Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the accompanying study, we report an in vitro culture system from bovine bone cells that can be applied to investigate bone cell growth and differentiation. In this system, bovine bone cells placed in mineralization medium formed multilayers (days 2-3), began deposition of mineral (days 5-6), and eventually acquired a mineralized matrix sheet (days 14-20) through the stages of mineralizing nodules and trabecular-like structure. In the current study we used this system to investigate the relative expression of bone matrix genes that may play an important role in bone development and metabolism. alpha 1(I)-collagen, alkaline phosphatase, osteonectin, biglycan (PgI), decorin (PgII), osteopontin, and bone sialoprotein mRNA gene expression were measured on days 0, 2, 6, 10, and 20 (date when the cells were placed in mineralization medium as day 0). Total RNA was purified and analyzed by northern blot using radiolabeled cDNA encoding these genes. To comprehend the relationship between gene expression and mineralization, total calcium content in the cultures was also measured. During the culture period we observed several very different gene expression profiles. The expression of both alpha 1(I)-collagen and biglycan increased 3- to 4-fold by day 6 and then returned to basal levels by day 20. The osteonectin gene was highly expressed throughout the culture, with no significant increase in induction found during any time of culture. A significant induction of alkaline phosphatase (13.8-fold) gene expression was observed by day 6. Osteopontin showed a similar profile to that of alkaline phosphatase but had a much greater level of relative expression (26-fold) compared to day 0. Interestingly, downregulation during mineral accumulation seemed a common occurrence among many of the genes measured. In contrast, the bone sialoprotein gene showed a significant and distinct expression pattern, increasing rapidly after the onset of mineralization on day 6 and ultimately reaching 140-fold that of day 0. Decorin (Pg II) showed an increasing pattern, with the final relative level of induction 5-fold on day 20. These data suggest that the development of the mature osteoblastic phenotype, complete with the ability to produce a thick mineralized matrix, requires the differential regulation of a series of genes and their gene products over the culture period.
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PMID:Bone matrix mRNA expression in differentiating fetal bovine osteoblasts. 164 43

The cartilagenous tissue of mandibular condyles of newborn mice contains progenitor cells as well as young and mature chondrogenic cells. During in vitro cultivation of the tissue, progenitor cells undergo osteogenic differentiation and form new bone (Silbermann, M., D. Lewinson, H. Gonen, M. A. Lizarbe, and K. von der Mark. 1983. Anat. Rec. 206:373-383). We have studied the expression of genes that typify osteogenic differentiation in mandibular condyles during in vitro cultivation. RNAs of the genes for collagen type I, osteonectin, alkaline phosphatase, and bone gla protein were sequentially expressed in progenitor cells and hypertrophic chondrocytes during culture. Osteopontin expression peaked in both the early and the late phase of the differentiation process. The data indicate a distinct sequence of expression of osteoblast-specific genes during osteogenic differentiation and new bone formation in mandibular condyles.
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PMID:Gene expression during osteogenic differentiation in mandibular condyles in vitro. 169 Nov 90

