Oestrogen is essential for maintaining bone mass, and it has been demonstrated to induce osteoblast proliferation and bone formation.In this study, complementary DNA（cDNA） microarrays were used to identify and study...Oestrogen is essential for maintaining bone mass, and it has been demonstrated to induce osteoblast proliferation and bone formation.In this study, complementary DNA（cDNA） microarrays were used to identify and study the expression of novel genes that may be involved in MC3T3-E1 cells’ response to 17-b estradiol. MC3T3-E1 cells were inoculated in minimum essential media alpha（a-MEM）cell culture supplemented with 17-b estradiol at different concentrations and for different time periods. MC3T3-E1 cells treated with1028mol？L2117-b estradiol for 5 days exhibited the highest proliferation and alkaline phosphatase（ALP） activity; thus, this group was chosen for microarray analysis. The harvested RNA was used for microarray hybridisation and subsequent real-time reverse transcription polymerase chain reaction（RT-PCR） to validate the expression levels for selected genes. The microarray results were analysed using both functional and pathway analysis. In this study, microarray analysis detected 5 403 differentially expressed genes,of which 1 996 genes were upregulated and 3 407 genes were downregulated, 1 553 different functional classifications were identified by gene ontology（GO） analysis and 53 different pathways were involved based on pathway analysis. Among the differentially expressed genes, a portion not previously reported to be associated with the osteoblast response to oestrogen was identified. These findings clearly demonstrate that the expression of genes related to osteoblast proliferation, cell differentiation, collagens and transforming growth factor beta（TGF-b）-related cytokines increases, while the expression of genes related to apoptosis and osteoclast differentiation decreases, following the exposure of MC3T3-E1 cells to a-MEM supplemented with 17-b estradiol. Microarray analysis with functional gene classification is critical for a complete understanding of complementary intracellular processes. This microarray analysis provides large-scale gene expression data that require further confirmatory studies.展开更多
The reinforcement of calcium phosphate materials with silk fibroin （SF） has been one of the strategies to overcome the brittleness. However, the lack of osteoinductivity may still restrict their further use. This st...The reinforcement of calcium phosphate materials with silk fibroin （SF） has been one of the strategies to overcome the brittleness. However, the lack of osteoinductivity may still restrict their further use. This study aimed to investigate the biocompatibility and osteogenesis capacity of a novel Semaphorin 3A-loaded chitosan microspheres/SF/a-tricalcium phosphate composite （Sema3A CMs/SF/a-TCP） in vitro. Sema3A was first incorporated into CMs, and the Sema3A CMs/SF/a-TCP composite was then prepared. The morphology of the CMs was observed using SEM. The in vitro release kinetics, cytotoxicity, and cell compatibility were evaluated, and the real-time quantitative polymerase chain reaction （RT-qPCR） and activity of alkaline phosphatase （ALP） were used to evaluate the osteogenesis capacity of the composite. The in vitro release of Sema3A from the Sema3A CMs/SF/a-TCP composite showed a temporally controlled manner. The extract of the Sema3A CMs/SF/a-TCP composite presented no obvious side effect on the MC3T3-E1 cell proliferation, nor promote cell proliferation. The MC3T3-E1 cells were well-spread and presented an elongated shape on the Sema3A CMs/SF/a-TCP composite surface; the ALP activity and the osteogenic-related gene expression were higher than those seeded on the surface of the CMs/SF/a-TCP and SF/a-TCP composites. In conclusion, Sema3A CMslSF/a-TCP has excellent biocompatibility and contributes to the osteoblastic differentiation of MC3T3-E1 cells.展开更多
