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)

When UMR201 cells, phenotypically preosteoblastic, were placed onto a type I collagen gel, expression of osteopontin (OP) mRNA and protein were strongly upregulated, compared to cells plated onto plastic. This upregulation was dose-dependent, with respect to the concentration of collagen gel, and was observable within hours of cells having attached and spread on the substrate. Retinoic acid (RA) acted synergistically with type I collagen at each concentration to induce a much greater increase in OP mRNA than in cells on plastic. In addition, RA increased the phosphorylation of secreted OP. The exogenous collagen substrate inhibited the growth of UMR201 cells, with the extent and duration of inhibition dependent on the collagen concentration. The effect of type I collagen was specific; plating cells on fibronectin, laminin or vitronectin did not upregulate OP expression. In contrast to the effects on OP expression, the strong RA induction of alkaline phosphatase (ALP) mRNA in cells on plastic was attenuated in cells plated on type I collagen. Growth on type I collagen did not change OP mRNA stability or transcription rate, although there was decreased stability of the ALP mRNA in cells on collagen.
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PMID:Regulation of osteopontin expression by type I collagen in preosteoblastic UMR201 cells. 883 49

Elastin molecules aggregate in the extracellular space where they are crosslinked by stable desmosine bridges. The resulting polymer is structurally organized as branched fibers and lamellae, which, in skin, are wider (a few microns) in the deep dermis and become progressively thinner (fraction of a micron) towards the papillary dermis. Several general and local factors seem to regulate elastin gene expression, deposition and degradation. In skin, the volume density of the elastin network increases from birth up to maturity, when it accounts for about 3-4% of the tissue. However, its amount and distribution depend on dermis areas, which are different among subjects and change with age. Several matrix molecules (glycosaminoglycans, decorin, biglycan, osteopontin) have been found to be associated with elastin into the normal fiber, and several others have been recognized within pathologic elastic fiber (osteonectin, vitronectin, alkaline phosphatase in PXE). With age, and in some pathologic conditions, skin elastin may undergo irreversible structural and compositional changes, which seem to progress from localized deposition of osmiophilic materials to the substitution of the great majority of the amorphous elastin with interwoven filaments negative for elastin specific antibodies.
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PMID:Elastic fiber during development and aging. 929 92

To clarify the function of osteopontin in osteoblast differentiation, we have examined the signal transduction pathway in an osteoblastic cell line (UMR106-6) bound to osteopontin, fibronectin, vitronectin and collagen type I surfaces. This was done by investigating the production and autophosphorylation of focal adhesion kinase (FAK) and the expression of alkaline phosphatase (ALP) at the transcription level. Results suggest that osteopontin was not only responsible for the autophosphorylation of FAK but regulated the expression of ALP, which was strongly correlated with FAK activity. These results suggest that osteopontin might act as a trigger in the early differentiation of osteoblasts.
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PMID:Osteopontin involvement in integrin-mediated cell signaling and regulation of expression of alkaline phosphatase during early differentiation of UMR cells. 945 May 60

The integrin alpha 8 beta 1 has been reported to bind to fibronectin, vitronectin, and tenascin-C in cell adhesion or neurite outgrowth assays. Here, we describe cDNA cloning of the murine alpha 8 subunit, purification of a recombinant soluble heterodimer consisting of the extracellular domains of the murine alpha 8 and beta1 subunits, and development of a sensitive binding assay using a modified form of this heterodimer fused to alkaline phosphatase (AP). In binding assays, the purified alpha 8 beta 1-AP chimera exhibited the same divalent ion requirements for activation and binding specificity as cell surface alpha 8 beta 1: in the presence of Mn2+ it bound to fibronectin and vitronectin in an RGDS-peptide inhibitable manner. Contrary to previous reports, we found no evidence that alpha 8 beta 1, expressed on K562 cells or as an AP chimera, interacts strongly with native tenascin-C. In binding, adhesion, and spreading assays, significant interactions were observed only to short fragments of tenascin-C containing the third fibronectin type III repeat which contains an RGD sequence. Full length tenascin-C and longer fragments containing this repeat did not appear to serve as ligands, implying that the RGD site in native tenascin-C is a cryptic binding site for this integrin, exposed by removal of adjacent domains. Soluble integrin-AP chimeras should be generally useful for identifying and characterizing integrin interactions with ligands.
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PMID:Utilization of a soluble integrin-alkaline phosphatase chimera to characterize integrin alpha 8 beta 1 receptor interactions with tenascin: murine alpha 8 beta 1 binds to the RGD site in tenascin-C fragments, but not to native tenascin-C. 954 28

