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9. Developmental aspects of bone: cellular differentiation of cartilage and bone
254 cytoplasmic concentration and distribution of ionized calcium. However, three peaks were consistently detected in the distribution of concentrations, and these peak ratios were constant in different osteoclasts, perhaps suggesting different spatial distribution of calcium in these cells. These preliminary experiments show that calcium concentrations are unequal throughout these cells with local areas of high calcium concentration. First attempts at ratio imaging of functioning, resorptive osteoclasts were hampered by the high levels of autofluorescence from the dentine substrates.
9. Developmental aspects of bone: cellular differentiation of cartilage and bone BK Hall Biology Department, Dalhousie Unizwsity, Halifax, NS, Canada B3H 411 Although bone and cartilage share many properties and may even arise from the same population of cells, bone may readily be distinguished from cartilage and only rarely does one find tissues (chondroid bone, secondary cartilage?) that are intermediate between bone and cartilage. The implication is that skeletal progenitor cells have two major choices with respect to cell fate, either because they are predetermined as chondroblasts or because the processes of embryonic development allow them to elect one fate over the other. These developmental events will be elaborated by discussing how bone is distinguished from cartilage and from the intermediate skeletal tissues, and then by discussing the evidence for the origin of bone and cartilage from mesoderm and from the embryonic neural crest. Whether cells are ‘determined’ as osteoblasts or chondroblasts early in embryonic life will be briefly discussed, briefly because we do not yet know. Before cytodifferentiation commences, progenitor cells accumulate at sites of future skeletogenesis via enhanced proliferation, migration and/or reduced cell death. The neural crest-derived craniofacial skeleton will be used to illustrate (a) the role that cell - cell and cell-matrix interactions play in ensuring that sufficient cells arise and become localized at the site of a future skeletal element, (b) that differentiation is initiated within condensations in response to epigenetic interaction with epithelial extracellular matrices or with other developing tissues such as muscles or blood vessels, and (c) that physical factors such as biomechanical stimuli, or molecules such as bone morphogenetic protein (BMP) within epithelial basement membranes may act in the embryo as signals for the differentiation of skeletogenic cells.
10. Transformation of chondrocytes to osteoblasts in organ culture CW Thesingh Laboratory of Cell Biology and Histology, Medical Faculty, Leiden. The Netherlands In vivo the first osteoclasts to enter the hypertrophic cartilaginous midshaft of the embryonic long bone from the periosteum expel the hypertrophic chondrocytes
Abstracts from the Bone and Tooth Society Meeting from their lacunae by extending long processes between the chondrocyte membrane and the lacunar wall. During this event the chondrocytes change appearance, their nucleus becomes dense and they sometimes even divide at the last moment. Once released in the excavated marrow cavity they escape recognition amidst the many invading cells. There are no indications that they die in great numbers. 3H-thymidine labeled hypertrophic chondrocytes, set free from their lacunae, turn up as ‘stromal’ cells and sometimes as endochondral osteoblasts inside the marrow cavity in co-cultured labeled periostless long bones with a nonlabeled osteoclast source. Hypertrophic chondrocytes, kept in confinement inside their lacunae in cultured periostless long bones transform into osteoblasts under certain stimuli, changing from alkaline phosphatase negative to strongly alkaline phosphatase positive cells. They produce a calcified bone matrix, which contains the course collagenic fibres of bone, and osteocalcin. The transformation process from hypertrophic chondrocytes to osteoblasts passes via a stage of stromallike, dividing cells. Brain tissue or brain extract from the same or from different species promote this transformation. Brain tissue in co-culture (but separated by a filter) with periostless (‘stripped’) long bones also stimulates the formation of a new ‘periosteum’, supposedly from chondrocytes abraded from the cartilage during the stripping procedure and still glued to the surface of the long bone. In this newly formed periosteum ‘bone’ formation and sometimes cartilage formation occurs.
11. Direct effects of intermittent compressive force on mineral metabolism in bone and cartilage in vitro J Klein-Nuland, JP Veldhuijzen and EH Burger Department of Oral Cell Biology, ACTA-Vrije Universiteit, De Boelelaan 1115, 1081 HVAmsterdam, The Netherlands There is increasing evidence that weightbearing exercise increases bone mass, but the cellular effector mechanism of mechanical stimulation of bone tissue is unknown. We have developed a tissue culture model which allows application of intermittent compressive force (ICF) to organ cultures of fetal bone rudiments, to study direct effects of mechanical stimulation on mineral metabolism. ICF is generated by intermittently compressing the gas phase (5% CO2 in air) of a closed culture chamber (humidity 98%) which is kept at 37°C. Fetal mouse long bone or calvarial rudiments were cultured for 3 or 5 days with or without ICF of 132glcm2, calculated to resemble maximal physiological pressure in vivo, and 0.3Hz pulse frequency. Culture under constant atmospheric pressure was considered to mimic disuse condition (DC). Osteoclastic resorption of the mineral phase was monitored by measuring release of radiocalcium from prelabeled fetal bone rudiments. Under ICF, release of radiocalcium was significantly lower than under DC.
