RESUMO
OBJECTIVE: To evaluate the osteogenic potential of bone morphogenetic protein (BMP)-2 gene transfected goat bone marrow-derived mesenchymal stem cells (MSCs). METHODS: Goat bone marrow-derived MSCs were transfected by Adv-human bone morphogenetic protein (hBMP)-2 gene (Group 1), Adv-beta gal transfected MSCs (Group 2) and uninfected MSCs (Group 3). Western blot analysis, alkaline phosphatase staining, Von Kossa staining and transmission electron microscopy were adopted to determine the phenotype of MSCs. Then the cells were injected into thigh muscles of the nude mice. Radiographical and histological evaluations were performed at different intervals. RESULTS: Only Adv-hBMP-2 transfected MSCs produced hBMP-2. These cells were positive for alkaline phosphatase staining at the 12th day and were positive for Von Kossa staining at the 16th day after gene transfer. Electron microscopic observation showed that there were more rough endoplasmic reticulum, mitochondria and lysosomes in Adv-hBMP-2 transfected MSCs compared to MSCs of other two groups. At the 3rd and 6th weeks after cell injection, ectopic bones were observed in muscles of nude mice of Group 1. Only fibrous tissue or a little bone was found in other two groups. CONCLUSIONS: BMP-2 gene transfected MSCs can differentiate into osteoblasts in vitro and induce bone formation in vivo.
Assuntos
Células da Medula Óssea/metabolismo , Proteínas Morfogenéticas Ósseas/genética , Terapia Genética , Células-Tronco Mesenquimais/metabolismo , Osteogênese/fisiologia , Engenharia Tecidual , Fator de Crescimento Transformador beta/genética , Animais , Western Blotting , Células da Medula Óssea/citologia , Proteína Morfogenética Óssea 2 , Diferenciação Celular , Cabras , Humanos , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Nus , Coloração e Rotulagem , TransfecçãoRESUMO
OBJECTIVE: To investigate the effects of bone morphogenetic protein-2 (BMP-2) gene therapy on the bone-implant interface in the reconstruction of periprosthetic bone defect. METHODS: Transverse defects were caused in the external condylae of both femurs of 14 adult Beagle dogs. Titanium alloy implants were inserted and a bone defect 3 mm wide around the titanium alloy implant was preserved. Then the total 28 defects were divided into 4 groups: 8 bone defects remained untreated (blank control group); 8 bone defects were implanted with heterogeneous freeze-dried bone by impaction grafting technique (non-cell group); 8 bone defects were implanted with heterogeneous freeze-dried bone loaded with autogenous bone marrow stromal cells (BMSCs) from the greater trochanter of the same dog (cell group); and 10 bone defects were implanted with freeze-dried allograft loaded with autogenous BMSCs from the greater trochanter of the same dog which were transfected by Adv-BMP-2 gene (gene group). Three, 6, and 12 weeks after implantation X-ray examination was carried out to observe the place of the implant and the absorption of the implants. Six and 12 weeks after the dogs were killed and their bone defects were taken out to undergo histological, histomorphometric and biomechanical examination to observe the healing and oseeointegration of the bone-implant interface. RESULTS: Histological examination showed that 6 weeks after implantation new bone formation was found on the implant surface and there was point contact between the bone and implant in the gene group with the bone-to-impact contact (BIC) of about 10%; and continuous soft tissue was found at bone-implant interface in all other groups. Twelve weeks after, there was thick soft tissue membrane between the new bone and implant in the blank control group; most of the interface was connective fibrous tissue in the non-cell group and cell group with point contact between the bone and implant and a BIC lower than 10%; and in the gene group the interface consisted mainly of bone tissue and continuous bone-implant contact was found with the BIC of 50%, significantly higher than those of the other 2 groups (both P < 0.01). The mechanical strength of interface increased time-dependently in all groups, that of the gene group being significantly higher than those of the other 2 groups at any time-points (both P < 0.01). CONCLUSION: BMP-2 gene therapy can improve the osseointegration of bone-implant interface.
Assuntos
Proteínas Morfogenéticas Ósseas/genética , Terapia Genética , Osseointegração/efeitos dos fármacos , Próteses e Implantes , Fator de Crescimento Transformador beta/genética , Animais , Células da Medula Óssea/citologia , Proteína Morfogenética Óssea 2 , Proteínas Morfogenéticas Ósseas/uso terapêutico , Regeneração Óssea/efeitos dos fármacos , Transplante Ósseo , Células Cultivadas , Cães , Fêmur/lesões , Implantes Experimentais , Masculino , Células Estromais/citologia , Fator de Crescimento Transformador beta/uso terapêuticoRESUMO
OBJECTIVE: To evaluate the effectiveness of the tissue-engineered bone substitute loaded with adenovirus mediated human bone morphogenetic protein-2 gene (Adv-hBMP-2) transfected bone marrow derived mesenchymal stem cells (BMSC) in the repair of diaphyseal segmental bone defect of large animal. METHODS: The right tibial bone defects (2.6 cm) model of 26 goats were established and divided into 5 groups: I. Adv-hBMP-2 transfected BMSC/calcined bone (CB) group (n = 9); II. adenovirus-beta-galactosidase (Adv-betagal) gene transfected BMSC/CB group (n = 6); III. untransfected BMSC/CB group (n = 6); IV. single CB group (n = 3); VI. untreated group (n = 2). The above tissue-engineered bone substitutes were implanted in the bone defects respectively except group VI. Roentgenography, histomorphometrical analysis and biomechanical measurement were studied at various times. RESULTS: X-ray: at 4 - 8th weeks after implantation, more bony callus was found in the bone defects of group I. The complete healing rates of group I, II, III, IV, and V were 5/8, 1/5, 0/5, 0/2, 0/1 respectively at 26th week after implantation. Histomorphometrical analysis showed much more new bony callus including cortical bone formed in group I than those of other groups. The compression strength of the implanted bone substitute of group I is significantly higher than those of group II and III. CONCLUSION: The tissue-engineered bone substitute loaded with human BMP-2 gene transfected BMSC can repair diaphyseal segmental bone defect of large animal (goat).
Assuntos
Doenças Ósseas/terapia , Proteínas Morfogenéticas Ósseas/genética , Terapia Genética , Transplante de Células-Tronco Mesenquimais , Tíbia , Engenharia Tecidual , Fator de Crescimento Transformador beta , Animais , Fenômenos Biomecânicos , Proteína Morfogenética Óssea 2 , Cabras , CicatrizaçãoRESUMO
Impaction bone allografting represents an attractive procedure for bone defects reconstruction in joint replacement. And it was found that bone morphogenetic protein-2(BMP-2) gene therapy can enhance bone healing. The purpose of this study was to determine if combined adenovirus mediated human BMP-2(Adv-hBMP-2) gene-modified bone marrow stromal cells(BMSCs) with allograft enhanced the defects healing and improved the strength of implant fixation in 3-mm bone defect around a titanium alloy implant. Using the impaction grafting technique, the defects were reconstructed using freeze-dried allograft, freeze-dried allografts loaded with autogenous BMSCs, or freeze-dried allografts loaded with autogenous BMSCs modified with the human bone morphogenetic protein-2 (hBMP-2) gene. At 6 and 12 weeks, the Bone-implant Contact rate and strength of the interface in the group with BMP-2 gene medication were significantly higher than those of the non-cell or cell groups. BMP-2 gene medication also showed significant effects on allograft healing and replacement compared with those of two other groups, as evidenced by increased new bone formation and reduced graft remnants. The results suggest that BMP-2 gene medication can enhance allograft healing and osseointegration of the bone-implant interface.