Carbon/carbon （C/C） composites were deposited with graphite-like carbon （GLC） coating, and then, Arg-Gly- Asp acid （RGD） peptides were successfully immobilized onto the functionalized GLC coating. GLC coating wa...Carbon/carbon （C/C） composites were deposited with graphite-like carbon （GLC） coating, and then, Arg-Gly- Asp acid （RGD） peptides were successfully immobilized onto the functionalized GLC coating. GLC coating was utilized to prevent carbon particles releasing and create a uniform surface condition for C/C composites. RGD peptides were utilized to improve biocompatibility of GLC coating. Surface chemical characterizations of functionalized GLC coating were detected by contact angle measurement, X-ray photoelectron spectroscopy and Raman spectra. Optical morphology of GLC coatings was observed by confocal laser scanning microscopy. In vitro biological performance was determined using samples seeded with MC3T3-E1 osteoblast-like cells and cultured for 1 week. Surface characterizations and morphological analysis indicated that C/C composites were covered by a dense and uniform GLC coating. Contact angle of GLC coating was reduced to 27.2° when it was functionalized by H202 oxidation at 40 ℃ for 1 h. In vitro cytological test showed that the RGD peptides immobilized GLC coating had a significant improvement in biocompatibility. It was suggested that RGD peptides provided GLC coating with a bioactive surface to improve cell adhesion and proliferation on C/C composites.展开更多
Eucommia ulmoides Oliv.(EuO),also known as Duzhong,native to China,has been reported to have antioxidative function,but its cellular mechanism is not fully examined yet.We investigated inhibitory effects of EuO leaf e...Eucommia ulmoides Oliv.(EuO),also known as Duzhong,native to China,has been reported to have antioxidative function,but its cellular mechanism is not fully examined yet.We investigated inhibitory effects of EuO leaf ethanol extracts on H2O2-induced apoptosis in rat osteoblastic MC3T3-E1 cells and underlying mechanisms.Locally-grown Duzhong leaves were extracted with ethanol.MC3T3-E1 cells were treated with EuO (6.25,12.5,25,50,and 100 μg/ml) for 24 h,and then H2O2 (800 μmol/L) for an additional 24 h.Cell survival rate,percentage of apoptosis,and expressions of caspases 3,6,7,and 9 were examined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay,microscopic analysis,Western blotting,and reverse transcription polymerase chain reaction (RT-PCR).The final EuO leaf ethanol extract powder was detected to contain caffeotannic acid at 58 mg/g and geniposide at 3.45 mg/g by high performance liquid chromatography (HPLC).EuO remarkably restrained cell oxidative damage and increased cell survival rate in a dose-dependent manner: 0 μg/ml,0.21;6.25 μg/ml,0.28;12.5 μg/ml,0.31;25 μg/ml,0.48;50 μg/ml,0.54;and 100 μg/ml,0.66 (P<0.05),with the half-effective concentration being around 25 μg/ml.MTT results were confirmed by microscopic analysis.Western blotting and RT-PCR analyses showed that the expressions of caspases 3,6,7,and 9 were significantly decreased in the EuO-treated cells compared with the control (EuOand H2O2-free) (P<0.05),with the half-effective concentration of EuO ranging from 12.5 to 25 μg/ml.We conclude that the ethanol-extracted EuO leaf extracts promoted the growth of MC3T3-E1 cells,and suppressed the H2O2-induced apoptosis in a rat MC3T3-E1 osteogenic cell model,likely due to the inhibition of caspases’ activities.The results indicate that EuO is a potent antioxidant,which may contribute to its many cellular protective functions,including the promotion of bone growth.展开更多
The purpose of this study was to test the hypothesis that the combination of micro-arc oxidation and alkali heatment （MAH） would improve the cytocompatibility of a newly designed Ti-24Nb-4Zr-8Sn alloy. In this study...The purpose of this study was to test the hypothesis that the combination of micro-arc oxidation and alkali heatment （MAH） would improve the cytocompatibility of a newly designed Ti-24Nb-4Zr-8Sn alloy. In this study, commercially pure titanium （cp Ti） and Ti-24Nb-4Zr-8Sn were used. Surface modification of Ti-24Nb- 4Zr-8Sn by a two-step treatment of micro-arc oxidation （MAO） and alkali heatment was reported. Surface characterizations were performed by scanning electron microscopy （SEM）, thin film X-ray diffraction （TF-XRD） and X-ray photoelectron spectroscopy （XPS）. The MAH layer consisted of finer crystals and possessed a higher degree of crystallity and stability than the MAO layer. A biocompatibility study on treated and untreated Ti- 24Nb-4Zr-8Sn in comparison with cp Ti was carried out to investigate the effect of the different surfaces on the bone integration property in vitro. The cellular assays revealed that the MAO and MAH layer favored the initial adhesion of MC3T3-E1 cells and that the growth rate of MC3T3-E1 cells on MAH layer was significantly higher than that on the conventional MAO-treated layer after 3-day and 5-day incubation, demonstrating the greater potential of the hybrid treatment of micro-arc oxidation followed with alkali heatment as a novel surface modification method for implanting materials.展开更多
