By combining coral with recombinant human bone morphogenetic protein-2 (rhBMP-2), rhBMP-2/coral composite was obtained in this study. Following implantation of the composite into the muscle pouches of mice, cartilage ...By combining coral with recombinant human bone morphogenetic protein-2 (rhBMP-2), rhBMP-2/coral composite was obtained in this study. Following implantation of the composite into the muscle pouches of mice, cartilage growth was induced in the pores or on the surface of the implants at one week, woven bone at three week and lamellar bone with bone marrow at six week, and coral was absorbed partially. The induced formation of endochondral bone was time-related and rhBMP-2 dose-related. The results of this study indicate that the composite possesses a superior ability of osteogenesis, and coral acts as one of the most suitable rhBMP-2 slowrelease carriers currently available. The composite will be a new type of bone substitute to be used in orthopaedics and maxillofacial surgery.展开更多
Objective To explore the feasibility and efficacy of lentivirus-mediated co-transfection of rat bone marrow mesenchymal stem cells （MSCs） with human vascular endothelial growth factor 165 （hVEGFI65） gene and human...Objective To explore the feasibility and efficacy of lentivirus-mediated co-transfection of rat bone marrow mesenchymal stem cells （MSCs） with human vascular endothelial growth factor 165 （hVEGFI65） gene and human bone morphogenetic protein 2 （hBMP2） gene. Methods The hVEGF165 and hBMP2 cDNAs were obtained from human osteosarcoma cell line MG63 and cloned into lentiviral expression vectors designed to co-express the copepod green fluorescent protein （copGFP）. The expression lentivector and packaging Plasmid Mix were co-transferred to 293TN cells, which produced the lentivirus carrying hVEGF165 （Lv-VEGF） or hBMP2 （ Lv-BMP） , respectively. MSCs of Wistar rats were co-transfected with Lv-BMP and Lv-VEGF （BMP ＋ VEGF group）, or each alone （BMP group and VEGF group）, or with no virus （ Control group）. The mRNA and protein expressions of hVEGF165 and hBMP2 genes in each group were detected by real-time PCR and enzyme linked immunosorbent assay （ELISA）. Results Lentiviral expression vectors carrying hVEGF165 or hBMP2 were correctly constructed and confirmed by restriction endonucleses analysis and DNA sequencing analysis. A transfer efficiency up to 90% was archieved in all the transfected groups detected by the fraction of fluorescent cells using fluorescent microscopy. From the results generated by real-time PCR and ELISA, VEGF165 and BMP2 genes were co-expressed in BMP ＋ VEGF group. No significant difference of BMP2 expression was detected between BMP ＋ VEGF and BMP groups （ P 〉 0. 05）. Similarly, there was no significant difference of VEGF165 expression between BMP ＋ VEGF and VEGF groups （ P 〉 0. 05）. Conclusion VEGF165 and BMP2 genes were successfully co-expressed in MSCs by lentivirus-mediated co-transfection, which provided a further foundation for the combined gene therapy of bone regeneration.展开更多
Objective: To identify eukaryotic expression vector of human bone morphogenetic protein 2 pcDNA3/BMP2, verify its expression in transfected human mesenchymal stem cells (hMSCs) and the effect on hMSCs differentiation....Objective: To identify eukaryotic expression vector of human bone morphogenetic protein 2 pcDNA3/BMP2, verify its expression in transfected human mesenchymal stem cells (hMSCs) and the effect on hMSCs differentiation. Methods: The BMP2 gene was cloned into a eukaryotic expression vector pcDNA3. Transfected the recombinant into hMSCs by liposome. Immunnohistochemistry and in situ hybridization methods were used to identify the expression of BMP2 mRNA and protein; ALP and Von Kossa stains were performed to identify the BMP2 gene differentiated effect on the hMSCs. Results: The pcDNA3/BMP2 fragments were as large as theory. BMP2 mRNA and protein were expressed and synthesized both in 48 h and 4 weeks after transfection, the ALP and Ca deposit exhibition, which marked the osteogenic lineage of hMSCs, were enhanced and sped. Conclusion: Transfection of pcDNA3/BMP2 is able to provide transient and persistent expression in hMSCs, and promote the MSCs differentiation to osteogenic lineage.展开更多
