| In the present study ,some author had observed osteogenic seed cells modified by rhBMPs gene could repair longer segmental bone defect by expressing and secreting rhBMPs protein ,but also inducing themselves to defferienciate to osteoblasts.Although Marrow mesenchymal stem cells were easier to be cultivated,the expand volume of them was not enough to meet the need of bone defect repair. Our aim was to search for a sort of new gene-modified osteogenic seed cells combined with scafford to construct Tissue-Engineering Bone,which was accessible to repair longer segmental bone defect. Therefore,we designed the experiment to evaluate the osteoinductivity of adenovirus and liposome mediated human bone morphogenetic protein-2 gene(adv-hBMP-2) transfer in skeletal muscle stem cells of rabbits :in vitro and ex vivoExperiment OneThe osteoinductivity of Recombinent Human BMP-2 Gene transfer in Skeletal Muscle Stem Cells of rabbits: in vitro[Abstract] objective To evaluate the osteoinductivity of adenovirus and liposome mediated human bone morphogenetic protein-2 gene (adv-rhBMP-2) transfer in skeletal muscle stem cells of rabbits. Methods (1) Adv-rhBMP-2 was amplified in 293 cells and titered by the plaque assay. The skeletal muscle stem cells from rabbit were cultivated in vitro . Cultured SMSCs were divided in 3 groups as follows: I , Adv-rhBMP-2 trusduced group. II ,Adv-GFP trusduced group. III, untransduced group. (1)To determined the transfection rate and biological change of SMSCs transferred by adv-hBMP-2 at a mutiplicity of infection(MOI) of 50. (2) Gene transferred SMSCs were evaluated by immunohistochemistry,ALP, ELISA, alizarin red staining (ARS), osteocalcin, ALP activity quantitation. The osteoinductivity of gene transferred SMSCs in muscle of rabbit was tested. (2) pAGFP -rhBMP-2 purified were combined with cationic Liposome. (1) SMSCs were transfered by pAGFPrh-BMP-2-Liposome complex. (2) Gene transferred SMSCs were evaluated by immunohistochemistry. Result (1) (1)The transfection rate of SMSCs transferred by adv-rhBMP-2 was 79% ; The expand speed of SMSCs transferred was slower than that of the SMSCs untransferred at 3th day but as soon as them at 7th day. (2) Positive stain of rhBMP-2 , ALP, Collegen I and expression of osteocalcin were achieved in Adv-rhBMP-2 mediated SMSCs;the quantities of rhBMP-2 was increased at the 4th day and reached peak at the 14th~ 16th days after rhBMP-2 gene ransduction;The calcifed nudes were formed at the 20th day after transduction; (2) The transfection rate of SMSCs transferred by cationic Liposome mediated pAGFP-rhBMP-2 was 14.18%. Positive stain of rhBMP-2, ALP, Collegen I were achieved in the SMSCs transferred . conclusion (1) Quatity of SMSCs is enough to be target cells of gene therapy. (2) Adenovirus mediated bone morphogenetic protein-2 gene transfected SMSCs have the osteoinductivity in vitro. (3) Comparied to AdrhBMP-2, the transfection rate of liposom mediated bone morphogenetic protein-2 gene transfected SMSCs is lower. As the advent of new generation Liposome, It's possible to be used safely in the clinical treatment of bone defect. [Keyword] Bone morphogenetic protein;Gene therapy;Skeletalmuscle stem cell Bone defectExperiment TwoExperiment study on osteoinductivity of rabbit Skeletal Muscle Stem Cells transduced by AdrhBMP-2: ex vivo [Abstract] objective To determine whether Skeletal Muscle Stem Cells (SMSCs) modified by Adenovirus mediated bone morphogenetic protcin-2 gene ex vivo in combination with DBM could induce bone formation in muscle of nude mice and repair radial longer segemental defect in rabbit. Methods (1) 9 BalB/c nude mice were divided into 3 groups: I . Adv-rhBMP-2 trusduced SMSCs/DBM group (n=3). II, DBM group (n=3). III, Adv-rhBMP-2 trusduced SMSCs group (n=3) Materials were implanted into the muscle of nude mice. Macroscopic, histologic and roentgenogiaphic examination were performed at the end of 3th week af...
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