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)

Human bone marrow stromal cells were examined for their osteogenic potential in an in vitro cell culture system. Dexamethasone (Dex) treatment induced morphological transformation of these cells from an elongated to a more cuboidal shape, increased their alkaline phosphatase activity and cAMP responses to PTH and prostaglandin E2, and was essential for mineralization of the extracellular matrix. Dex-induced differentiation of human bone marrow stromal cells was apparent after 2-3 days of treatment and reached a maximum at 7-14 days, as judged by alkaline phosphatase activity, although induction of osteocalcin by 1,25-dihydroxyvitamin D3 was attenuated by Dex. Withdrawal of Dex resulted in an enhancement of the 1,25-dihydroxyvitamin D3-induced secretion of osteocalcin, whereas alkaline phosphatase activity and the cAMP response to PTH remained at prewithdrawal levels. The steady state mRNA level of osteonectin was not affected by Dex. Our results, which demonstrate that Dex conditions the differentiation of human bone marrow osteogenic stromal cells into osteoblast-like cells, support the hypothesis of a permissive effect of glucocorticoids in ensuring an adequate supply of mature osteoblast populations. Furthermore, the established human bone marrow stromal cell culture provides a good model of an in vitro system to study the regulation of differentiation of human bone osteoprogenitor cells.
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PMID:Differentiation of human bone marrow osteogenic stromal cells in vitro: induction of the osteoblast phenotype by dexamethasone. 827 45

Numerous studies of experimental hypo- and hypervitaminosis A have long suggested that retinoic acid (RA) is involved in chondrocyte maturation during endochondral ossification and skeletogenesis. However, the specific and direct roles of RA in these complex processes remain unclear. Based on recent studies from our laboratories, we tested the hypothesis that RA induces the expression of genes associated with the terminal mineralization phase of chondrocyte maturation and promotes apatite deposition in the extracellular matrix. Cell populations containing chondrocytes at advanced stages of maturation were isolated from the upper portion of Day 18 chick embryo sterna and grown for 2 weeks in monolayer until confluent. The cells were then treated with low doses (10-100 nM) of RA for up to 6 days in the presence of a phosphate donor (beta-glycerophosphate) but in the absence of ascorbic acid. Within 4 days of treatment, RA dramatically induced expression of the alkaline phosphatase (APase), osteonectin, and osteopontin genes, caused a several-fold increase in APase activity, and provoked massive mineral formation while it left type X collagen gene expression largely unchanged. The mineral had a mean Ca/Pi molar ratio of 1.5; Fourier transform infrared spectra confirmed that it represented hydroxyapatite. Mineralization was completely abolished by treatment with parathyroid hormone; this profound effect confirmed that RA induced cell-mediated mineralization and not nonspecific precipitation. When cultures were treated with both RA and ascorbic acid, there was a slight further increase in APase activity and increased calcium accumulation. The effects of RA were also studied in cultures of immature chondrocytes isolated from the caudal portion of sternum; however, RA only had minimal effects on mineralization and gene expression in these cells. Thus, RA appears to be a rapid, potent, maturation-dependent, ascorbate-independent promoter of terminal maturation and matrix calcification in chondrocytes.
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PMID:Retinoic acid induces rapid mineralization and expression of mineralization-related genes in chondrocytes. 834 89

Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) has been used to map the extracellular secretory activity of normal osteoblasts. The proteins osteonectin, bone sialoprotein, and both the C-telopeptide of collagen I together with collagen I have now been positionally identified. In addition the secretory differences which exist between normal and Pagetic osteoblasts have been mapped with the Pagetic osteoblasts shown to consistently secrete an altered 30 kDa C-telopeptide of collagen type I. The use of the diphosphonate Pamidronate in the treatment of Paget's disease of bone has beneficial effects with suppression of the bone isoenzyme marker alkaline phosphatase. It has been reported that diphosphonates directly inhibit human osteoblast secretory function as well as osteoclast metabolism. The effects of Pamidronate on the secretory activity of normal and Pagetic osteoblast cultures was also investigated. The extracellular protein secretion of normal and Pagetic osteoblasts was not affected by Pamidronate treatment as assessed by 2-D PAGE. This technique allows a comprehensive multiparameter assessment of extracellular secretory activity in normal and diseased states. The findings show that the Pagetic osteoblasts cultured in vitro are functionally abnormal and they support the hypothesis that the underlying problem in Paget's disease is characterised by disorder of osteoblast and osteoclast interactions.
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PMID:Extracellular protein secretion of cultured normal and Pagetic osteoblasts. 839 80

