Adrenomedullin促进组织工程骨血管化构建的实验研究

Artifical bone scaffold has a poor destiny due to the insufficient angiogenesis, which mostly induced by the inner hypoxia environment and metabolic product accumulation. The key point in bone tissue engineering is how to promote the rapid angiogenesis in the scaffolds. To aim at this problem, this project will introduce a multifuctional peptide - adrenomedullin into artificial tissue-engineering bone using ionic crosslinking emulsion and thermally induced phase separation methods, and make the microsphere-scaffold composite heparin-functionalized, which was based on preliminary work. In vitro, we will implant the vascular endothelial cells onto the scaffolds to evaluate the cell migration, adhesion and growing into the scaffolds loaded with ADM by SEM observation, transwell assay, caculation with microscopes and detect related gene and protein expression levels as well by real-time PCR and immunohistochemistry. To combining with the animal experiments, we will clarify the in vivo effect of ADM on angiogenesis around or into the scaffold via intravital microscopic montages, immunohistochemistry. In addtion, the new bone formation around the scaffold related to angiogenesis was evaluated by microCT, histology, immunohistochemistry. This project will bring an apealing candidate-adrenomedullin for angiogenesis of tissue-engineering bone, and provide theoretic and experimental proof for angiogenesis and bone regeneration of artifical tissue-engineering bone loaded with ADM, which will has a promising future for clinical application.

人工骨支架材料由于其内部缺氧环境及局部代谢物堆积易导致其血管化不足，从而影响最终成骨效果。如何使骨支架材料快速血管化成为骨组织工程的关键问题。本项目以此为背景，以研究多功能调节肽Adrenomedullin（ADM）在骨组织工程中的作用及其机理为目的，在前期工作的基础上，将ADM载入骨支架材料并对其进行肝素化改性。在此基础上建立与血管内皮细胞共培养体系，利用Transwell迁徙实验、扫描电镜观察负载ADM的组织工程骨对细胞迁徙、粘附、生长的促进作用，通过实时定量PCR和免疫组化技术研究成血管相关基因、蛋白表达水平的变化；并结合动物实验，从体内水平研究ADM的引入对组织工程骨血管化构建及骨化速度与质量的促进作用，阐明ADM促进组织工程骨血管化构建可能的机理。本课题为骨组织工程治疗寻找了新靶点，为ADM应用于临床骨缺损的修复提供理论基础和实验依据，具有广阔的临床应用前景。

人工骨支架材料由于其内部缺氧环境及局部代谢物堆积易导致其血管化不足，从而影响最终成骨效果。如何使骨支架材料快速血管化成为骨组织工程的关键问题。本项目以此为背景，以研究多功能调节肽Adrenomedullin（ADM）在骨组织工程中的作用及其机理为目的，在前期工作的基础上，将ADM载入骨支架材料并对其进行肝素化改性。在此基础上建立与血管内皮细胞共培养体系，利用Transwell迁徙实验、扫描电镜观察负载ADM的组织工程骨对细胞迁徙、粘附、生长的促进作用，通过实时定量PCR和免疫组化技术研究成血管相关基因、蛋白表达水平的变化；并结合动物实验，从体内水平研究ADM的引入对组织工程骨血管化构建及骨化速度与质量的促进作用，阐明ADM促进组织工程骨血管化构建可能的机理。Heparin接枝后可以明显改善多肽的释放，同时我们证明ADM在多种支架材料中均能促进血管化进程，同时动物实验也证明了含有ADM的支架具有更好的骨修复效果。本课题为骨组织工程治疗寻找了新靶点，为ADM应用于临床骨缺损的修复提供理论基础和实验依据，具有广阔的临床应用前景。