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)

The ultrastructure of crystal formation in vitro associated with extracellular membrane-bound matrix vesicles (MV) isolated from rat incisor pulp was studied in Dulbecco's modified Eagle's medium (DMEM) supplemented with an organic phosphate, Na-beta-glycerophosphate (BGP). Matrix vesicles were isolated from basal regions of the pulps using a collagenase digestion and ultra-centrifugation method. Isolated MV contained alkaline phosphatase (ALP) activity and had diameters of 30-200 nm. Membrane structures of the isolated MV were well preserved. Incubation of MV in DMEM in the presence of BGP caused the development of bilaminar electron densities associated with the vesicle membrane. These preceded crystal deposition which was observed in the culture medium after 3 days. Both heat-inactivated MV incubated with BGP, and fresh MV incubated in the absence of BGP failed to show crystal formation, even after 3 days. Staining of demineralized sections of mineralized MV with uranyl acetate and lead citrate, revealed numerous needle-like structures similar in shape to the untreated crystals. Electron diffraction patterns of the newly formed crystals revealed a pattern consistent with hydroxyapatite. The requirement of BGP for mineralization of these MV and the long lag time before crystal formation is probably due to the low calcium (Ca) x inorganic phosphate (Pi) ion product in the original medium. The requirement of ALP activity which would cause hydrolysis of BGP and a rise in Pi would favor the precipitation of biologic apatite from the culture medium.
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PMID:Matrix vesicles isolated from apical pulp of rat incisors: crystal formation in low Ca x Pi ion-product medium containing beta-glycerophosphate. 196 82

High levels of interleukin-6 (IL-6) have been detected in synovial fluid from patients with inflammatory arthropathies associated with local bone resorption, suggesting a role for IL-6 as a local regulator of bone resorption and remodeling. In the present study we examined the effects of IL-6 on [3H]thymidine ([3H]TdR) incorporation, collagen synthesis, and alkaline phosphatase activity in UMR-106-01 rat osteoblastic osteosarcoma cells. IL-6 stimulated a dose-dependent increase in [3H]TdR incorporation that was maximal at 1000 U/ml (-147% of basal, p less than 0.005) in osteoblastlike cells that were in a logarithmic phase of growth. The increase in [3H]TdR incorporation was maximal between 12 and 24 h and was neutralized by pretreatment with the polyclonal rabbit antibody to IL-6. IL-6 also increased cell number and the secretion of prostaglandin E2 in UMR-106-01 cells in logarithmic growth phase. The stimulation of [3H]TdR incorporation and release of PGE2 into the culture medium by IL-6 was inhibited by indomethacin. A 24 h exposure of the osteoblastlike cells to 1000 U/ml of IL-6 reduced [3H]proline incorporation into collagenase-digestible (CDP) protein to 73% of control values (p less than 0.01). Noncollagen protein (NCP) synthesis was inhibited to 80% of control values (p less than 0.01) by 1000 U/ml of IL-6. The inhibitory effect was relatively greater on CDP than on NCP and consequently resulted in a decrease in the percentage of collagen synthesis. Alkaline phosphatase activity was not altered in these cells after a 24 h exposure to 1-1000 U/ml of IL-6.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of interleukin-6 on cellular function in UMR-106-01 osteoblastlike cells. 202 35

