Objective: To purify and identify the osteoclasts from the tissue of humangiant cell tumor of bone. Methods: We have developed a new method that allows the purification oflarge numbers of authentic osteoclasts (OCs). ...Objective: To purify and identify the osteoclasts from the tissue of humangiant cell tumor of bone. Methods: We have developed a new method that allows the purification oflarge numbers of authentic osteoclasts (OCs). The OCs were isolated from tissue of human giant celltumor of bone by 0.25% trypsin and collagenase. We characterized OCs in terms of the expression ofdifferent phenotypic markers of OCs. The phenotypic markers of OC included Tartrate-resistant acidphosphatase staining (TRAP). The expression of calcitonin receptor (CTR), cathepsin K and receptoractivator of necrosis factor κB (RANK) mRNA were examined by RT-PCR. Results: The OC cell purifiedby above method functioned normally in vitro. The purity was about 79.7%. They showed the normalosteoclast phenotypes markers of OC. Conclusion: The method provides a system for performingbiochemical and molecular studies of OCs. The study indicates that the method of purifying theosteoclasts from human GCT cell can be used for research of bone metabolism.展开更多
Purpose： To investigate the influence of the same mechanical loading on osteogenesis and osteoclastogenesis in vitro. Methods： Primary osteoblasts, bone marrow-derived mesenchymal stem cells （BMSCs, cultured in ost...Purpose： To investigate the influence of the same mechanical loading on osteogenesis and osteoclastogenesis in vitro. Methods： Primary osteoblasts, bone marrow-derived mesenchymal stem cells （BMSCs, cultured in osteoinductive medium） and RAW264.7 cells cultured in osteoclast inductive medium were all subjected to a 1000μstrain （μs） at 1 Hz cyclic mechanical stretch for 30 min （twice a day）. Results： After mechanical stimulation, the alkaline phosphatase （ALP） activity, osteocalcin protein level of the osteoblasts and BMSCs were all enhanced, and the mRNA levels of ALP and collagen type I increased. Additionally, extracellular-deposited calcium of both osteoblasts and BMSCs increased. At the same time, the activity of secreted tartrate-resistant acid phosphatase, the number of tartrate-resistant acid phosphatase-positive multinucleated cells, matrix metalloproteinase-9 protein levels of RAW264.7 cells and the extracellular calcium solvency all decreased. Conclusion： The results demonstrated that 1000 μs cyclic mechanical loading enhanced osteoblasts activity, promoted osteoblastic differentiation of BMSCs and restrained osteoclastogenesis of RAW264.7 cells in vitro.展开更多
文摘Objective: To purify and identify the osteoclasts from the tissue of humangiant cell tumor of bone. Methods: We have developed a new method that allows the purification oflarge numbers of authentic osteoclasts (OCs). The OCs were isolated from tissue of human giant celltumor of bone by 0.25% trypsin and collagenase. We characterized OCs in terms of the expression ofdifferent phenotypic markers of OCs. The phenotypic markers of OC included Tartrate-resistant acidphosphatase staining (TRAP). The expression of calcitonin receptor (CTR), cathepsin K and receptoractivator of necrosis factor κB (RANK) mRNA were examined by RT-PCR. Results: The OC cell purifiedby above method functioned normally in vitro. The purity was about 79.7%. They showed the normalosteoclast phenotypes markers of OC. Conclusion: The method provides a system for performingbiochemical and molecular studies of OCs. The study indicates that the method of purifying theosteoclasts from human GCT cell can be used for research of bone metabolism.
基金This work was financially supported by the National Natural Science Foundation of China （No.11372351, No.31370942, No.81160223）, and Scientific Research Foundation of Guangxi Higher Education （No.KY2015LX241）.
文摘Purpose： To investigate the influence of the same mechanical loading on osteogenesis and osteoclastogenesis in vitro. Methods： Primary osteoblasts, bone marrow-derived mesenchymal stem cells （BMSCs, cultured in osteoinductive medium） and RAW264.7 cells cultured in osteoclast inductive medium were all subjected to a 1000μstrain （μs） at 1 Hz cyclic mechanical stretch for 30 min （twice a day）. Results： After mechanical stimulation, the alkaline phosphatase （ALP） activity, osteocalcin protein level of the osteoblasts and BMSCs were all enhanced, and the mRNA levels of ALP and collagen type I increased. Additionally, extracellular-deposited calcium of both osteoblasts and BMSCs increased. At the same time, the activity of secreted tartrate-resistant acid phosphatase, the number of tartrate-resistant acid phosphatase-positive multinucleated cells, matrix metalloproteinase-9 protein levels of RAW264.7 cells and the extracellular calcium solvency all decreased. Conclusion： The results demonstrated that 1000 μs cyclic mechanical loading enhanced osteoblasts activity, promoted osteoblastic differentiation of BMSCs and restrained osteoclastogenesis of RAW264.7 cells in vitro.