Osteoblasts, the bone-forming cells, synthesize the macromolecules of the bone matrix including: type I collagen; osteocalcin; osteonectin; osteopontin; proteoglycan I and II; bone sialoprotein; matrix gla-protein; bone glycoprotein 75; several other proteins, which have not been extensively characterized; growth factors, including transforming growth factor beta and fibroblast growth factor. Osteoblasts also have high levels of the membrane-bound enzyme, alkaline phosphatase, which plays a role in matrix mineralization, and receptors for tissue-specific hormones, such as parathyroid hormone, as well as many other hormones, cytokines and growth factors, which regulate bone growth, differentiation and metabolism. The expression of these various proteins, most of which are not unique to bone but which together characterize the bone phenotype, is induced during osteoblastic differentiation in a stepwise fashion, suggestive of multiple regulatory factors. The detailed sequence of the expression of osteoblastic genes in situ has not been fully characterized. It appears that type I collagen and alkaline phosphatase are expressed early during the commitment to the osteoblastic phenotype, whereas osteopontin and osteocalcin appear late during osteoblastic differentiation. Diversity among "osteoblastic" cells is also apparent, probably not all osteoblastic cells express all the features. A large number of osteoblastic models are currently available to study the expression of osteoblast-related genes in vitro. These include primary cultures from calvaria or trabecular bone from several species, including humans, osteosarcoma-derived cell lines, and experimentally immortalized cells. Some of these in vitro models, especially the calvaria-derived cultures, undergo changes which mimic osteoblastic differentiation in vivo. The study of these and other cell models started providing insights into the regulation of gene expression in osteoblastic cells. In addition to a vast body of information on the conditions required for the expression of various proteins in culture and their regulation by hormones and growth factors, more detailed information on specific genes has recently been obtained. For example, regulation of type I collagen gene expression has been studied in osteosarcoma cell lines where 1,25(OH)2 vitamin D3 was shown to act via specific DNA segment(s) in the 5' flanking region of the gene, while parathyroid hormone affected gene expression by altering the stability of the transcripts. TGF beta 1, which stimulates osteogenesis, was shown to promote the transcription of osteopontin and type I collagen, the latter effect requiring the binding site for the transactivating protein, nuclear factor I.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Gene expression in osteoblastic cells. 180 5

This study provides data concerning the cells and their extracellular matrix in prenatal human mandibular condylar cartilage. The latter cartilage represents a secondary type of cartilage since it develops late in the morphogenesis of the craniofacial skeleton. The cartilage of the mandibular condyle is actively involved in endochondral ossification, thus showing all the phases of cartilage growth, maturation, and mineralization that precedes de novo bone formation. The present study focused on the localization and distribution of the major macromolecules that are normally encountered in cartilage and bone, including collagens, proteoglycans, fibronectin, osteonectin, osteocalcin, alkaline phosphatase, and anchorin CII. It became clear that the mineralized zone of the cartilage already contained bone-specific antigens; thus the above zone might serve as an essential propagative predecessor in the ossification process.
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PMID:Immunohistochemical studies of the extracellular matrix in the condylar cartilage of the human fetal mandible: collagens and noncollagenous proteins. 184 41

We have examined the ability of dexamethasone, retinoic acid, and vitamin D3 to induce osteogenic differentiation in rat marrow stromal cell cultures by measuring the expression of mRNAs associated with the differentiated osteoblast phenotype as well as analyzing collagen secretion and alkaline phosphatase activity. Marrow cells were cultured for 8 days in primary culture and 8 days in secondary culture, with and without 10 nM dexamethasone or 1 microM retinoic acid. Under all conditions, cultures produced high levels of osteonectin mRNA. Cells grown with dexamethasone in both primary and secondary culture contained elevated alkaline phosphatase mRNA and significant amounts of type I collagen and osteopontin mRNA. Addition of 1,25-dihydroxyvitamin D3 to these dexamethasone-treated cultures induced expression of osteocalcin mRNA and increased osteopontin mRNA. The levels of alkaline phosphatase, osteopontin, and osteocalcin mRNAs in Dex/Dex/VitD3 cultures were comparable to those of 1,25-dihydroxyvitamin D3-treated ROS 17/2.8 osteosarcoma cells. Omitting dexamethasone from either primary or secondary culture resulted in significantly less alkaline phosphatase mRNA, little osteopontin mRNA, and no osteocalcin mRNA. Retinoic acid increased alkaline phosphatase activity to a greater extent than did dexamethasone but did not have a parallel effect on the expression of alkaline phosphatase mRNA and induced neither osteopontin or osteocalcin mRNAs. In all conditions, marrow stromal cells synthesized and secreted a mixture of type I and III collagens. However, dexamethasone-treated cells also synthesized an additional collagen type, provisionally identified as type V. The synthesis and secretion of collagens type I and III was decreased by both dexamethasone and retinoic acid. Neither dexamethasone nor retinoic acid induced mRNAs associated with the chondrogenic phenotype. We conclude that dexamethasone, but not retinoic acid, promotes the expression of markers of the osteoblast phenotype in cultures of rat marrow stromal fibroblasts.
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PMID:Dexamethasone induction of osteoblast mRNAs in rat marrow stromal cell cultures. 202 91

