骨形态发生蛋白9复合胶原基骨修复材料体外成骨性能与体内修复骨缺损  

Osteogenesis in vitro and bone defect repair in vivo of bone morphogenetic protein 9 composite collagen based bone repair material

作　　者：庄传记 陈文昭[2] 江新民 Zhuang Chuanji;Chen Wenzhao;Jiang Xinmin(Department of Orthopedics,Second People’s Hospital of Jingdezhen,Jingdezhen 333000,Jiangxi Province,China;Department of Orthopedics,the First Affiliated Hospital of Nanchang University,Nanchang 330006,Jiangxi Province,China)

机构地区：[1]景德镇市第二人民医院骨科,江西省景德镇市333000 [2]南昌大学第一附属医院骨科,江西省南昌市330006

出　　处：《中国组织工程研究》2021年第10期1489-1494,共6页Chinese Journal of Tissue Engineering Research

基　　金：江西省健康委员会支持项目(20204356),课题名称:临床阳性体征与膝关节半月板损伤的相关性研究,项目负责人:庄传记。

摘　　要：背景:胶原基骨修复材料具备良好的骨传导性、骨诱导性、降解性、机械性能与生物相容性,目前已被广泛应用于临床骨修复中,但其诱导成骨效果仍不及自体骨。目的:将骨形态发生蛋白9(bone morphogenetic protein 9,BMP-9)复合于胶原基骨修复材料中,观察其诱导成骨的能力。方法:将编码BMP-9的质粒DNA负载到胶原基骨修复材料中,构建复合BMP-9的胶原基骨修复材料。①体外实验:将SD大鼠脂肪间充质干细胞分别接种于复合BMP-9的胶原基骨修复材料与胶原基骨修复材料中,利用MTT法检测细胞增殖,扫描电镜观察细胞黏附,DAPI染色后计数细胞黏附数量,碱性磷酸酶活力试剂盒检测成骨诱导后细胞中的碱性磷酸酶活性,qRT-PCR检测成骨诱导后细胞中的骨桥蛋白与骨钙素mRNA表达量;②体内实验:在15只SD大鼠右后肢股骨内上髁制作直径3 mm、深3 mm的骨缺损,空白组不植入任何材料,对照组植入胶原基骨修复材料,观察组植入复合BMP-9的胶原基骨修复材料,术后8周取材,分别进行Micro-CT检查与组织学观察。实验方案经南昌大学实验动物科学中心伦理委员会批准。结果与结论:①体外实验显示,复合BMP-9胶原基骨修复材料中培养1,3,5,7 d的细胞增殖与胶原基骨修复材料比较差异无显著性意义(P﹥0.05);②体外共培养24 h后,在复合BMP-9胶原基骨修复材料表面黏附的细胞较多,并伸出较多的伪足,黏附细胞计数与铺展情况优于胶原基骨修复材料;③体外成骨诱导3,7 d后,复合BMP-9胶原基骨修复材料表面细胞的碱性磷酸酶活性高于胶原基骨修复材料(P <0.05),成骨诱导7 d后的骨桥蛋白与骨钙素mRNA表达量高于胶原基骨修复材料(P <0.05);④体内实验Micro-CT检查显示,观察组新骨生成量高于对照组、空白组(P <0.05),对照组高于空白组(P <0.05);⑤体内实验苏木精-伊红与Masson三色染色显示,观察组的骨修复效果优于对照组BACKGROUND: Collagen-based bone repair materials have been widely used in clinical bone repair due to their excellent osteoconductivity, osteoinduction, degradation, mechanical properties and biocompatibility, but their osteoinduction effect is not as good as that of autogenous bone.OBJECTIVE: Bone morphogenetic protein 9(BMP-9) was incorporated into collagen matrix to investigate its osteoinductive ability.METHODS: The plasmid DNA encoding BMP-9 was loaded into collagen-based bone repair materials to construct a composite of collagen-based bone repair materials.(1) In vitro experiment: The Sprague-Dawley rat adipose mesenchymal stem cells were inoculated with BMP-9 composite collagen-based bone repair material and collagen-based bone repair material. Cell proliferation was detected by MTT assay, and the cell adhesion was observed by scanning electron microscope. Cell adhesion was counted after DAPI staining. Alkaline phosphatase activity was detected by alkaline phosphatase kit. Osteopontin and osteocalcin mRNA expression levels were detected by qRT-PCR.(2) In vivo experiment: Bone defects of 3 mm in diameter and 3 mm in depth were made in the medial epicondyle of the right hind limb of 15 Sprague-Dawley rats. No material was implanted in the blank group. Collagen-based bone repair material was implanted in the control group. BMP-9 composite collagen-based bone repair material was implanted in the observation group. Samples were obtained at 8 weeks after surgery. Micro-CT examination and histological observation were performed. This study was approved by the Ethics Committee of Laboratory Animal Science Center of Nanchang University. RESULTS AND CONCLUSION:(1) In vitro experiment showed that there was no significant difference between the proliferation of adipose tissue mesenchymal stem cell cultured for 1, 3, 5, and 7 days in collagen-based bone repair materials compounded with BMP-9(P > 0.05).(2) After co-culture in vitro for 24 hours, more cells and more pseudopodia appeared on the surface of collagen-based b

关 键 词：骨 材料 胶原基骨 骨形态发生蛋白 骨缺损 成骨 骨诱导 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学]