镁金属结合骨牵引技术通过CGRP促进大段骨缺损修复的机制研究

Despite investigations focusing on the motivation of intrinsic potentials have been intensively conducted, the repair of segmental bone defect of critical size remains a challenging problem for orthopaedic surgeons. Distraction osteogenesis has been developed to generate new bone formation to fill the defect site, however, major drawbacks do also exist, including that it is required to maintain the affected bone externally fixed for a relatively long period, as the new bones usually consolidate very slowly, which therefore increases the risk of infection. With the degradable capacity, the bioactive magnesium-based implants have become innovative orthopaedic fixators. Our team previously published an original study of milestone significance in the prestigious Nature Medicine, reporting that magnesium stimulates the production as well as secretion of calcitonin gene-related peptide (CGRP) to promote the osteogenic differentiation of stem cells which leads to significant improvement in the healing of fracture established at weight-bearing site (i.e. femur) in ovariectomy-induced osteoporotic rat model. In the currently initiated project, to validate the scientific hypotheses: magnesium and distraction osteogenesis would synergistically promote the functional repair of segmental bone defect of critical size through elevating CGRP-coordinated coupling between angiogenesis and osteogenesis, we will comprehensively employ approaches of histology, immunohistochemistry, X-ray, micro-CT, angiography, biomechanical test, micro-indentation, flow cytometry, PCR, and western blotting to assess the outcomes at histological, cellular and molecular levels. In addition, capsaicin-pretreated and Ramp1-knockdown animals will be used to further confirm the crucial roles of CGRP. In conclusion, this project will be of great scientific value as well as translational potential. The combined application of magnesium and distraction osteogenesis will likely serve as a promising alternative for bedside treating critical size bone defect, which may help reduce the morbidity and the burdens to patients, healthcare system, and our society.

如何通过动员机体内在潜能来修复大段骨缺损是长期以来的热点问题。骨牵引技术可诱导新骨长入，但缺点是新生骨组织塑形慢，患者需长时间携带外固定架，故感染风险高。镁金属正迅速发展为具生物活性的可降解骨科内固定物。本课题组曾在Nature Medicine发表具里程碑意义的研究，报道镁可刺激感觉神经末梢CGRP的产生和释放而显著增强大鼠股骨骨折的愈合。因此，我们推测镁金属与骨牵引术可通过CGRP调控的血管形成和干细胞成骨分化偶联而协同性促进骨缺损的功能性修复。本项目拟综合运用组织学、免疫组化（荧光）、X-ray、micro-CT、血管造影、生物力学及微米压痕测试、流式细胞术、PCR及蛋白印迹来实现在组织、细胞及分子水平验证以上科学假说。同时，将应用辣椒素及敲减Ramp1以进一步确定CGRP的重要作用。本项目具良好的转化前景，为临床治疗骨缺损提供新方案，有助降低致残率，减轻患者、医疗系统及社会的负担。

大段骨缺损由于缺乏自我修复能力，致残率很高，给患者、医疗系统及社会留下各种巨大的负担。其治疗十分棘手，目前主要依赖于填充自体骨或同种异体骨，但这些方案都有一些明显的不足。如何通过动员机体内在潜能来修复骨缺损，是长期以来的热点问题。骨牵引（Distraction Osteogenesis, DO）技术用于骨科临床已有百余年历史，可诱导新骨长入到牵引所致的缺损区，但缺点是耗时长，新生骨组织塑形慢，患者需长时间携带外固定架，由此也常增加感染风险。镁金属正迅速发展为具生物活性的可降解骨科内固定物。本课题组曾在Nature Medicine报道镁可刺激感觉神经末梢CGRP的产生和释放，CGRP再作用于骨膜干细胞，从而明显增加骨质疏松大鼠受力部位（股骨）骨折愈合。在本项目中，我们综合运用了组织学、免疫组化（荧光）、X-ray、micro-CT、血管灌注显影、PCR及蛋白印迹等技术在组织、细胞及分子水平证实了镁金属与骨牵引术可通过CGRP调控的血管形成和干细胞成骨分化偶联而协同性促进骨缺损的功能性修复。在 5 mm 股骨节段缺损大鼠模型中，术后两周时，我们发现与单独使用 DO 组相比，镁（Mg）钉联合DO治疗组的新骨形成增加约 4 倍，新血管形成增加5 倍。本项目具有显著的科学意义及转化前景，为临床治疗大段骨缺损提供新方案。