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)

Cultured pulmonary artery smooth muscle cells derived from the medial vessel layer of weanling rabbits were grown in the presence or absence of sodium ascorbate. The connective tissue elements insoluble elastin and collagen were identified and quantified. Formation and accumulation of alpha-aminoadipic acid gamma-semialdehyde (allysine) and the intermolecular cross-links desmosine (Des), isodesmosine (Ides), and aldol condensation product (Aldol) were evaluated from [14C]lysine pulse-chase experiments. [14C]Des, [14C]Ides, peptide-bound [14C]lysine, [14C]allysine, and [14C]Aldol were determined from amino acid analysis. The latter two components were determined after reduction with NaBH4. [14C]Proline conversion to hydroxy[14C]proline and collagenase susceptibility were used to identify and quantify collagen synthesis. Ascorbate dramatically affects insoluble elastin synthesis, accumulation, and cross-link formation. Cells grown in the presence of ascorbate synthesize and accumulate significantly less insoluble elastin than non-ascorbate cultures. Those elastin molecules which do become incorporated into the extracellular matrix in the presence of ascorbate contain a slightly elevated content of hydroxyproline and lysine and, most importantly, are turned over more rapidly.
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PMID:Effects of ascorbate on insoluble elastin accumulation and cross-link formation in rabbit pulmonary artery smooth muscle cultures. 681 21

The addition of 0.2 mM Na L-ascorbate increased the incorporation of 3H-thymidine by rabbit articular chondrocytes in cell and organ culture. The stimulatory response of explants to ascorbate was potentiated by pretreatment of the cartilage with 0.2% clostridial collagenase (type 1) or trypsin for 15-30 minutes. In explants there was a latent period of 3 to 4 days before increased labeling of the nuclei could be detected. The effect was transient and declined after 8 days of culture. It was more evident in organ cultures of immature (3-month-old) than 2- to 3-year-old rabbits. Age differences were not detected in cell cultures. Explants of adult human articular cartilage were stimulated by ascorbate when the medium was supplemented with 10% fresh human serum but not by fetal bovine serum. The findings indicated that synthesis of DNA by articular chondrocytes in situ is regulated by responsiveness of the cells proper to compounds such as vitamin C, by properties of the extracellular matrix, and by factors in the serum. Ascorbate was cytotoxic at concentrations greater than 0.2 mM in the presence of certain batches of serum.
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PMID:Stimulation of DNA synthesis by ascorbate in cultures of articular chondrocytes. 706 59

Several events are associated with cellular aging: alterations in the extracellular matrix, loss of the cell's proliferative capacity, and decreased responsiveness to growth factors. In skin, a major component of the extracellular matrix is collagen; an important regulator of collagen synthesis is ascorbic acid, which may also have growth factor-like properties. To investigate the relationship of the extracellular matrix and proliferative capacity to aging, we examined the effects of ascorbic acid on cell proliferation and collagen expression in dermal fibroblasts from donors of two age classes, newborn (3-8 d old) and elderly (78-93 years old). In the absence of ascorbic acid (control) proliferative capacities were inversely related to age; newborn cell lines proliferated faster and reached greater densities than elderly cell lines. However, in the presence of ascorbic acid both newborn and elderly cells proliferated at a faster rate and reached higher densities than controls. To determine whether there are age-related differences in extracellular matrix production and ascorbic acid responsiveness we examined and found that collagen biosynthesis (collagenase-digestible protein) was inversely related to age, but the stimulation by ascorbic acid appeared age independent. The increase in collagen synthesis was reflected by coordinate increases in steady-state pro alpha 1(I) and pro alpha 1(III) collagen mRNAs, suggesting a pretranslational mechanism. Ascorbic acid appears capable of overcoming the reduced proliferative capacity of elderly dermal fibroblasts, as well as increasing collagen synthesis in elderly cells by similar degrees as in newborn cells even though basal levels of collagen synthesis are age dependent.
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PMID:Effects of ascorbic acid on proliferation and collagen synthesis in relation to the donor age of human dermal fibroblasts. 751 57

