如何有效促进骨再生治疗骨折不愈合是骨科棘手问题，miR-196a因具有促成骨作用而备受关注，但其调控机制仍不十分清楚。申请人前期已发现miR-196a能促进大鼠颅骨缺损再生，且在骨折不愈合患者血样中呈低表达。本项目拟继续扩大临床样本量，明确骨折不愈合与miR-196a的科学关系及临床价值；依据前期发现HoxA9的3’UTR1210-1216：-ctacct-为关键结合位点，利用RNA干涉和CO-IP技术阐明miR-196a-HoxA9-TGF-β互作机制，及该分子轴对BMSC增殖、迁徙和分化的影响；采用Cre/LoxP系统和CRISPR-Cas9技术获得miR-196a条件敲除模式动物，通过三维CT、IHC及IF等技术证实miR-196a-HoxA9-TGF-β为骨再生关键调控轴。本项目将深入探索miR-196a的临床意义，阐明骨折不愈合发病机制，并为将来针对性治疗策略的研发提供理论依据。

To effectively promote bone regeneration is in urgent clinic need. miR-196a stimulates osteogenesis; however, the mechanism remains unknown. We have found that miR-196a promoted rat calvarial bone defect regeneration and was lower expressed in bone nonunion patients. In this project, we plan to explore the clinic and scientific relation between bone nonunion disease and miR-196a through testing the clinic blood samples; Based on the finding of the miR-196a binding site in HoxA9 3’ UTR 1210-1216：-ctacct-, we will elucidate the communication among miR-196a-HoxA9-TGF-β axis via RNA interfering and CO-IP, and explore how this axis regulates BMSC migration, proliferation and differentiation; Upon Cre/LoxP and CRISPR-Cas9 technique, we successfully acquired miR-196a conditional knockout mouse. MicroCT, Immunohistochemistry and immunofluorescent staining analysis will be studied to explore the role of miR-196a-HoxA9-TGF-β in bone regeneration in vivo. In this project, we will further explore the clinic value of miR-196a, elucidate the pathological mechanism of bone nonunion, and provide theorical proof for future treatment.