This study reports the selection and characterization of osteogenic precursors from human bone marrow which were isolated by two "clonings" and successive subculturing. These cell lines express alkaline phosphatase activity. Gel electrophoresis of [3H]-proline labeled cultures showed that the cloned cells produce only type I collagen. They synthetize osteocalcin and osteonectin. They respond to 1,25 dihydroxy vitamin D3 by increasing osteocalcin synthesis and secretion, and to parathyroid hormone by increasing cyclic AMP synthesis. After the third subculture in the absence of beta-glycerophosphate, these cell lines formed lots of clusters which exhibit high alkaline phosphatase activity and positive von Kossa staining. X-ray energy spectrum shows that these cells are surrounded by "budding" structures containing calcium and phosphorus with a ratio Ca:P identical to those of pure hydroxyapatite. This process was associated with 45Ca uptake into the cells. All these data support the selection of osteogenic cells which may be of considerable clinical importance.
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PMID:Human bone marrow stromal cells express an osteoblastic phenotype in culture. 840 13

We established cultures of cells growing out from adult bone chips and maintained them through 12 passages in culture. The cultures showed osteoblastic phenotype accompanied by synthesis of collagen type I, osteonectin, alkaline phosphatase, and osteocalcin. We report the characterization of 21 clones obtained from three different individual primary cultures. We studied the expression of osteonectin, alkaline phosphatase, collagen, and osteocalcin in the clones. Metabolic labeling showed production of type I collagen and of osteonectin in all clones studied. In two-thirds of the clones and in mass cultures alkaline phosphatase was not detected at passage 2, but it was detected in increasing amounts at later passages in culture. The clones attained different but detectable levels of expression of this marker by passage 8. The different levels in the expression of alkaline phosphatase in positive clones may be because they were derived from cells at different stages of osteoblastic maturation or due to small changes in microenvironment. The alkaline phosphatase-positive clones were tested for osteocalcin, and they showed measurable expression only at passage 10. A third of the clones obtained were negative for alkaline phosphatase during 12 passages in culture. The obtainment of clones unable to produce alkaline phosphatase may be due to loss of differentiating potential under the in vitro culture conditions. The growth rate and potential of all clones studied were similar through 12 passages in culture, regardless of their potential for expression of alkaline phosphatase.
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PMID:Differential expression of alkaline phosphatase in clones of human osteoblast-like cells. 845 85

Endosteal bone surface cells were previously shown to be involved in the regulation of bone formation in humans. In this study, we have characterized the cells isolated from the endosteal bone surface in adult rats. Fragments of periosteum-free tibia were obtained from 4-, 6- and 9-month-old rats by collagenase digestion, and the phenotypic characteristics of the osteoblastic cells migrating from the endosteal bone surface were evaluated in culture. Endosteal bone surface cells present a strong alkaline phosphatase (ALP) activity as shown by cytochemistry and measured biochemically. The cells synthesize high levels of osteocalcin as measured by radioimmunoassay. Osteocalcin production was increased after stimulation with 10 nM 1,25 dihydroxyvitamin D (1,25(OH)2 D) and the response to 1,25(OH)2 D was similar at all ages. Endosteal cells from young adult rats (4 months old) but not from older rats (6 and 9 months old) showed increased cAMP production in response to 10 nM parathyroid hormone (PTH), suggesting an age-related decrease in the PTH-responsiveness of the bone surface cells. Immunocytochemistry using specific antibodies showed that preconfluent endosteal bone cells from adult rats expressed collagen and noncollagenous bone proteins in culture in the absence of inducers. The cells synthesized mostly type-I collagen which remained localized intracellularly. Type-III collagen was only expressed at low levels. The bone surface cells also expressed osteocalcin and bone sialoprotein, two markers of differentiated osteoblasts, as well as osteonectin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Cells isolated from the endosteal bone surface of adult rats express differentiated osteoblastic characteristics in vitro. 847 7

The effects of skeletal unloading on osteoblastic cells were evaluated in tail-suspended rats. Hindlimb elevation for 14 days induced osteopenia, decreased histomorphometric indexes of bone formation in tibial metaphysis, and reduced plasma osteocalcin and alkaline phosphatase (ALP) levels compared with controls. The in vitro proliferation of osteoblastic cells isolated from the endosteal bone surface of suspended tibias was decreased by 42 and 31% at 2 and 4 days of culture, respectively, compared with controls, as shown by [3H]thymidine labeling and cell number. The proliferation of ALP-positive marrow stromal cells was also decreased by 20-24% at 1 and 2 days of culture. However, ALP activity in bone-derived cells and marrow stromal cells was not different in unloaded and control rats, and the number of bone cells synthesizing osteocalcin, osteonectin, and type I or type III collagen was identical in the two groups. The results indicate that the inhibition of bone formation induced by skeletal unloading is related to a decreased proliferation of putative osteoblast precursor cells present along the endosteal bone surface and in the marrow stroma.
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PMID:Skeletal unloading in rat decreases proliferation of rat bone and marrow-derived osteoblastic cells. 849