Objective:To observe the clinical efficacy of heat-sensitive moxibustion plus medications on senile osteoporosis(SOP),and to explore the related mechanisms.Methods:A total of 70 elderly participants with osteoporosis ...Objective:To observe the clinical efficacy of heat-sensitive moxibustion plus medications on senile osteoporosis(SOP),and to explore the related mechanisms.Methods:A total of 70 elderly participants with osteoporosis were randomly divided into an observation group and control group,with 35 cases in each group.The control group was treated with conventional drugs,and the observation group was treated with heat-sensitive moxibustion on the basis of the conventional drugs.Both groups were treated for 3 months.Before and after treatment,assessed the visual analog scale(VAS)and Oswestry disability index(ODI)scores,determined the bone mineral density of the participants'lumbar spine(L2-L4)and left femoral neck,and detected the participants'serum bone morphogenetic protein-2(BMP-2)and osteoprotegerin(OPG)levels.Results:After treatment,the VAS scores of both groups were lower than before treatment(both P<0.05),and the VAS score of the observation group was significantly lower than that of the control group(P<0.05).After treatment,the bone mineral density values of the lumbar spine and left femoral neck in both groups were significantly higher than before treatment(both P<0.05),and the bone mineral density values of the observation group were higher than those of the control group(P<0.05).After treatment,the ODI scores of the two groups were lower than those before treatment(both P<0.05),and the ODI score of the observation group was lower than that of the control group(P<0.05).After treatment,the serum BMP-2 and OPG levels in the observation group were significantly higher than those in the control group(both P<0.05).Conclusion:Heat-sensitive moxibustion plus medications for SOP can significantly relieve patients'pain,improve dysfunction,and increase bone density,which may be related to the improvement of the serum BMP-2 and OPG levels.展开更多
Objective: To investigate the effect of nano hydroxyapatite/collagen (nHA/collagen) composite as a graft extender and enhancer when combined with recombinant human bone morphogenetic protein 2 (rhBMP 2) on lumbar inte...Objective: To investigate the effect of nano hydroxyapatite/collagen (nHA/collagen) composite as a graft extender and enhancer when combined with recombinant human bone morphogenetic protein 2 (rhBMP 2) on lumbar intertransverse fusion in rabbits. Methods: Sixty four adult female New Zealand white rabbits, aged 1 year and weighing 3.5 4.5 kg, underwent similar posterolateral intertransverse process arthrodesis and were randomly divided into 4 groups based on different grafts: autogenous cancellous bone alone (ACB group), nHA/collagen alone (HAC group), half autogenous cancellous bone and half nHA/collagen (ACB+HAC group) and nHA/collagen combined with rhBMP 2 (HAC+BMP group). The fusion masses were analyzed by manual palpation, radiography, biomechanical testing and histological examination. Results: Fusion was observed in 4 cases in the 6th week and in 5 cases in the 10th week after surgery in ACB group. No case showed fusion in HAC group. In ACB+HAC group, there was fusion in 3 cases in the 6th week and in 4 cases in the 10th week after surgery. In HAC+BMP group, fusion in 1 case was found in the 4th week, in 5 cases in the 6th week and in 6 cases in the 10th week after surgery. It suggested that ACB, ACB+HAC and HAC+BMP groups showed similar fusion ratio and mechanical strength in the 6th and 10th week after surgery. According to the microstructure analysis of the samples, nHA/collagen had no negative effect when implanted together with ilium autograft. In HAC+BMP group, new bone like tissue was observed in the 2nd week postoperatively, and nearly all of the implanted composites were replaced by mature bone matrix and new bones in 10th week postoperatively. Conclusions: The nHA/collagen, especially combined with rhBMP 2, is a promising bone substitute, for it has quick biodegradation, fine bone bending ability, and high osteoconductivity on posterolateral spinal fusion in rabbits.展开更多
