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:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of ascorbic acid deficiency on growth and calcification of bone were studied in whole 18-day fetal rat radii and ulnae cultured in a chemically defined medium. Ascorbic acid deficiency decreased the formation of labeled hydroxyporline from labeled proline in both bone shafts and cartilage ends while incorporation of tryptophan was maintained. Dry weights and collagen content of bone and cartilage were decreased, but calcification was not affected. The optimun initial concentration of ascorbic acid for collagen synthesis was 200 mug/ml. The effect of ascorbic acid was not antagonized by glucoascorbic acid or replaced by dithiothreitol. Decreased collagen synthesis in ascorbic acid deficiency could not be ascribed to loss of available peptidyl proline hydorxylase. Formation of underhydroxylated collagen and its release into the medium accounted for much of the decrease in hydroxylated collagen in ascorbic acid deficient bones. Nevertheless, the total newly synthesized collagen, as measured by collagenase digestion, was still decreased. Similar effects were exerted by alpha, alpha'-dipyridyl which also inhibited general protein synthesis. Ascorbic acid did not stimulate proline incorporation into collagen in the presence of alpha, alpha'-dipyridyl.
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PMID:The effects of ascorbic acid deficiency on calcium and collagen metabolism in cultured fetal rat bones. 16 34

Collagen is produced by WI-38 diploid human fibroblast cultures throughout their life cycle. It is examined by a sensitive method based on the analysis of specific peptides obtained after digestion with bacterial collagenase. The production and hydroxylation of the collagen is strongly dependent upon the age (population doublings) of the culture and the presence of ascorbic acid. Young cultures (passage 26) produce large amounts of collagen in the absence of ascorbic acid, and this collagen is about 50% hydroxylated compared to that produced by young cultures in the presence of ascorbic acid. Ascorbic acid reduces to about one-half the amount of collagen produced by these young cultures. The young confluent cultures also depend strongly on ascorbic acid for hydroxylation of proline. The dependence declines rapidly with the age of the culture. The collagen produced by young cultures supplied with ascorbic acid is very similar to the type I collagen produced by normal individuals and has about the same degree of hydroxylation of its prolyl residues. The amount of collagen produced by "older" cultures is unaffected by ascorbic acid, but the degree of hydroxylation is normal only if ascorbic acid is present, and is decreased to about 60 to 70% in the absence of the vitamin. "Senescent" cultures showed little, if any, dependency on ascorbic acid, and the collagen produced, with and without the vitamine, is about 80% hydroxylated. The prolyl hydroxylation system of the WI-38 cells and the various controls on the system are age-dependent.
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PMID:Collagen synthesized and modified by aging fibroblasts in culture. 118 37

Interstitial collagenase either obtained from human neutrophils by phorbol myristate acetate (PMA) induced degranulation or isolated from human gingival crevicular fluid was found to be activated by addition of an oxidative agent, hypochlorous acid (HOCl). Collagenase released by PMA stimulated neutrophils was completely in latent form but underwent partial autoactivation during 16 h incubation at 22 degrees C in the presence of soy bean trypsin inhibitor. The partial autoactivation was potentiated to complete activation of released collagenase after addition of exogenous HOCl. Ascorbate prevented this activation of neutrophil collagenase. Isolated human gingival crevicular fluid collagenase represented an apparent Mr of 70 kD in completely latent form, whereas 70/54 kD enzyme species were detected for partially autoactive form of the enzyme. Western blot analysis of gingival crevicular fluid using a polyclonal antibody raised against purified human neutrophil collagenase revealed the same 70/54 kD molecular forms of the enzyme. The latent gingival crevicular fluid collagenase was also activated by HOCl and this activation could be prevented by ascorbate. Activation of the 70 kD latent collagenase by HOCl as well as by other non-proteolytic activators such as an organomercurial compound (phenylmercuric chloride) and a gold(I) compound (gold thioglucose) was not associated with detectable changes in apparent Mr, whereas trypsin activation resulted in fragmentation of 70 kD enzyme to 54 kD species. Our results provide further evidence for the neutrophil origin of gingival crevicular fluid collagenase and suggest that, in addition to proteolytic activation, oxidative and antioxidative agents seem to be able to regulate neutrophil collagenase activity.
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PMID:Hypochlorous acid induced activation of human neutrophil and gingival crevicular fluid collagenase can be inhibited by ascorbate. 166 35