基金supported by grants from the Natural Science Fund (ZR2010HM035) of Shandong Provincethe Shandong Provincial Health Development Project Fund (2011WSB19002) in China
文摘Oestrogen is essential for maintaining bone mass, and it has been demonstrated to induce osteoblast proliferation and bone formation.In this study, complementary DNA（cDNA） microarrays were used to identify and study the expression of novel genes that may be involved in MC3T3-E1 cells’ response to 17-b estradiol. MC3T3-E1 cells were inoculated in minimum essential media alpha（a-MEM）cell culture supplemented with 17-b estradiol at different concentrations and for different time periods. MC3T3-E1 cells treated with1028mol？L2117-b estradiol for 5 days exhibited the highest proliferation and alkaline phosphatase（ALP） activity; thus, this group was chosen for microarray analysis. The harvested RNA was used for microarray hybridisation and subsequent real-time reverse transcription polymerase chain reaction（RT-PCR） to validate the expression levels for selected genes. The microarray results were analysed using both functional and pathway analysis. In this study, microarray analysis detected 5 403 differentially expressed genes,of which 1 996 genes were upregulated and 3 407 genes were downregulated, 1 553 different functional classifications were identified by gene ontology（GO） analysis and 53 different pathways were involved based on pathway analysis. Among the differentially expressed genes, a portion not previously reported to be associated with the osteoblast response to oestrogen was identified. These findings clearly demonstrate that the expression of genes related to osteoblast proliferation, cell differentiation, collagens and transforming growth factor beta（TGF-b）-related cytokines increases, while the expression of genes related to apoptosis and osteoclast differentiation decreases, following the exposure of MC3T3-E1 cells to a-MEM supplemented with 17-b estradiol. Microarray analysis with functional gene classification is critical for a complete understanding of complementary intracellular processes. This microarray analysis provides large-scale gene expression data that require further confirmatory studies.
文摘The reinforcement of calcium phosphate materials with silk fibroin （SF） has been one of the strategies to overcome the brittleness. However, the lack of osteoinductivity may still restrict their further use. This study aimed to investigate the biocompatibility and osteogenesis capacity of a novel Semaphorin 3A-loaded chitosan microspheres/SF/a-tricalcium phosphate composite （Sema3A CMs/SF/a-TCP） in vitro. Sema3A was first incorporated into CMs, and the Sema3A CMs/SF/a-TCP composite was then prepared. The morphology of the CMs was observed using SEM. The in vitro release kinetics, cytotoxicity, and cell compatibility were evaluated, and the real-time quantitative polymerase chain reaction （RT-qPCR） and activity of alkaline phosphatase （ALP） were used to evaluate the osteogenesis capacity of the composite. The in vitro release of Sema3A from the Sema3A CMs/SF/a-TCP composite showed a temporally controlled manner. The extract of the Sema3A CMs/SF/a-TCP composite presented no obvious side effect on the MC3T3-E1 cell proliferation, nor promote cell proliferation. The MC3T3-E1 cells were well-spread and presented an elongated shape on the Sema3A CMs/SF/a-TCP composite surface; the ALP activity and the osteogenic-related gene expression were higher than those seeded on the surface of the CMs/SF/a-TCP and SF/a-TCP composites. In conclusion, Sema3A CMslSF/a-TCP has excellent biocompatibility and contributes to the osteoblastic differentiation of MC3T3-E1 cells.