The method of plasma-spray coating of hy- droxyapatite （HA） onto pure titanium has been demon- strated to be effective to enhance the osteogenic differentiation and accelerate bone regeneration. Yet it is still a bi...The method of plasma-spray coating of hy- droxyapatite （HA） onto pure titanium has been demon- strated to be effective to enhance the osteogenic differentiation and accelerate bone regeneration. Yet it is still a big challenge to figure out the interplay among im- plant surface properties, adsorbed proteins and cell-surface interactions. In this study, the plasma-sprayed HA-coated titanium （HA-Ti） surface was compared with the titanium substrate in terms of protein adsorption, cell adhesion and differentiation. The phase composition, wettability and to- pography were characterized. Compared to the Ti substrate, the HA-Ti had a smaller water contact angle, but larger micro-scale roughness, and showed a poorer ability to ad- sorb fibronectin （Fn）, bovine serum albumin （BSA） and serum proteins. However, it could adsorb larger amount of recombinant human bone morphogenetic protein 2 （BMP- 2）. The osteoblasts and bone marrow mesenchymal stem cells （BMSCs） tended to adhere on the Ti substrate. By contrast, the BMSCs cultured on the HA-Ti showed a stronger tendency toward osteogenesis differentiation.展开更多
OBJECTIVE:This study was conducted to evaluate the effect of Amomum villosum on longitudinal bone growth.METHODS:Adolescent female Sprague-Dawley rats were divided into 3 groups and treated for 4 days:control(distille...OBJECTIVE:This study was conducted to evaluate the effect of Amomum villosum on longitudinal bone growth.METHODS:Adolescent female Sprague-Dawley rats were divided into 3 groups and treated for 4 days:control(distilled water,p.o.),recombinant human growth hormone(rhGH;100 μg/kg,s.c.),and A.villosum(500 mg/kg,p.o.) groups.On day 3,tetracycline(20 g/kg,i.p.) was injected for growth plate identification.On days 2,3 and 4,5-bromo-2'-deoxyuridine(BrdU)(50 mg/kg,i.p.) was injected to label proliferating cells.On day 5,tibias were dissected and fixed in 4% paraformaldehyde,dehydrated,and sectioned for immunohistochemistry and histomorphometry.RESULTS:The rate of bone growth in the A.villosum and rhGH groups increased to(410 ± 44) and(389 ± 46) μm/day(P<0.01),respectively,as compared with the control(330.7 ± 34.7) μm/day.The thickness of the growth plates also increased to(591 ± 37) and(598 ± 32) μm,respectively,as compared with the control(524± 89) μm(P<0.001).The number of BrdU-positive cells in the chondrocytes of the A.villosum and rhGH groups was also significantly higher(126±24) and(143±18) cells/mm 2,respectively) than in the control(109 ± 25) mm 2(P<0.05).Insulin-like growth factor-1 and bone morphogenetic protein-2 in the A.villosum and rhGH groups were highly expressed in the growth plate as compared with the control samples,indicating increased bone formation.CONCLUSIONS:A.villosum could be used to treat growth retardation during adolescence.展开更多
Objective： To investigate the effect of a new biomaterial combining calcium citrate and recombinant human bone morphogenetic protein-2 （rhBMP-2） on bone regeneration in a bone defect rabbit model. Methods： Totall...Objective： To investigate the effect of a new biomaterial combining calcium citrate and recombinant human bone morphogenetic protein-2 （rhBMP-2） on bone regeneration in a bone defect rabbit model. Methods： Totally 30 male New Zealand white rabbits were randomly and equally divided into calcium citraterhBMP-2 （CC-rhBMP-2） group and rhBMP-2 only group. Two 10 ram-long and 5 ram-deep bone defects were respectively created in the left and right femoral condyles of the rabbits. Subsequently 5 pellets of calcium citrate （10 mg） combined with rhBMP-2 （2 rag） or rhBMP-2 alone were implanted into the bone defects and compressed with cotton swab. Bone granules were obtained at 2, 4 and 6 weeks after procedure and received histological analysis. LSD t-test and a subsequent t-test were adopted for statistical analysis. Results： Histomorphometric analysis revealed newlyformed bones, and calcium citrate has been absorbed in the treatment group. The percent of newly formed bone area in femoral condyle in control group and CC-rhBMP-2 group was respectively 31.73%±1.26% vs 48.21%±2.37% at 2 weeks; 43.40%±1.65% vs 57.32%±1.47% at 4 weeks, and 51.32%±7.80% vs 66.74%±4.05% at 6 weeks （P〈0.05 for all）. At 2 weeks, mature cancellous bone was observed to be already formed in the treatment group. Conclusion： From this study, it can be concluded that calcium citrate combined with rhBMP-2 signifcantly enhances bone regeneration in bone defects. This synthetic gelatin matrix stimulates formation of new bone and bone marrow in the defect areas by releasing calcium ions.展开更多