Rat frontonasal and mandibular mesenchyme was isolated from day-12 1/2 (stage-22) rat embryos and cultured at high density for up to 12 days. The stage chosen was based on the observation that mandibular mesenchyme at this stage became independent of its epithelium with respect to the production of both cartilage and bone. Frontonasal cultures developed aggregates of anastomosing columns of cells within 2 days. These grew as the cells enlarged, laying down an Alcian-blue-positive matrix by day 3 of culture. Significant mineral was detected by von Kossa staining by day 5 at which time the aggregates covered a large portion of the culture, eventually covering the entire micromass by day 10-12. Mandibular cultures developed centrally located nodular aggregates by 3 days of culture. These nodules increased in number, spreading outwards as the cells enlarged, laying down an Alcian-blue-positive matrix by day 4 and mineral by days 6-7. At this time the nodules began to elongate and coalesce, but never covered the entire culture over the 12-day period. Antibody staining revealed that in both cultures the cells were initially positive for type I collagen. Subsequently, the aggregates began expressing type II collagen, followed by type X, which coincided with the onset of mineralization. At this time some cells were negative for these cartilage markers, but positive for osteoblast markers, bone sialoprotein II, osteocalcin and type I collagen. In addition osteonectin and alkaline phosphatase were demonstrable in all of the aggregate cells late in the culture period. This provided clear evidence that chondroblast and osteoblast differentiation was proceeding within these cultures. The culture of rat facial mesenchyme should prove very useful, not only for the analysis of bone and cartilage induction and lineage relationships, but also in furthering our knowledge of craniofacial differentiation, growth and pattern formation by extending our analysis to a mammalian system.
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PMID:Formation of chondrous and osseous tissues in micromass cultures of rat frontonasal and mandibular ectomesenchyme. 227 11

Estrogen is important for both the sexual dimorphism of the skeleton during growth and the maintenance of bone balance in adults. This report describes the in vivo effects of estrogen on bone formation and gene expression in the tibial diaphysis of ovariectomized rats. Rats were ovariectomized at 8 weeks of age and were given diethylstilbestrol (DES) or placebo 1 week later as sc sustained release pellets. Histomorphometry revealed that that the periosteal bone formation and apposition rates were reduced at the tibial diaphysis 1 week after beginning estrogen treatment and further reduced after 2 weeks. Interestingly, DES treatment had no effect on endosteal bone formation, but suppressed endosteal bone resorption. Northern analysis of freshly isolated periosteal cells from tibiae and femora revealed that DES treatment resulted in dramatic decreases in steady state mRNA levels for the bone matrix proteins osteocalcin, prepro alpha 2(I) chain of type 1 collagen, osteonectin, and osteopontin as well as the osteoblast marker enzyme alkaline phosphatase. The results suggest that the inhibitory effects of estrogen on radial bone growth in rats are mediated, or at least accompanied, by the inhibition of the expression of bone matrix protein genes in periosteal cells.
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PMID:Estrogen inhibition of periosteal bone formation in rat long bones: down-regulation of gene expression for bone matrix proteins. 238 57

