LRP5核转位调控circRNA_19081/miR-466k/BMP7轴在种植体骨结合中的作用及机制研究

The mechanisms how the roughened surfaces on dental implants promote bone marrow mesenchymal stem cells into osteoblasts to form osseointegration have not been well elucidated. Our previous study has found that low density lipoprotein receptor-related protein 5 (LRP5), as one of the membrane receptors of classical Wnt signaling pathway, underwent nuclear translocation accidently. The results of microarrays showed that the expressions of circRNA_19081 sponging with miR-466k was upregulated, and the expression of BMP7 was upregulated when nuclear LRP5 (nLRP5) was overexpressed. This project proposes that nLRP5 can establish rapid and stable osseointegration through promoting the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), and presents that nLRP5 can regulate the circRNA_19081/miR-466k/BMP7 axis and contribute to enhancing osseointegration. This project constructs LRP5-LOV2 nuclear transport system to establish precise spatial and temporal control of LRP5, and explores the possibility of using nLRP5 overexpressing adenoviruses to enhance osseointegration in LRP5 knock-out mice. Our project can provide a new strategy and therapeutic target to establish more rapid and stable osseointegration.

种植体粗化表面如何促进BMSCs成骨分化，从而形成种植体骨结合，具体的形成机制研究较为薄弱。本课题组前期研究意外发现，LRP5作为经典的Wnt信号通路的细胞膜上受体之一，存在细胞膜受体核转位现象，且能促进BMSCs成骨分化，全转录组基因芯片发现核内LRP5（Nuclear LRP5，nLRP5）使circRNA_19081、BMP7表达增加，circRNA_19081对miR-466k有海绵吸附作用，据此提出利用nLRP5的促成骨功能建立更加快速、稳定的种植体骨结合，并研究nLRP5调控circRNA_19081/miR-466k/BMP7轴在种植体骨结合中的机制。本项目通过构建LRP5-LOV2核转运系统，精准调控LRP5的时空位置，并探索体内通过转染nLRP5腺病毒加快在LRP5敲除小鼠体内种植体骨结合的可能性。本研究可为建立更加快速、稳定的种植体骨结合提供新的思路和治疗靶点。

种植体粗化表面如何促进BMSCs成骨分化，从而形成种植体骨结合，具体的形成机制研究较为薄弱。本项目按照课题计划书所设计的研究方案，利用免疫荧光、免疫印迹、Flag标记等方法明确了跨膜受体LRP5进入核内的现象。全转录组芯片检测发现LRP5核内过表达后circRNA_19081、circRNA422明显上升，RT-qPCR进一步验证了circRNA422的促成骨能力优于circRNA_19081，因此选择circRNA422进行后续研究。体内外实验验证了circRNA422通过SP7/LRP5环路加快骨形成的机制。本项目通过深入探究LRP5的亚细胞定位，构建出可调控LRP5全长蛋白出核与入核的光遗传工具，证实LRP5出核上调BMP7、Runx2等成骨指标，促进BMSCs成骨分化，揭示了LRP5核转位的新机制。本研究为建立更加快速、稳定的种植体骨结合提供了新的思路和治疗靶点。本项目圆满完成研究任务，共发表基金资助论文17篇（其中高水平论文9篇），申请发明专利4项，协助培养硕士研究生2名、博士研究生1名。项目负责人以该项目为基础获得硕士研究生招生资格，获中华口腔医学会2项会议一等奖。