9. Developmental aspects of bone: cellular differentiation of cartilage and bone BK Hall Biology Department, Dalhousie Unizwsity, Halifax, NS, Canada B3H 411 Although bone and cartilage share many properties and may even arise from the same population of cells, bone may readily be distinguished from cartilage and only rarely does one find tissues (chondroid bone, secondary cartilage?) that are intermediate between bone and cartilage. The implication is that skeletal progenitor cells have two major choices with respect to cell fate, either because they are predetermined as chondroblasts or because the processes of embryonic development allow them to elect one fate over the other. These developmental events will be elaborated by discussing how bone is distinguished from cartilage and from the intermediate skeletal tissues, and then by discussing the evidence for the origin of bone and cartilage from mesoderm and from the embryonic neural crest. Whether cells are ‘determined’ as osteoblasts or chondroblasts early in embryonic life will be briefly discussed, briefly because we do not yet know. Before cytodifferentiation commences, progenitor cells accumulate at sites of future skeletogenesis via enhanced proliferation, migration and/or reduced cell death. The neural crest-derived craniofacial skeleton will be used to illustrate (a) the role that cell - cell and cell-matrix interactions play in ensuring that sufficient cells arise and become localized at the site of a future skeletal element, (b) that differentiation is initiated within condensations in response to epigenetic interaction with epithelial extracellular matrices or with other developing tissues such as muscles or blood vessels, and (c) that physical factors such as biomechanical stimuli, or molecules such as bone morphogenetic protein (BMP) within epithelial basement membranes may act in the embryo as signals for the differentiation of skeletogenic cells.
10. Transformation of chondrocytes to osteoblasts in organ culture CW Thesingh Laboratory of Cell Biology and Histology, Medical Faculty, Leiden. The Netherlands In vivo the first osteoclasts to enter the hypertrophic cartilaginous midshaft of the embryonic long bone from the periosteum expel the hypertrophic chondrocytes
Abstracts from the Bone and Tooth Society Meeting from their lacunae by extending long processes between the chondrocyte membrane and the lacunar wall. During this event the chondrocytes change appearance, their nucleus becomes dense and they sometimes even divide at the last moment. Once released in the excavated marrow cavity they escape recognition amidst the many invading cells. There are no indications that they die in great numbers. 3H-thymidine labeled hypertrophic chondrocytes, set free from their lacunae, turn up as ‘stromal’ cells and sometimes as endochondral osteoblasts inside the marrow cavity in co-cultured labeled periostless long bones with a nonlabeled osteoclast source. Hypertrophic chondrocytes, kept in confinement inside their lacunae in cultured periostless long bones transform into osteoblasts under certain stimuli, changing from alkaline phosphatase negative to strongly alkaline phosphatase positive cells. They produce a calcified bone matrix, which contains the course collagenic fibres of bone, and osteocalcin. The transformation process from hypertrophic chondrocytes to osteoblasts passes via a stage of stromallike, dividing cells. Brain tissue or brain extract from the same or from different species promote this transformation. Brain tissue in co-culture (but separated by a filter) with periostless (‘stripped’) long bones also stimulates the formation of a new ‘periosteum’, supposedly from chondrocytes abraded from the cartilage during the stripping procedure and still glued to the surface of the long bone. In this newly formed periosteum ‘bone’ formation and sometimes cartilage formation occurs.
11. Direct effects of intermittent compressive force on mineral metabolism in bone and cartilage in vitro J Klein-Nuland, JP Veldhuijzen and EH Burger Department of Oral Cell Biology, ACTA-Vrije Universiteit, De Boelelaan 1115, 1081 HVAmsterdam, The Netherlands There is increasing evidence that weightbearing exercise increases bone mass, but the cellular effector mechanism of mechanical stimulation of bone tissue is unknown. We have developed a tissue culture model which allows application of intermittent compressive force (ICF) to organ cultures of fetal bone rudiments, to study direct effects of mechanical stimulation on mineral metabolism. ICF is generated by intermittently compressing the gas phase (5% CO2 in air) of a closed culture chamber (humidity 98%) which is kept at 37°C. Fetal mouse long bone or calvarial rudiments were cultured for 3 or 5 days with or without ICF of 132glcm2, calculated to resemble maximal physiological pressure in vivo, and 0.3Hz pulse frequency. Culture under constant atmospheric pressure was considered to mimic disuse condition (DC). Osteoclastic resorption of the mineral phase was monitored by measuring release of radiocalcium from prelabeled fetal bone rudiments. Under ICF, release of radiocalcium was significantly lower than under DC.