衰老骨基质外泌体通过转运miR-483-5p抑制H型血管再生参与老年骨质疏松症发病

H-type vessel (HV) couples angiogenesis and osteogenesis. Aging is coupled with decrease of both HV and bone formation, leading to osteoporosis. Increasing HV can alleviate senile osteoporosis via promoting bone regeneration. However, the mechanism for the reduction of HV in aging body is unclear. We have found that the aged bone matrix exosomes (AB-Exo) are highly enriched with miR-483-5p, which has angiogenic inhibitory function, and inhibit the formation of HV in mice. We then raised scientific hypothesis: in aged body, bone matrix exosomes (B-Exo) released during bone resorption inhibit HV formation via transport miR-483-5p, thus result in senile osteoporosis. To verify this hypothesis, we will (1) use exosome tracing mouse model (CD63-GFP transgenic mice) to reveal the source cell and intraosseous distribution of B-Exo; (2) verify the signaling pathway of miRNA-483-5p induced HV reduction; (3) deliver antagomiR-483-5p to AB-Exo in the bone marrow cavity via tail-intravenous injection of antagomiR-483-5p conjugated with an AB-Exo-targeted aptamer, and then verify its protective effect on HV and bone in a senile OP animal model. The aim of this project is to explore the mode of action and molecular mechanism of AB-Exo induced reduction of HV, and to provide new targets and new ideas for the prevention and treatment of senile OP.

H型血管（HV）偶联血管生成与骨形成，衰老伴随HV减少与骨形成减少，导致骨质疏松症（OP）；增加HV通过促进骨再生防止老年OP。衰老机体HV减少的机制不明。我们前期发现，衰老骨基质外泌体（AB-Exo）高度富集具有血管生成抑制功能的miR-483-5p，抑制小鼠HV生成。我们据此假设：衰老机体骨吸收过程中释放的骨基质外泌体（B-Exo）转运miRNA-483-5p抑制HV生成从而导致老年OP。为验证此假设，我们拟：采用外泌体示踪模型（CD63-GFP转基因小鼠）探明B-Exo的来源细胞与骨内分布；验证miRNA-483-5p作用于HV的分子机制；借助适配体，通过尾静脉注射，把antagomiR-483-5p靶向递送到骨髓腔中的AB-Exo，在老年OP动物模型中验证其对HV与骨的保护功效。该项目旨在探索AB-Exo导致HV减少的作用模式与分子机制，为老年OP防治提供新靶点与新思路。

骨质疏松症是中老年人常见病，多发病。它是以骨量减少、骨微结构破坏为特征的全身性骨骼代谢性疾病。成骨细胞骨形成不足和（或）破骨细胞骨吸收过多是导致骨质疏松症发生发展的细胞学基础。2014年，骨骼特异性H型血管被发现，它通过偶联成血管和成骨营造有利于骨生成的骨髓微环境。同年，项目负责人率先探明破骨前体细胞通过分泌PDGF-BB促进H型血管形成与骨形成，提出基于破骨前体细胞的“骨血管-骨吸收-骨形成偶联”的骨代谢三元调控模式（Nat Med, 2014）。H型血管随增龄而减少，相反，骨丢失则随增龄而显著增加。该项目按原计划明确了骨基质胞外囊泡的骨细胞来源，探明骨细胞胞外囊泡可转运到H型血管，发现年轻骨细胞胞外囊泡（YO-EVs）促进H型血管生成与骨形成，而衰老骨细胞胞外囊泡（AO-EVs）则抑制H型血管生成与骨形成。另外，由于原计划研究的miRNA-483-5p没有如预期在体内发挥对H型血管生成的显著抑制作用，我们通过蛋白质组学技术另筛选出了高度富集于AO-EVs中的Lumican蛋白，探明其介导AO-EVs抑制血管内皮细胞成管，可能是抑制H型血管生成的关键功能分子。我们将在2023年完成Lumican特异性敲除转基因小鼠的体内表型验证实验。该研究为老年骨质疏松症防治提供了新靶点与新思路。