文摘By combining coral with recombinant human bone morphogenetic protein-2 (rhBMP-2), rhBMP-2/coral composite was obtained in this study. Following implantation of the composite into the muscle pouches of mice, cartilage growth was induced in the pores or on the surface of the implants at one week, woven bone at three week and lamellar bone with bone marrow at six week, and coral was absorbed partially. The induced formation of endochondral bone was time-related and rhBMP-2 dose-related. The results of this study indicate that the composite possesses a superior ability of osteogenesis, and coral acts as one of the most suitable rhBMP-2 slowrelease carriers currently available. The composite will be a new type of bone substitute to be used in orthopaedics and maxillofacial surgery.
基金Supported by Key Program of Shanghai Science and Technology Committee (054119520)
文摘Objective To explore the feasibility and efficacy of lentivirus-mediated co-transfection of rat bone marrow mesenchymal stem cells （MSCs） with human vascular endothelial growth factor 165 （hVEGFI65） gene and human bone morphogenetic protein 2 （hBMP2） gene. Methods The hVEGF165 and hBMP2 cDNAs were obtained from human osteosarcoma cell line MG63 and cloned into lentiviral expression vectors designed to co-express the copepod green fluorescent protein （copGFP）. The expression lentivector and packaging Plasmid Mix were co-transferred to 293TN cells, which produced the lentivirus carrying hVEGF165 （Lv-VEGF） or hBMP2 （ Lv-BMP） , respectively. MSCs of Wistar rats were co-transfected with Lv-BMP and Lv-VEGF （BMP ＋ VEGF group）, or each alone （BMP group and VEGF group）, or with no virus （ Control group）. The mRNA and protein expressions of hVEGF165 and hBMP2 genes in each group were detected by real-time PCR and enzyme linked immunosorbent assay （ELISA）. Results Lentiviral expression vectors carrying hVEGF165 or hBMP2 were correctly constructed and confirmed by restriction endonucleses analysis and DNA sequencing analysis. A transfer efficiency up to 90% was archieved in all the transfected groups detected by the fraction of fluorescent cells using fluorescent microscopy. From the results generated by real-time PCR and ELISA, VEGF165 and BMP2 genes were co-expressed in BMP ＋ VEGF group. No significant difference of BMP2 expression was detected between BMP ＋ VEGF and BMP groups （ P 〉 0. 05）. Similarly, there was no significant difference of VEGF165 expression between BMP ＋ VEGF and VEGF groups （ P 〉 0. 05）. Conclusion VEGF165 and BMP2 genes were successfully co-expressed in MSCs by lentivirus-mediated co-transfection, which provided a further foundation for the combined gene therapy of bone regeneration.
文摘Objective: To identify eukaryotic expression vector of human bone morphogenetic protein 2 pcDNA3/BMP2, verify its expression in transfected human mesenchymal stem cells (hMSCs) and the effect on hMSCs differentiation. Methods: The BMP2 gene was cloned into a eukaryotic expression vector pcDNA3. Transfected the recombinant into hMSCs by liposome. Immunnohistochemistry and in situ hybridization methods were used to identify the expression of BMP2 mRNA and protein; ALP and Von Kossa stains were performed to identify the BMP2 gene differentiated effect on the hMSCs. Results: The pcDNA3/BMP2 fragments were as large as theory. BMP2 mRNA and protein were expressed and synthesized both in 48 h and 4 weeks after transfection, the ALP and Ca deposit exhibition, which marked the osteogenic lineage of hMSCs, were enhanced and sped. Conclusion: Transfection of pcDNA3/BMP2 is able to provide transient and persistent expression in hMSCs, and promote the MSCs differentiation to osteogenic lineage.