Epithelio-mesenchymal interactions during kidney organogenesis are disrupted in integrin alpha8 beta1-deficient mice. However, the known ligands for integrin alpha8 beta1-fibronectin, vitronectin, and tenascin-C-are not appropriately localized to mediate all alpha8 beta1 functions in the kidney. Using a method of general utility for determining the distribution of unknown integrin ligands in situ and biochemical characterization of these ligands, we identified osteopontin (OPN) as a ligand for alpha8 beta1. We have coexpressed the extracellular domains of the mouse alpha8 and beta1 integrin subunits as a soluble heterodimer with one subunit fused to alkaline phosphatase (AP) and have used the alpha8 beta1-AP chimera as a histochemical reagent on sections of mouse embryos. Ligand localization with alpha8 beta1-AP in developing bone and kidney was observed to be overlapping with the distribution of OPN. In "far Western" blots of mouse embryonic protein extracts, bands were detected with sizes corresponding to fibronectin, vitronectin, and unknown proteins, one of which was identical to the size of OPN. In a solid-phase binding assay we demonstrated that purified OPN binds specifically to alpha8 beta1-AP. Cell adhesion assays using K562 cells expressing alpha8 beta1 were used to confirm this result. Together with a recent report that anti-OPN antibodies disrupt kidney morphogenesis, our results suggest that interactions between OPN and integrin alpha8 beta1 may help regulate kidney development and other morphogenetic processes.
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PMID:Identification of osteopontin as a novel ligand for the integrin alpha8 beta1 and potential roles for this integrin-ligand interaction in kidney morphogenesis. 961 84

Lipopolysaccharide (LPS) in solution primes neutrophils for enhanced release of superoxide in response to N-formyl-methionyl-leucyl-phenylalanine. We show that LPS immobilized on polystyrene or polypropylene acted on neutrophils by a mechanism different from that of LPS in solution. Coating the surface with 1% plasma, either before coating with LPS (plasma/LPS) or after coating with LPS (LPS/plasma), was essential to induce the LPS response in neutrophils. However, plasma could be replaced by fibrinogen, type I collagen or type IV collagen, or, to a lesser extent, by fibronectin or vitronectin, which was not true for LPS in solution. About 20% of the LPS added was immobilized on the plastic surfaces, based on its ability to adsorb anti-LPS antibody after extensive washing. The amount of soluble LPS that might have been released from surfaces during the incubation with neutrophils was too low to account for the priming by immobilized LPS. About 13-20 min was needed for neutrophils to become primed after incubation with immobilized LPS. Immobilized LPS induced up-regulation of CD11b/CD18 and latent alkaline phosphatase and also enhanced the adhesive response of neutrophils. Priming by immobilized LPS was inhibited by anti-CD14 antibody or by treatment of neutrophils with the LPS antagonist LA-14-PP. When immobilized LPS was treated with anti-LPS-binding protein (LBP) antibody, the response of neutrophils to LPS/plasma was inhibited but the response to plasma/LPS or fibrinogen/LPS was not. Thus, the LPS in plasma/LPS or fibrinogen/LPS acted on neutrophils in an LBP-independent manner. We conclude that the CD14-dependent LPS receptor system of neutrophils was capable of working in the absence of LBP, but only when LPS was immobilized on a surface coated with protein.
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PMID:Neutrophils responded to immobilized lipopolysaccharide in the absence of lipopolysaccharide-binding protein. 971 56

Calcification of vascular tissue is a common complication in aging, atherosclerosis, diabetes, renal failure, aortic stenosis, and prosthetic valve replacement. Osteopontin is a noncollagenous adhesive protein routinely found at sites of dystrophic calcification and synthesized at high levels by macrophages in calcified aortic valves and atherosclerotic plaques. In the present study, we have characterized the calcification of bovine aortic smooth muscle cell (BASMC) cultures in vitro and have studied the effects of exogenous osteopontin on mineral deposition. Induction of calcification in BASMC cultures was alkaline phosphatase-dependent and was characterized by a multilayer cell morphology. Mineral deposition occurred in the basal matrix of multilayered areas as indicated by von Kossa staining, and transmission electron microscopy and electron diffraction identified the mineral as apatite. Ultrastructural analysis of the cultures showed the presence of extracellular matrix vesicles, calcifying collagen fibrils, and nodular-type calcifications similar to those found in calcified heart valves and atherosclerotic plaques. Purified osteopontin (0.05 to 5 microgram/mL) dose dependently inhibited calcification of BASMC cultures, whereas vitronectin and fibronectin had no effect. In contrast to the inhibitory mechanism of levamisole on mineral deposition, osteopontin did not inhibit alkaline phosphatase activity or reduce phosphorus levels in the culture medium. Addition of calcium to the cultures overcame the inhibitory effect of osteopontin on BASMC culture calcification and resulted in decreased levels of calcium in the culture medium and increased levels in the cell layer. Moreover, using high-resolution, colloidal-gold immunocytochemistry, osteopontin was found intimately associated with growing apatite crystals. These data indicate that the effect of osteopontin, although calcium-dependent, was not mediated by simple calcium chelation but most likely by direct interaction of osteopontin with crystal surfaces. These studies suggest that BASMCs can be used to model vascular calcification in vitro and that soluble osteopontin released near sites of vascular calcification may represent an adaptive mechanism aimed at preventing vascular calcification.
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PMID:Calcification of vascular smooth muscle cell cultures: inhibition by osteopontin. 993 48

