疗实现目的基因表达促炎性骨缺损的快速再生是攻关前沿。我们前期采用病毒载体体外投递BMP-2成功修复犬种植体周围炎，但该法存在安全隐患且步骤繁琐，提示制备高效、安全的载体并选择体内投递是推进基因治疗的关键。我们已验证新型非病毒载体PEI-LA经体外间接法投递BMP-2修复大鼠标准颅骨缺损的效果，提示了体内直接投递的可行性；并观察到载体/基因复合体能降低小鼠体内Th17细胞的分化，而后者在破骨中起主导作用，但相关机制尚不明确。因此，本研究拟以种植体周围炎基因治疗为主线，以增加周围骨形成为重点，评估炎症条件下基因转染的效率；并围绕免疫与成骨的相互作用，探索PEI-LA载体和载体/基因复合体对CD4+T细胞分化的影响，阐明PEI-LA介导的基因治疗对促进炎性骨缺损修复的影响因素，为实现基因治疗改善种植体周围炎奠定基础。

种植体周围炎可导致支持骨的不可逆性丧失，严重威胁种植体的长期成功。应用基因治疗实现目的基因表达促炎性骨缺损的快速再生是攻关前沿。课题组首次比较了新型非病毒载体PEI-LA经体外间接法和体内直接法投递BMP-2基因修复大鼠标准颅骨缺损的效果，发现两者均能诱导局部骨组织的再生，在此基础上初步分析了非病毒载体PEI-LA系统对免疫相关细胞增殖、分化的影响；在病原菌诱导大鼠种植体周围炎模型上，系统评估了周围骨及牙龈软组织再生水平；发现低氧炎症环境中脂肪干细胞和BMP-2间接投递法可通过激活AMPK促进上颌窦中心骨再生，进而实现种植体快速骨结合；明确了机械力诱导的局部炎症环境中，使用他汀类药物可抑制牙周膜干细胞中AMPK/MAPK/NF-kB通路，实现被迁移磨牙拉升侧的成骨，抑制局部破骨吸收。本研究以种植体周围炎性骨缺损的治疗为主线，围绕骨再生与植体-骨再结合，初步探讨了AMPK活化调控下免疫与成骨的相互作用，综合性阐明PEI-LA介导BMP-2直接投递促进骨再生的机理，对优化非病毒载体设计，提高疾病状态下载体的转染效率，优化载体免疫属性等方面具有重要的指导意义，并为治疗其它骨炎性疾病的治疗提供新的思路和策略。