文摘Objective:To observe the clinical efficacy of heat-sensitive moxibustion plus medications on senile osteoporosis(SOP),and to explore the related mechanisms.Methods:A total of 70 elderly participants with osteoporosis were randomly divided into an observation group and control group,with 35 cases in each group.The control group was treated with conventional drugs,and the observation group was treated with heat-sensitive moxibustion on the basis of the conventional drugs.Both groups were treated for 3 months.Before and after treatment,assessed the visual analog scale(VAS)and Oswestry disability index(ODI)scores,determined the bone mineral density of the participants'lumbar spine(L2-L4)and left femoral neck,and detected the participants'serum bone morphogenetic protein-2(BMP-2)and osteoprotegerin(OPG)levels.Results:After treatment,the VAS scores of both groups were lower than before treatment(both P<0.05),and the VAS score of the observation group was significantly lower than that of the control group(P<0.05).After treatment,the bone mineral density values of the lumbar spine and left femoral neck in both groups were significantly higher than before treatment(both P<0.05),and the bone mineral density values of the observation group were higher than those of the control group(P<0.05).After treatment,the ODI scores of the two groups were lower than those before treatment(both P<0.05),and the ODI score of the observation group was lower than that of the control group(P<0.05).After treatment,the serum BMP-2 and OPG levels in the observation group were significantly higher than those in the control group(both P<0.05).Conclusion:Heat-sensitive moxibustion plus medications for SOP can significantly relieve patients'pain,improve dysfunction,and increase bone density,which may be related to the improvement of the serum BMP-2 and OPG levels.
文摘Objective: To investigate the effect of nano hydroxyapatite/collagen (nHA/collagen) composite as a graft extender and enhancer when combined with recombinant human bone morphogenetic protein 2 (rhBMP 2) on lumbar intertransverse fusion in rabbits. Methods: Sixty four adult female New Zealand white rabbits, aged 1 year and weighing 3.5 4.5 kg, underwent similar posterolateral intertransverse process arthrodesis and were randomly divided into 4 groups based on different grafts: autogenous cancellous bone alone (ACB group), nHA/collagen alone (HAC group), half autogenous cancellous bone and half nHA/collagen (ACB+HAC group) and nHA/collagen combined with rhBMP 2 (HAC+BMP group). The fusion masses were analyzed by manual palpation, radiography, biomechanical testing and histological examination. Results: Fusion was observed in 4 cases in the 6th week and in 5 cases in the 10th week after surgery in ACB group. No case showed fusion in HAC group. In ACB+HAC group, there was fusion in 3 cases in the 6th week and in 4 cases in the 10th week after surgery. In HAC+BMP group, fusion in 1 case was found in the 4th week, in 5 cases in the 6th week and in 6 cases in the 10th week after surgery. It suggested that ACB, ACB+HAC and HAC+BMP groups showed similar fusion ratio and mechanical strength in the 6th and 10th week after surgery. According to the microstructure analysis of the samples, nHA/collagen had no negative effect when implanted together with ilium autograft. In HAC+BMP group, new bone like tissue was observed in the 2nd week postoperatively, and nearly all of the implanted composites were replaced by mature bone matrix and new bones in 10th week postoperatively. Conclusions: The nHA/collagen, especially combined with rhBMP 2, is a promising bone substitute, for it has quick biodegradation, fine bone bending ability, and high osteoconductivity on posterolateral spinal fusion in rabbits.