Ascorbic acid is the active component of fetal brain extract that induces increased acetylcholine receptor (AChR) expression in L5 rat clonal muscle cell cultures. The induction of AChR expression, as determined by 125I-alpha-bungarotoxin binding, occurs with a delay of 20-25 h. We report that the delayed increase in AChR can be triggered by a 5-h exposure to ascorbic acid. These studies suggest that intermediary processes may be involved. Ascorbic acid treatment also causes a threefold increase in collagen secretion in L5 cultures by 3 h. The rapid increase in collagen secretion and the delayed induction of surface AChR suggested that there may be a link between these two responses. However, although bacterial collagenase eliminates secreted collagen, it had no effect on the increase in surface AChR. Thus, the ascorbic acid effect on elevating AChR expression is independent of its effect on collagen secretion.
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PMID:Acetylcholine receptor regulation in L5 muscle cells is independent of increases in collagen secretion induced by ascorbic acid. 196 67

The effect of ascorbate on the glycosaminoglycans synthesized by normal and simian virus 40(SV40)-transformed human skin fibroblasts was examined. Cells were incubated in the presence or absence of ascorbate, and radiolabelled with [3H]glucosamine and [35S]sulphate for 48 h, 3 days after reaching confluence. Glycosaminoglycans were analysed in the medium, a collagenase extract, and in the trypsin/cell-associated fraction. Hyaluronic acid was the main 3H-labelled glycosaminoglycan in all but the collagenase extracts, and showed a large decrease in normal fibroblast cultures, but a significant increase in SV40-transformed fibroblast cultures following feeding with ascorbate. Incorporation of [3H]glucosamine into sulphated glycosaminoglycans was reduced in normal fibroblast cultures but increased slightly in SV40-transformed cultures following ascorbate supplementation. [35S]sulphate incorporation remained essentially unaltered in both cell cultures. Ascorbate stimulated the deposition of glycosaminoglycans into the insoluble matrix of normal fibroblasts while reducing the deposition in SV40-transformed fibroblast cultures. The observed changes may in part be related to ascorbate-induced deposition of collagen in normal fibroblast cultures and the inability of the transformed fibroblast cells to deposit an extensive extracellular matrix, in addition to possible changes in the specific activity of the UDP-N-acetyl-[3H]hexosamine pool.
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PMID:Ascorbate induced changes in glycosaminoglycan synthesis and distribution of normal and SV40-transformed fibroblasts. 302 33

Ascorbate and beta-aminopropionitrile (BAPN) have direct, but diverse affects on collagen matrix production. Ascorbate is necessary for the intracellular hydroxylation of prolyl and lysyl residues during collagen biosynthesis whereas BAPN inhibits the enzyme lysyl oxidase in the extracellular space thus preventing collagen crosslink formation. To study the influence of these two agents on fibroplasia, an in vitro model was used to analyze fibroblast migration, proliferation, and collagen synthesis. Biopsies of chicken tendon were covered with a fibrin clot to simulate an in vivo wound environment, and then they were exposed to either ascorbate or BAPN for up to 7 days. Fibroblast migration into the fibrin clot was measured using a Zeiss Mopp II planimeter, DNA synthesis by 125IUDR incorporation, and collagen synthesis by [3H]proline incorporation into collagenase-digestible protein. Tendon biopsies treated daily with fresh ascorbate (0.1 mM) had significantly greater fibroblast migration than controls without ascorbate (P less than 0.05). Cellular proliferation, collagen synthesis, and total protein synthesis were not significantly altered by ascorbate treatment. In contrast, BAPN inhibited fibroblast migration in a dose-dependent fashion without inhibiting proliferation (0.25 and 0.5 mM), collagen, and noncollagen protein synthesis. Therefore, the effect of BAPN on migration does not appear to be due to generalized cytotoxicity. These combined studies suggest that compounds such as ascorbate and BAPN which can modify collagen may also modify fibroblast migration.
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PMID:Effect of beta-aminopropionitrile and ascorbate on fibroblast migration. 339 46

1. Cell cultures propagated from foetal bovine ligamentum nuchae synthesized and secreted two glycoproteins, designated MFP I and MFP II, that are closely related to elastic-fibre microfibrils. Glycoproteins MFP I (apparent mol.wt. 150 000) and MFP II (apparent mol.wt. 300 000) were metabolically labelled, separated from other culture-medium components by immunoprecipitation with a specific anti-(microfibrillar protein) serum, and analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and sodium dodecyl sulphate/gel-filtration chromatography. 2. Ligament cells also synthesized and secreted fibronectin, but salt-fractionation and immunoprecipitation studies with a specific anti-(cold-insoluble globulin) serum established that neither glycoprotein MFP I nor glycoprotein MFP II was related to fibronectin. 3. The secretion of glycoprotein MFP I, but not that of glycoprotein MFP II, was enhanced by the addition of ascorbate to the culture medium. 4. Ascorbate-supplemented ligament cells incorporated [3H]proline into glycoprotein MFP I, and 36% of the nondiffusible proline residues were hydroxylated, exclusively as 4-hydroxy[3H]proline. Less than 1% of the total proline residues in [3H]proline-labelled glycoprotein MFP II were hydroxylated. 5. Ascorbate-supplemented cells incorporated [14C]lysine into glycoprotein MFP I and 30% of the non-diffusible lysine residues were hydroxylated. 6. Newly secreted glycoprotein MFP I was digested by highly purified bacterial collagenase to yield polypeptide fragments of apparent mol.wts. 50 000 and 30 000. Glycoprotein MFP II was not digested by bacterial collagenase. 7. The results suggest that elastic-fibre microfibrils are composed of a novel collagenous glycoprotein MFP I in association, as yet undefined, with a non-collagenous glycoprotein MFP II.
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PMID:The nature of the microfibrillar glycoproteins of elastic fibres. A biosynthetic study. 627 35