Human osteoblast-like cells were examined for the presence of the Ca2+-Mg2+ ATPase pump. The osteoblast-like cells had characteristic features of the osteoblast phenotype, including the presence of osteonectin, bone GLA protein, and type I collagen. The cells were able to mineralize matrix, their production of cAMP increased in response to PTH, and their alkaline phosphatase activity increased in response to 1,25-dihydroxyvitamin D3. Immunocytochemical staining of the osteoblast-like cells with a monoclonal antibody against human red cell Ca2+-Mg2+ ATPase demonstrated the presence of an epitope of the Ca2+-Mg2+ ATPase in these cells; staining of paraffin-embedded osteoblast-like cell sections demonstrated anti-Ca2+-Mg2+ ATPase staining only in cell plasma membranes. Western blot analysis of osteoblast-like cell homogenates showed that the monoclonal antibody to human erythrocyte Ca2+-Mg2+ ATPase bound to a major band at 140,000 mol wt, similar to the mol wt of known plasma membrane Ca2+-Mg2+ ATPases. The presence in the osteoblast-like cells of a Ca2+-Mg2+ ATPase similar to the human red cell calcium pump suggests that this enzyme may play a role in osteoblast intracellular calcium homeostasis.
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PMID:Epitopes of the human erythrocyte Ca2+-Mg2+ ATPase pump in human osteoblast-like cell plasma membranes. 246 88

We have used in situ hybridization to examine expression of collagen type I, II, and X mRNA and osteonectin mRNA in the chick epiphysis. Tissue samples from the proximal tibial growth cartilage were fixed in modified Carnoy's solution, dehydrated in ethanol, and embedded in paraffin. Longitudinal and transverse sections were demineralized with HCl and digested with hyaluronidase and proteinase K. In situ hybridization was carried out using biotinylated cDNA probes; the hybridized probe was detected using a streptavidin-biotinylated alkaline phosphatase conjugate. This procedure permitted detection of the corresponding mRNAs in cartilage with high sensitivity and low background. Osteonectin mRNA was detected in proliferating cartilage; lower levels of osteonectin mRNA were seen in the mid-hypertrophic region. This mRNA species was also expressed in cells that border the vascular canals in the premineralized region of the epiphysis. Collagen type X mRNA was detected throughout the hypertrophic zone. As localization of collagen type X mRNA corresponded to the site of maximal synthesis of the protein, reported in other studies, our results would further support the suggestion that this protein is associated with mineralization of cartilage. Collagen type II mRNA was seen in both the proliferating and the hypertrophic regions of the cartilage. Highest levels of expression were observed in the proliferative region. The results suggest that the transcriptional control of collagen type II and X by cells of the proliferating and hypertrophic regions of the growth cartilage may be related.
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PMID:Developmental expression of genes in chick growth cartilage detected by in situ hybridization. 250 10

Cells derived from embryonic rat calvariae were immortalized by retroviral delivery of cDNA for the SV-40 large T antigen and the bacterial neomycin resistance gene. After selection with G418, cells were cloned by limiting dilution and screened for expression of osteoblast characteristics. One clone (RCT-3), derived from cells collected during the third period of enzymatic digestion, showed high constitutive expression of alkaline phosphatase (ALP), synthesized type I collagen in the virtual absence of type III and exhibited a parathyroid hormone (PTH)-responsive adenylate cyclase (EC50, 10 nM). Messenger RNAs for osteonectin and osteopontin were present in RCT-3 cells and osteopontin mRNA was enhanced by 1,25 (OH)2 vitamin D3 treatment. The other cell line (RCT-1), derived from cells released during the first 10 min of digestion, expressed osteoblast features only after 3 d treatment with 1 microM retinoic acid (RA). ALP activity increased from 0.003 to 0.25 mumole/min/mg protein, there was a substantial increase in the steady-state level of type I collagen mRNA and a dose-dependent and saturable response to PTH was induced (EC50, 10 nM). Osteopontin mRNA was induced by 1,25 (OH)2D3. This study has provided two new cell lines which may be useful models for studies of differentiation-related gene expression in bone cells.
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PMID:SV-40 large-T immortalization of embryonic bone cells: establishment of osteoblastic clonal cell lines. 261 49


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