The nature and tissue distribution of non-collagenous bone proteins synthesized by adult rat bone marrow cells, induced to differentiate in the presence of dexamethasone (DEX) and beta-glycerophosphate (beta-GP), was studied in vitro to determine the potential role of these proteins in bone formation. Northern hybridization analysis revealed a strong induction of bone sialoprotein (BSP) and osteocalcin in DEX-treated cultures, whereas the constitutive expression of secreted phosphoprotein I (SPP-1), type I collagen, SPARC, and alkaline phosphatase was stimulated 6-, 5-, 3-, and 2.5-told, respectively. Metabolic labeling of proteins showed that the sialoproteins (SPP-1 and BSP) were mostly secreted into the culture medium in the non-mineralizing (-beta-GP) cultures, but were the predominant non-collagenous proteins associated with the hydroxyapatite of the bone nodules in mineralizing cultures (+ beta-GP). Extraction of the tissue matrix with 4 M GuHCl and digestion of the demineralized tissue matrix with bacterial collagenase revealed that some BSP was also associated non-covalently and covalently with the collagenous matrix. SPP-1 was present in two distinct, 44 kDa and 55 kDa, forms in the conditioned medium of all cultures and was preferentially associated with the hydroxyapatite in the mineralizing cultures. In comparison, SPARC was abundant in culture media but could not be detected in de-mineralizing extracts of the mineralized tissue. Radiolabeling with [35SO4] demonstrated that both SPP-1 and BSP synthesized by bone cells are sulfated, and that a 35 kDa protein and some proteoglycan were covalently associated with the collagenous matrix in +DEX cultures. Labeling with [32PO4] was essentially confined to the sialoproteins; the 44 kDa SPP-1 incorporating significantly more [32PO4] than the 55 kDa SPP-1 and the BSP. These studies demonstrate that BSP and osteocalcin are only expressed in differentiated osteoblasts and that most of the major non-collagenous bone proteins associate with the bone mineral. However, some novel proteins together with some of the BSP are associated with the collagenous matrix where they can influence hydroxyapatite formation.
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PMID:Expression of bone matrix proteins associated with mineralized tissue formation by adult rat bone marrow cells in vitro: inductive effects of dexamethasone on the osteoblastic phenotype. 203 18

Human bone cells were obtained as the outgrowth from cancellous bone fragments pretreated with collagenase and DNase. The osteogenic potential of cells in primary culture was assessed upon intramuscular transplantation into young mice pretreated with cortisone. Transplants were recovered after 2 weeks and examined by light microscopy. Of 34 transplants, 6 showed evidence of osteogenesis and 12 the production of unmineralized matrix. Only cells were observed in the other transplants. In an attempt to find a biochemical marker for osteogenic cells we have assayed medium osteocalcin and alkaline phosphatase activity levels in cultures before transplantation. No correlation was found between the level of expression of the two osteoblast markers and the osteogenic potential of the cells.
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PMID:In vivo osteogenic activity of isolated human bone cells. 204 31

Abnormalities of tubular membrane structure and composition have been proposed as the primary defect in nephronophthisis (NEF). In order to characterize the protein composition of tubular cells in NEF, in vitro methods were developed to culture and propagate tubular cells obtained from biopsy fragments. Accordingly, microdissected cortical slices (1 x 3 mm) were first digested with collagenase and DNAse and then grown in RPMI medium supplemented with 10% NU serum and conditioned serum deriving from 3T3 cultures. At confluence, cultured cells from NEF showed characteristics which were typical of normal tubules, i.e. presence of cytokeratin and positivity for succinic dehydrogenase and alkaline phosphatase stainings, and presented no morphological alterations compared to cultured cells from normal tubular epithelium. Moreover, no difference was observed for fibronectin, collagen IV and laminin stains. Analysis by two-dimensional electrophoresis of cellular extracts revealed several changes in protein composition of NEF, the main one being the decrease in NEF cells of a polypeptide with a molecular weight of 120 kD and a pI of 4.8; this polypeptide was a constant finding in normal kidneys. These observations demonstrated that human tubular epithelial cells can be successfully cultured from very small biopsy fragments, which represents a new approach to the study of molecular disorders involving tubular cells in inherited disease. Cultured cells from NEF maintain the same morphological, immunological and cytochemical characteristics as normal tubular cells, but present a few alterations in polypeptide composition which may have pathogenetic relevance. A more careful analysis of these alterations is needed to define the molecular disorder(s) involving the tubule in NEF.
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PMID:Tubular epithelium culture from nephronophthisis-affected kidneys: a new approach to molecular disorders of tubular cells. 207 4

