人骨形态发生蛋白-2基因腺病毒载体异位成骨

目的：评价人骨形态发生蛋白2(hBMP-2)基因腺病毒表达载体(recombinant adenoviral vector carrying the human BMP-2 gene, AdBMP-2)成骨能力并探讨其成骨机制。方法：应用 AdBMP-2体外感染兔骨髓间质干细胞（BMSC），7 d后观察hBMP-2和碱性磷酸酶(ALP)的表达,21 d后观察感染细胞钙结节形成能力。同时将AdBMP-2行裸鼠腓 肠肌组织内注射，术后20 d取材，观察hBMP-2的表达及肌组织内异位成骨情况。结果：AdBMP-2可高效感 染BMSC并获得hBMP-2的有效表达，ALP表达增高并形成钙结节。AdBMP-2在体内同样表达hBMP-2， 并且以软骨化骨的方式启动成骨。结论：感染AdBMP-2的BMSC向成骨细胞分化，AdBMP-2在体内有着良好的成骨能力。

Ectopic bone formation using bone morphogenetic protein 2 adenoviral vector

Objective To evaluate the osteoinductive activity of recombinant adenoviral vector carrying the human BMP-2 gene(AdBMP-2) and investigate the mechanism of bone formation induced by the bone morphogenetic protein 2(BMP-2).Methods AdBMP-2 infected rabbit bone messenchymal stem cells(BMSC) were cultivated in vitro.The expression of hBMP-2 was determined by in situ hybridization and immunohistochemical analysis,and ALP was determined by enzyme cytochemistry 7 d after infection.At 21th day,Alizarin red S staining was used to evaluate calcium-rich deposits by cells transfected in culture.30 μL AdBMP-2 solution(2×10~(12)pfu·mL~(-1)) was injected into the muscle of the left hind limb of the nude rat,and 30 μL Adβ gal solution(2×10~(12)pfu·mL~(-1)) was injected into the muscle of the right hind limb as control.Animals were sacrificed at 20th day following injection to investigate the bone formation histologically and the expression of BMP-2 in the specimens of the injection sites by immunohistochemistry and in situ hybridization method.Results The expressions of BMP-2 and ALP in the transduced BMSC were confirmed at 7 th day after infection.A deeply stained mineralized layer was evident at 21th day in transduced cells.In vivo experiment,BMP-2 gene therapy induced mesenchymal monocytes chemotaxis,proliferation, differentiation into chondrocytes,with subsequent endochondral bone formation.At the same time,hBMP-2 expression was observed in the chondrocytes,osteoblasts and mesenchymal monocytes.(Conclusion AdBMP-2) posseses the osteoinductive activity in vitro and in vivo.