基金supported by the National Basic Research Program of China(2011CB606203)the National Natural Science Foundation of China(21434006,21374097)
文摘The method of plasma-spray coating of hy- droxyapatite （HA） onto pure titanium has been demon- strated to be effective to enhance the osteogenic differentiation and accelerate bone regeneration. Yet it is still a big challenge to figure out the interplay among im- plant surface properties, adsorbed proteins and cell-surface interactions. In this study, the plasma-sprayed HA-coated titanium （HA-Ti） surface was compared with the titanium substrate in terms of protein adsorption, cell adhesion and differentiation. The phase composition, wettability and to- pography were characterized. Compared to the Ti substrate, the HA-Ti had a smaller water contact angle, but larger micro-scale roughness, and showed a poorer ability to ad- sorb fibronectin （Fn）, bovine serum albumin （BSA） and serum proteins. However, it could adsorb larger amount of recombinant human bone morphogenetic protein 2 （BMP- 2）. The osteoblasts and bone marrow mesenchymal stem cells （BMSCs） tended to adhere on the Ti substrate. By contrast, the BMSCs cultured on the HA-Ti showed a stronger tendency toward osteogenesis differentiation.
基金Supported by the Fund of Amomum villosum in bone growth
文摘OBJECTIVE:This study was conducted to evaluate the effect of Amomum villosum on longitudinal bone growth.METHODS:Adolescent female Sprague-Dawley rats were divided into 3 groups and treated for 4 days:control(distilled water,p.o.),recombinant human growth hormone(rhGH;100 μg/kg,s.c.),and A.villosum(500 mg/kg,p.o.) groups.On day 3,tetracycline(20 g/kg,i.p.) was injected for growth plate identification.On days 2,3 and 4,5-bromo-2'-deoxyuridine(BrdU)(50 mg/kg,i.p.) was injected to label proliferating cells.On day 5,tibias were dissected and fixed in 4% paraformaldehyde,dehydrated,and sectioned for immunohistochemistry and histomorphometry.RESULTS:The rate of bone growth in the A.villosum and rhGH groups increased to(410 ± 44) and(389 ± 46) μm/day(P<0.01),respectively,as compared with the control(330.7 ± 34.7) μm/day.The thickness of the growth plates also increased to(591 ± 37) and(598 ± 32) μm,respectively,as compared with the control(524± 89) μm(P<0.001).The number of BrdU-positive cells in the chondrocytes of the A.villosum and rhGH groups was also significantly higher(126±24) and(143±18) cells/mm 2,respectively) than in the control(109 ± 25) mm 2(P<0.05).Insulin-like growth factor-1 and bone morphogenetic protein-2 in the A.villosum and rhGH groups were highly expressed in the growth plate as compared with the control samples,indicating increased bone formation.CONCLUSIONS:A.villosum could be used to treat growth retardation during adolescence.
文摘Objective： To investigate the effect of a new biomaterial combining calcium citrate and recombinant human bone morphogenetic protein-2 （rhBMP-2） on bone regeneration in a bone defect rabbit model. Methods： Totally 30 male New Zealand white rabbits were randomly and equally divided into calcium citraterhBMP-2 （CC-rhBMP-2） group and rhBMP-2 only group. Two 10 ram-long and 5 ram-deep bone defects were respectively created in the left and right femoral condyles of the rabbits. Subsequently 5 pellets of calcium citrate （10 mg） combined with rhBMP-2 （2 rag） or rhBMP-2 alone were implanted into the bone defects and compressed with cotton swab. Bone granules were obtained at 2, 4 and 6 weeks after procedure and received histological analysis. LSD t-test and a subsequent t-test were adopted for statistical analysis. Results： Histomorphometric analysis revealed newlyformed bones, and calcium citrate has been absorbed in the treatment group. The percent of newly formed bone area in femoral condyle in control group and CC-rhBMP-2 group was respectively 31.73%±1.26% vs 48.21%±2.37% at 2 weeks; 43.40%±1.65% vs 57.32%±1.47% at 4 weeks, and 51.32%±7.80% vs 66.74%±4.05% at 6 weeks （P〈0.05 for all）. At 2 weeks, mature cancellous bone was observed to be already formed in the treatment group. Conclusion： From this study, it can be concluded that calcium citrate combined with rhBMP-2 signifcantly enhances bone regeneration in bone defects. This synthetic gelatin matrix stimulates formation of new bone and bone marrow in the defect areas by releasing calcium ions.