The effect of transforming growth factor-beta (TGF-beta) on proliferation and differentiation of mouse clonal osteoblastic cells (MC3T3-E1) was examined in vitro in three different stages of their differentiation. Stage I (1-3 days after plating) was characterized by rapid cell growth, negligible alkaline phosphatase (ALP) activity and high proteoglycan synthesis, but low collagen production. In stage II (3-5 days after plating), proteoglycan synthesis sharply decreased and ALP activity and collagen synthesis began to increase. Stage III (7-9 days after plating) was characterized by maximal osteoblastic phenotypes. Treating MC3T3-E1 cells with 1 ng/ml of TGF-beta greatly inhibited DNA synthesis in stage I but not in stage II. In contrast, TGF-beta dose-dependently stimulated the synthesis of collagenase digestible proteins (CDP), noncollagenous proteins (NCP) and proteoglycan, especially in stage II. The minimum effective dose of TGF-beta in this stage was as low as 0.04-0.2 ng/ml. In stages I and III, the MC3T3-E1 cells were rather insensitive to TGF-beta in increasing three osteoblastic phenotypes. The increase in ALP activity in stages II and III was inhibited by 1 ng/ml of TGF-beta. These results indicate that the response to TGF-beta of mouse clonal osteoblastic MC3T3-E1 cells changes depending on their maturation stages.
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PMID:Transforming growth factor-beta modulates proliferation and differentiation of mouse clonal osteoblastic MC3T3-E1 cells depending on their maturation stages. 208 82

Bone remodeling requires bone resorption and formation. Bone formation involves osteoblastic activity. Both intermittent and continuous mechanical forces have been proved to affect osteoblast differentiation in vivo. It is evident in the present investigation that periodically applied tension alter cell division, protein synthesis and alkaline phosphatase activity. An initial seeding density of 50,000 cells/well was used for both the mechanically stimulated cells and controls. For the experimental cultures, the cells were subjected to different levels of tension applied at different frequencies; 1. Low frequency (0.3 cycles/minute) at moderate tension (17% elongation); 2. Moderate frequency (3 cycles/minute) at moderate tension (17% elongation); 3. High frequency (30 cycles/minute) at moderate tension (17% elongation); 4. Low tension (9% elongation at moderate frequency (3 cycles/minute); 5. High tension (22% elongation) at moderate frequency (3 cycles/minute). Cell numbers, protein synthesis and alkaline phosphatase activity were measured for each treatment combination, as well as for the unstressed controls. The cells were harvested at 24, 72, and 120 hours after the start of force application. Analysis of variance revealed that periodically applied tension stimulated cell activity in cultured UMR 106-01 cells: 1. Seventeen percent elongation at 3 and 30 cycles/minute was the optimum condition for stimulating osteoblastic cell division; and 2. High frequency stimulation at moderate tension and high tension at moderate frequency stimulated alkaline phosphatase activity. Given these findings, the following conclusions seem warranted: 1. There may be an optimal magnitude of tensional force that stimulates osteoblast activity. Beyond this optimal level, osteoblasts may lose potential to have cell divisions; 2. The optimal condition of tension might be different for cell division and other cell functions. Although high tension (22% elongation) did not stimulate cell division, alkaline phosphatase activity showed significantly high measurement at this condition; and 3. in the light of both the present study and previous investigations, periodically applied tension may stimulate both collagenase (a parameter of bone resorption) and alkaline phosphatase (a parameter of bone formation) production.
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PMID:[Effects of periodic tension on osteoblast-like cells for cell differentiation and alkaline phosphatase activity]. 213 80