Integrin heterodimers sharing the common alphaV subunit are receptors for adhesion glycoproteins such as vitronectin and fibronectin. They are suggested to play an essential role in cell anchoring, differentiation, and survival. Here, we describe the construction of an expression plasmid coding for an intracellular single-chain antibody against alphaV integrin subunit. Saos-2 osteosarcoma cells transfected with this DNA construct showed an approximately 70-100% decrease in the cell surface expression of alphaVbeta3 and alphaVbeta5 integrins as shown by flow cytometry. Intracellular antibody expression had no effect on the mRNA levels of alphaV integrin. Pulse chase experiments of metabolically labeled integrins showed that the translation of precursor alphaV integrin subunit was not affected. However, the maturation of alphaV integrins as glycoproteins was slow suggesting that the transport from endoplasmic reticulum to Golgi complex was partially prevented. Depletion of alphaV integrins from Saos-2 cells led to a decreased ability to spread on fibronectin and vitronectin. Furthermore, the expression of osteoblast differentiation marker genes, alkaline phosphatase and osteopontin, was induced and concomitantly the expression of matrix metalloproteinase-2 decreased. Thus, alphaV integrins seem to be important regulators of osteosarcoma cell phenotypes. Our data also indicate that the expression of intracellular antibodies is an effective strategy to study the significance of specific integrins for cell phenotype and differentiation.
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PMID:Depletion of alphaV integrins from osteosarcoma cells by intracellular antibody expression induces bone differentiation marker genes and suppresses gelatinase (MMP-2) synthesis. 1042 43

Osteoclasts from a patient affected by osteopetrosis were examined in vivo and in vitro. Iliac crest biopsy revealed an osteosclerotic pattern, with prominent numbers of osteoclasts noted for hypernuclearity and incomplete adherence to the bone surface. A population comprising tartrate-resistant acid phosphatase (TRAP)-positive, multinucleated and mononuclear cells, and alkaline phosphatase-positive stromal fibroblasts was obtained in vitro from bone marrow. Mononuclear TRAP-positive precursors spontaneously fused in culture to form giant osteoclast-like cells. These cells expressed the osteoclast marker MMP-9 and calcitonin receptor, and lacked the macrophage marker, Fc receptor. Expression and distribution of c-src, c-fms, and CD68, and response to steroid hormones relevant to osteoclast differentiation and function were apparently normal, whereas cell retraction in response to calcitonin was impaired. TRAP-positive multinucleated cells did not form osteoclast-specific adhesion structures (clear zone, podosomes, or actin rings). Bone resorption rate was severely reduced in vitro. Focal adhesions and stress fibers were observed en lieu of podosomes and actin rings. Adhesion structures contained low levels of immunoreactive vitronectin receptor, most of this integrin being retained in cytoplasmic vesicles. These data provide the first characterization of abnormal differentiation and function of human osteopetrotic osteoclast-like cells.
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PMID:Mechanisms of osteoclast dysfunction in human osteopetrosis: abnormal osteoclastogenesis and lack of osteoclast-specific adhesion structures. 1062 70

Human monocytes (Mo) in culture can be differentiated into macrophages (M phi), dendritic cells (DC) and osteoclasts. In addition, we have established a Mo-derived in vitro granuloma model which here was compared with ex-vivo isolated foreign body granuloma cells. In these models overlapping phenotypes developed between monocyte-derived dendritic cells (MoDC), osteoclasts, M phi, and osteoblasts. In Mo cultures granulomas were induced by immobilized particulate material. AP activity (osteoblast marker) was found to be co-expressed with cytoplasmic tartrate resistant acid phosphatase (TRAP) as a marker of osteoclasts. While proliferating, the number of AP+ cells decreased, being replaced by cells co-expressing the osteoclast markers vitronectin receptor (VNR) and TRAP. Coexpression of the Mo/M phi marker CD68 with AP or VNR confirmed the monocytic origin of the cells. When Mo were treated with interleukin-4 (IL-4), the number of AP+ cells markedly increased and remained stably expressed over 12 days. In explants from ex vivo granulomas obtained from endoprosthetic revisions the major cell type was the AP+ cell co-expressing CD68. The bone-specific alkaline phosphatase (BAP) as a marker of osteoblasts was detected by FACS analysis in the ex vivo granuloma cells. By RT-PCR the mRNA for osteocalcin, which is a highly specific marker for osteoblasts, was detected. From our results we conclude an ontogenetic relationship between macrophages, DC and osteoclasts. Furthermore, the data suggest a transdifferentiation between Mo and osteoblasts.
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PMID:Alkaline phosphatase expression during monocyte differentiation. Overlapping markers as a link between monocytic cells, dendritic cells, osteoclasts and osteoblasts. 1087 91


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