基金supported by the National Natural Science Foundation of China under Grant Nos.51202194 and 51221001the Programme of Introducing Talents of Discipline to Universities(‘‘111’’project of China)under Grant No.B08040
文摘Carbon/carbon （C/C） composites were deposited with graphite-like carbon （GLC） coating, and then, Arg-Gly- Asp acid （RGD） peptides were successfully immobilized onto the functionalized GLC coating. GLC coating was utilized to prevent carbon particles releasing and create a uniform surface condition for C/C composites. RGD peptides were utilized to improve biocompatibility of GLC coating. Surface chemical characterizations of functionalized GLC coating were detected by contact angle measurement, X-ray photoelectron spectroscopy and Raman spectra. Optical morphology of GLC coatings was observed by confocal laser scanning microscopy. In vitro biological performance was determined using samples seeded with MC3T3-E1 osteoblast-like cells and cultured for 1 week. Surface characterizations and morphological analysis indicated that C/C composites were covered by a dense and uniform GLC coating. Contact angle of GLC coating was reduced to 27.2° when it was functionalized by H202 oxidation at 40 ℃ for 1 h. In vitro cytological test showed that the RGD peptides immobilized GLC coating had a significant improvement in biocompatibility. It was suggested that RGD peptides provided GLC coating with a bioactive surface to improve cell adhesion and proliferation on C/C composites.
基金Project (No.2007C33030) supported by the Science and Technology Program of Zhejiang Province,China
文摘Eucommia ulmoides Oliv.(EuO),also known as Duzhong,native to China,has been reported to have antioxidative function,but its cellular mechanism is not fully examined yet.We investigated inhibitory effects of EuO leaf ethanol extracts on H2O2-induced apoptosis in rat osteoblastic MC3T3-E1 cells and underlying mechanisms.Locally-grown Duzhong leaves were extracted with ethanol.MC3T3-E1 cells were treated with EuO (6.25,12.5,25,50,and 100 μg/ml) for 24 h,and then H2O2 (800 μmol/L) for an additional 24 h.Cell survival rate,percentage of apoptosis,and expressions of caspases 3,6,7,and 9 were examined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay,microscopic analysis,Western blotting,and reverse transcription polymerase chain reaction (RT-PCR).The final EuO leaf ethanol extract powder was detected to contain caffeotannic acid at 58 mg/g and geniposide at 3.45 mg/g by high performance liquid chromatography (HPLC).EuO remarkably restrained cell oxidative damage and increased cell survival rate in a dose-dependent manner: 0 μg/ml,0.21;6.25 μg/ml,0.28;12.5 μg/ml,0.31;25 μg/ml,0.48;50 μg/ml,0.54;and 100 μg/ml,0.66 (P<0.05),with the half-effective concentration being around 25 μg/ml.MTT results were confirmed by microscopic analysis.Western blotting and RT-PCR analyses showed that the expressions of caspases 3,6,7,and 9 were significantly decreased in the EuO-treated cells compared with the control (EuOand H2O2-free) (P<0.05),with the half-effective concentration of EuO ranging from 12.5 to 25 μg/ml.We conclude that the ethanol-extracted EuO leaf extracts promoted the growth of MC3T3-E1 cells,and suppressed the H2O2-induced apoptosis in a rat MC3T3-E1 osteogenic cell model,likely due to the inhibition of caspases’ activities.The results indicate that EuO is a potent antioxidant,which may contribute to its many cellular protective functions,including the promotion of bone growth.
基金supported by the National Natural Science Foundation of China under grant No. 30973354
文摘The purpose of this study was to test the hypothesis that the combination of micro-arc oxidation and alkali heatment （MAH） would improve the cytocompatibility of a newly designed Ti-24Nb-4Zr-8Sn alloy. In this study, commercially pure titanium （cp Ti） and Ti-24Nb-4Zr-8Sn were used. Surface modification of Ti-24Nb- 4Zr-8Sn by a two-step treatment of micro-arc oxidation （MAO） and alkali heatment was reported. Surface characterizations were performed by scanning electron microscopy （SEM）, thin film X-ray diffraction （TF-XRD） and X-ray photoelectron spectroscopy （XPS）. The MAH layer consisted of finer crystals and possessed a higher degree of crystallity and stability than the MAO layer. A biocompatibility study on treated and untreated Ti- 24Nb-4Zr-8Sn in comparison with cp Ti was carried out to investigate the effect of the different surfaces on the bone integration property in vitro. The cellular assays revealed that the MAO and MAH layer favored the initial adhesion of MC3T3-E1 cells and that the growth rate of MC3T3-E1 cells on MAH layer was significantly higher than that on the conventional MAO-treated layer after 3-day and 5-day incubation, demonstrating the greater potential of the hybrid treatment of micro-arc oxidation followed with alkali heatment as a novel surface modification method for implanting materials.