Ultrathin sections from the dermis of five normal subjects and from 10 patients suffering from pseudoxanthoma elasticum (PXE) were analyzed by immunoelectron microscopy with the aim of identifying and localizing proteins associated with the mineral precipitates within the altered elastic fibers. Serial sections were processed by indirect immunogold cytochemistry using primary antibodies against human fibronectin, vitronectin, bone sialoprotein, alkaline phosphatase, osteonectin, and osteopontin. In the latter two cases, antibodies against synthetic peptides were also used. The results indicate that normal elastic fibers contained osteopontin, and that this protein was associated with the apparently normal elastin as well as with the needle-shaped mineral precipitates in the elastic fibers of patients. On the contrary, significant amounts of vitronectin, alkaline phosphatase and, less, of bone sialoprotein were associated with the polymorphous mineral precipitates inside the elastic fibers. Large amounts of osteonectin and fibronectin, together with vitronectin, were localized on the microfilament aggregates, which were often associated with altered elastic fibers in PXE dermis and were never visualized in the dermis of control subjects. The results seem to indicate once more that PXE is a complex disorder of the fibroblast synthetic control. Elastic fiber mineralization might be considered a secondary event, which could depend on the abnormal synthesis and accumulation within the elastic fibers of proteins that are normally involved in mineralization processes.
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PMID:Matrix proteins with high affinity for calcium ions are associated with mineralization within the elastic fibers of pseudoxanthoma elasticum dermis. 857 19

Gene expression patterns have been investigated in prolonged cultures of rat mandibular condylar cartilage (MCC) cells to examine the possibility of culture-induced phenotypic changes. MCC cells were isolated from newborn rats and grown in the presence of 10% fetal bovine serum (FBS) and basic fibroblast growth factor (bFGF). MCC cells were passaged and cultured in the presence of 10% FBS and bFGF until confluent. After confluence, the medium was changed to that supplemented with 10% FBS, ascorbate, and beta-glycerophosphate (day 1). Mineralization and gene expression of MCC cells have been investigated. Mineralization, visualized by staining with Alizarin red, was observed to begin at day 13 in culture, and increased up to day 22 in culture, which was the length of this study. Type II collagen and aggrecan mRNAs were highly expressed at the start of culture (day 1-4) and decreased in a time-dependent manner. Type I collagen, alkaline phosphatase, and osteopontin mRNAs expressed biphasic patterns that peaked at the start of culture and the beginning of mineralization (day 13-16). Osteonectin mRNA was expressed throughout the culture period. Osteocalcin mRNA was expressed before the beginning of mineralization peaking at day 7. These observations suggest that the gene expression patterns of MCC cells can be categorized into two different periods in prolonged culture: maturation (day 1-10) and mineralization (day 13-22). The day culture system of MCC represents a new model system in which the differentiation of embryonic MCC cells can be examined.
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PMID:Changes in phenotypic gene expression in rat mandibular condylar cartilage cells during long-term culture. 859 45

The effect of recombinant Pasteurella multocida toxin (PMT) on primary cultures of embryonic chick bone-derived osteoblastic cells was investigated. It was found that PMT was a potent mitogen for primary derived chicken osteoblasts. The toxin stimulated DNA synthesis and cell proliferation in quiescent osteoblasts at the first passage and accelerated cell growth in subconfluent cultures. Cell viability was not affected by PMT, even at relatively high concentrations. Osteoblast numbers increased in a dose-dependent manner in response to PMT. Intracellular inositol phosphates were elevated in response to PMT, but no elevation in cyclic AMP (cAMP) levels was evident. Indeed, PMT inhibited cAMP elevation in osteoblasts in response to cholera toxin at a stage before other PMT-mediated events take place. In addition to increased cell turnover, PMT down-regulated the expression of several markers of osteoblast differentiation. Both alkaline phosphatase and type I collagen were reduced, but osteonectin was not affected. The in vitro deposition of mineral in cultures of primary osteoblasts and osteoblast-like osteosarcoma cells was also inhibited by the presence of PMT. This suggests that PMT interferes with differentiation at a preosteoblastic stage.
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PMID:Pasteurella multocida toxin is a mitogen for bone cells in primary culture. 864 7


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