Effects of transforming growth factor (TGF)-beta, epidermal growth factor (EGF), insulin, 1, 25-dihydroxyvitamin D3 (1, 25 (OH)2D3), and parathyroid hormone (PTH) on the proliferation and differentiation of clonal dental pulp cells of rats were investigated. Interaction between growth factors (TGF-beta and EGF) and two hormones insulin and 1, 25 (OH)2D3, which have been noticed to accelerate the differentiation of the cells, were also studied, and the following results were obtained: 1) TGF-beta decreased alkaline phosphatase (ALPase) activity in a dose-dependent manner, and the inhibitory effect was not blocked by indomethacin, suggesting that the effect of TGF-beta on the cells may not be mediated by prostaglandins. Inhibitory effects of ALPase antagonists (L-phenylalanine, L-homoarginine, levamisole) on the activity were not affected by TGF-beta. TGF-beta showed no evident effect on the DNA synthesis (incorporation of [3H] thymidine) and collagen synthesis (incorporation of [2, 3-3H] proline into the collagenase-digestible protein) of the cells. 2) EGF stimulated the incorporation of [3H] thymidine and inhibited the ALPase activity. The inhibitory effect was not blocked by indomethacin, indicating that the EGF effect is not mediated by prostaglandins. Collagen synthesis was significantly inhibited by EGF. 3) Insulin showed a weak but significant inhibition of the DNA synthesis. Insulin increased the ALPase activity evidently, and accelerated the collagen synthesis significantly. 4) The vitamin 1, 25 (OH)2D3 significantly increased the ALPase activity though no significant changes were observed in the DNA synthesis and collagen synthesis. 5) PTH had no evident effect on the DNA synthesis and ALPase activity, but did tend to accelerate the collagen synthesis. 6) A study on the interaction between insulin and EGF or TGF-beta revealed that the acceleration of DNA synthesis induced by EGF was inhibited when the factor was combined with insulin, and the increase in ALPase activity elicited by insulin was inhibited by EGF and weakened by TGF-beta significantly when these factors were added simultaneously with the insulin. Or viewed another way, the inhibitory effect of EGF or TGF-beta on the ALPase activity was antagonized by insulin. The accelerative action of insulin on collagen synthesis was antagonized by EGF and potentiated by TGF-beta. 7) A study on the interaction between 1, 25 (OH)2D3 and EGF or TGF-beta revealed that 1, 25 (OH)2D3 inhibited the accelerating effect of EGF on the DNA synthesis and that the increasing effect of 1, 25 (OH)2D3 on ALPase activity was strongly inhibited by EGF.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:[Effects of various growth factors and hormones on clonal rat pulp cells]. 213 79

Two test teeth, anteriors with greater than or equal to 6 mm deep periodontal pockets from each of 10 patients with advancing periodontitis were included in this study. The clinical signs of advancing periodontitis, generalized moderate to deep pockets and to severe loss of alveolar bone, were observed in young adult. There have been several reports on factors, which reflect the conversion clinically from infection by highly pathogenic plaque bacteria to a from of periodontitis displaying relatively rapid loss of clinical attachment. The purpose of this investigation was to detect parameters in fluid, which could leak from the underlying inflamed connective tissue into the gingival crevice, and which could shown correlatively the progressive variations of periodontal disease by recurrent acute stage. In order to determine active disease sites and to monitor guantitatively response to therapy or to measure degree to susceptibility of future breakdown. Examinations on following parameters at pre- and post- periodontal treatment stages were carried out. Endotoxin, collagenase, alkaline phosphatase, beta-glucuronidase, interleukin-1 alpha, IgG antibody levels to Bacteroides gingivalis, Bacteroides intermedius were measured in gingival exudate samples, which were collected by the microtips technique from periodontal pockets. The following results were obtained: 1) Considering the effect of periodontal therapy, pathogenic responses on total colony forming unit (CFU), interleukin-1 alpha and changes of endotoxin and beta-glucuronidase levels after the treatment have indicated that specific changes in humoral responses. 2) There was not significant relation between alkaline phosphatase, collagenase, IgG antibodies level to Bacteroides gingivalis, Bacteroides intermedius and responses in active and also inactive disease sites. 3) This study has been resulted in the development of diagnostic techniques which requires strict criteria on the disease activity of the periodontal disease very specific in order to permit a more scientific approach to the care of periodontitis patients and to speculate the prognosis of the patients after the treatment.
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PMID:[Changes of factors on disease activity in advancing periodontitis]. 213 9

Cells were isolated by sequential collagenase digestion from the parietal segments of one day old mice (Swiss albino BNL strain) and characterized for osteoblast parameters by alkaline phosphatase histochemistry and bovine parathyroid hormone (bPTH-(1-34] induced cAMP activity (protein binding assay). Phenytoin (DPH) reduced PTH stimulated cAMP activity nearly 3-fold in the presence and nearly 1.5-fold in the absence of added calcium. In the absence of PTH, DPH exerted no significant effect. Bay-K-8644, a calcium channel activator, appeared to approximate the PTH stimulation of cAMP activity, even in the presence of DPH. This study demonstrates that DPH has a direct effect on PTH stimulated cAMP activity in cultured murine osteoblasts.
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PMID:The effect of phenytoin on parathyroid hormone stimulated cAMP activity in cultured murine osteoblasts. 215 57


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