聚乙二醇-骨形态发生蛋白-2-聚乳酸/聚已内酯载基因仿生骨促进骨缺损修复的实验观察  

Bone morphogenetic protein-2-encupsulated PEG-grafted-poly-lactic acid-polycaprolactone nanoparticles promote bone repair

作　　者：王湛[1] 许晓军[1] 杨军[1] 丁立峰[1] 李建军[1] 

机构地区：[1]中国医科大学附属盛京医院骨科,沈阳110004

出　　处：《中华医学杂志》2015年第11期865-869,共5页National Medical Journal of China

基　　金：辽宁省自然科学基金（20082118）;辽宁省教育厅科学技术研究基金（L2010652）

摘　　要：目的 观察聚乙二醇（PEG）/骨形态发生蛋白（BMP）-2纳米基因复合物及载基因仿生骨与骨髓间充质干细胞（BMSCs）对骨缺损修复的共同作用.方法 通过离子交联法制备PEG/BMP-2纳米基因复合物,通过溶液共混法制备聚乳酸（PLA）/聚已内酯（PCL）载基因仿生骨并接种BMSCs于仿生骨之上,应用免疫组化及免疫印迹检测BMSCs转染后BMP-2蛋白表达;酶联免疫方法检测转染后细胞培养上清中BMP-2分泌情况;实时聚合酶链反应检测转染后细胞BMP-2及骨钙素mRNA表达水平;截除新西兰大耳白兔双侧桡骨中段,植入载基因仿生骨材料,对骨缺损修复部位摄X线片、行HE染色和BMP-2免疫组织化学染色.结果 制备出PEG/BMP-2纳米颗粒及PEG-BMP-2-PLA/PCL载基因仿生骨.PEG/BMP-2纳米颗粒转染BMSCs和载基因仿生骨BMSCs内BMP-2表达量明显上调,骨钙素mRNA表达和碱性磷酸酶活性有所增加;体内实验中,PEG-BMP-2-PLA/PCL载基因仿生骨组与对照组比较BMP-2表达升高,新生骨在骨缺损区域所占面积比也明显增加.结论 PEG-BMP-2-PLA/PCL对骨缺损修复具有良好的效果.Objective To explore the efficacy of a novel tissue engineered bone in repairing bone defects using poly-lactic acid-polycaprolactone （PLA-PCL） scaffolding seeded with PEG-bone morphogenetic protein-2 （BMP-2） transfected rBMSCs （rabbit bone marrow stromal cells）.Methods rBMSCs were harvested,transfected with PEG/BMP-2 or liposome/BMP-2 and then implanted into PLA-PCL tissue engineered bone.The protein level of BMP-2 was assessed by Western blot and immunohistochemistry.Enzyme-linked immunosorbent assay （ELISA） was employed to measure the amount of BMP-2 in culture media.The mRNA levels of BMP-2 and osteocalcin were assayed quantitatively by real-time polymerase chain reaction （PCR）.The middle portion of bilateral radius in New Zealand rabbits was excised and implanted with tissue engineered bone.And the modified areas were monitored by radiology,hematoxylin-eosin staining and immunohistochemical staining of BMP-2.Results PEG-BMP-2 nanoparticles （NPs） and BMP-2 loaded PEG-PLA-PCL tissue engineered bones were successfully constructed.The novel PEG-PLA-PCL NPs/DNA complex was superior for transfecting BMP-2 in rBMSCs compared to traditional liposomes.Moreover,the mRNA level of osteocalcin and alkaline phosphatase activity also increased after a transfection of BMP-2 encapsulated NPs.Conclusion PEG-PLA-PCL NPs/BMP-2 complex facilitates bone repair so that it provides theoretic rationales for potential clinical treatments.

关 键 词：修复外科手术 髓样组细胞 骨形态发生蛋白质类 仿生学 基因 纳米颗粒 

分 类 号：R687.3[医药卫生—骨科学] R318.08[医药卫生—外科学]