In confluent human skin fibroblasts maintained in 0.5% serum-supplemented medium. L-ascorbate specifically stimulated the rate of incorporation of labeled proline into total collagenase-sensitive protein, without changing the specific activity of the intracellular free proline. This influence of ascorbate reached a maximum at 30 microM and continued for at least 4 days, resulting in a 4-fold increase. The ascorbate effect occurred in cells at both confluent and subconfluent densities and was evident at all serum concentrations from 0.5-20%. The effect was independent of duration of the radioactive pulse between 2-6 h. D-Ascorbate, D-isoascorbate, and L-dehydroascorbate also stimulated collagen synthesis but at considerably higher concentrations, i.e., 250-300 microM. The stimulation of collagen synthesis by ascorbate and its analogs was accompanied by a decline in prolyl hydroxylase activity and a rise in lysyl hydroxylase activity; again L-ascorbate was found to be most effective. Dimethyltetrahydropterine and L-lactate failed to produce these effects.
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PMID:Collagen synthesis in cultured human skin fibroblasts: effect of ascorbic acid and its analogs. 630 3

The question whether ascorbate regulates collagen production solely through its direct role in proline hydroxylation was investigated. Proteins in calvarial bones from control and scorbutic weanling guinea pigs were labeled in short-term cultures with radioactive proline. Proteins were digested with purified bacterial collagenase to distinguish between effects on collagen polypeptide production and hydroxyproline formation. There was a preferential decrease in the absolute rate of collagen biosynthesis beginning after 2 wk of ascorbate deficiency, and this effect was temporally dissociated from decreased proline hydroxylation. There were no significant changes in the absolute rates of collagen degradation or noncollagen protein production. In vitro inhibition of proline hydroxylation in normal bone with alpha, alpha'-dipyridyl did not affect the relative rate of collagen synthesis, further dissociating these functions. Ascorbate added to scorbutic bone cultures reversed defective proline hydroxylation but not defective collagen synthesis, suggesting that the latter was an indirect effect of scurvy. There was a linear correlation between the extent of body weight lost during the 3rd and 4th wk of scurvy and the rate of collagen synthesis in scorbutic bone. This correlation also applied to control animals receiving ascorbate, but with weight loss induced by food restriction. These studies establish for the first time that ascorbate deficiency in guinea pigs leads to a specific decrease in collagen polypeptide synthesis and suggest that this decrease results from the reduced food intake and/or weight-loss characteristic of scurvy.
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PMID:Specifically decreased collagen biosynthesis in scurvy dissociated from an effect on proline hydroxylation and correlated with body weight loss. In vitro studies in guinea pig calvarial bones. 630 11

The effect of ascorbic acid on the synthesis, distribution and sulphation of glycosaminoglycans by human skin fibroblasts has been examined. Medium was supplemented with ascorbate over several days, and cultures incubated with [3H]glucosamine and Na2(35)SO4 for 48 h, followed by analysis of the glycosaminoglycans in the medium, in collagenase and trypsin extracts, and in cell fractions. Ascorbate feeding resulted in a reduction in hyaluronate synthesis, which was the main 3H-labelled component and was distributed mainly in the medium fractions. Sulphated glycosaminoglycans showed a reduction in incorporation of 3H label, but increased sulphation following ascorbate feeding. In control cultures 53% of 3H-labelled sulphated glycosaminoglycans and 63% of 35S-labelled glycosaminoglycans were present in the medium fraction, while in ascorbate-fed cultures, 41% of 3H label and 38% 35S label were incorporated into medium-sulphated glycosaminoglycans. Ascorbate also caused an increase in cell density and in collagen production and deposition.
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PMID:Changes in the synthesis, distribution and sulphation of glycosaminoglycans of cultured human skin fibroblasts upon ascorbate feeding. 642 Apr 22


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