To define the influence of time in culture on gene expression for extracellular matrix proteins we have examined the progression of changes in gene expression for chondrocyte extracellular matrix proteins from the time the chondrocytes are initially isolated from their native cartilaginous matrix through 13 days of high-density culture. We have also determined the effect of matrix depletion and shape change by enzymatically resuspending cells after 6 days in culture and sampling the replated cells at intervals up to 13 days. Northern blots of chondrocyte RNA were hybridized with probes for collagen alpha 1(II), alpha 1(I), aggrecan, link protein, and decorin mRNA. The steady-state level of alpha 1(II) collagen mRNA dropped to 45% of the initial value within 24 h, with a further decrease to 21% by Day 3. A similar decline occurred, but less rapidly in ascorbate supplemented cultures with values of 78%, at 24 h and 62% at Day 3. Very low levels of alpha 1(I) collagen mRNA were detectible in cells maintained for 2 weeks without ascorbate supplementation. Type I collagen alpha 1(I) mRNA was not detected in freshly isolated chondrocytes or at the earliest times in culture but was increasingly abundant from Days 5-13 in the presence of ascorbate. Ascorbic acid supplementation altered the pattern of aggrecan expression over time. Without ascorbate there was an increase in steady-state aggrecan mRNA with time in culture, but in the presence of ascorbate, aggrecan mRNA levels peaked at early culture times and progressively diminished. Decorin steady-state mRNA levels in cultures not supplemented with ascorbic acid steadily increased over time in culture following a lag of several days. In cultures treated with ascorbate, however, there was a progressive increase in decorin steady-state mRNA levels from the first day in culture. Resuspending chondrocytes by digestion of the cell layer with pronase and collagenase at Day 6, which resulted in a transient shape change as well as matrix depletion, resulted in a greater than 2-fold increase in alpha 1(II) mRNA at Day 7 in ascorbate supplemented cultures. Only with ascorbate was there a small increase in decorin mRNA at Day 7, after resuspension. Aggrecan mRNA, however, showed a 3-fold increase without ascorbate and a 10-fold increase with ascorbate within 24 h of resuspension. Similarly, link protein steady-state mRNA showed an 8-fold increase without ascorbate and a 9-fold increase with ascorbate within 24 h after resuspension.
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PMID:Modulation of extracellular matrix gene expression in bovine high-density chondrocyte cultures by ascorbic acid and enzymatic resuspension. 794 10

Matrix vesicles, media vesicles, and plasma membranes from three well-characterized, osteoblast-like cells (ROS 17/2.8, MG-63, and MC-3T3-E1) were evaluated for their content of enzymes capable of processing the extracellular matrix. Matrix vesicles were enriched in alkaline phosphatase specific activity over the plasma membrane and contained fully active neutral, but not acid, metalloproteinases capable of digesting proteoglycans, potential inhibitors of matrix calcification. Matrix vesicle enrichment in neutral metalloproteinase varied with the cell line, whereas collagenase, lysozyme, hyaluronidase, and tissue inhibitor of metalloproteinases (TIMP) were not found in any of the membrane fractions examined. MC-3T3-E1 cells were cultured for 32 days in the presence of ascorbic acid (100 micrograms/ml), beta-glycerophosphate (5 mM), or a combination of the two, to assess changes in matrix vesicle enzymes during calcification. Ascorbate or beta-glycerophosphate alone had no effect, but in combination produced significant increases in both active and total neutral metalloproteinase in matrix vesicles and plasma membranes, with the change seen in matrix vesicles being the most dramatic. This correlated with an increase in the formation of von Kossa-positive nodules. The results of the present study indicate that osteoblast-like cells produce matrix vesicles enriched in proteoglycan-degrading metalloproteinases. In addition, the observation that matrix vesicles contain significantly increased metalloproteinases under conditions favorable for mineralization in vitro lends support to the hypothesis that matrix vesicles play an important role in extracellular matrix processing and calcification in bone.
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PMID:Matrix vesicles produced by osteoblast-like cells in culture become significantly enriched in proteoglycan-degrading metalloproteinases after addition of beta-glycerophosphate and ascorbic acid. 806 58

Treatment of mouse MC3T3-E1 cells with ascorbic acid initiates the formation of a collagenous extracellular matrix and synthesis of several osteoblast-related proteins. We recently showed that ascorbic acid dramatically increases alkaline phosphatase and osteocalcin mRNAs and that this induction is blocked by inhibitors of collagen triple-helix formation (Franceschi and Iyer, J Bone Miner Res 7:235). In the present study, the relationship between collagen matrix formation and osteoblast-specific gene expression is explored in greater detail. Kinetic studies revealed that ascorbic acid increased proline hydroxylation in the intracellular procollagen pool within 1 h and stimulated the cleavage of type I collagen propeptides beginning at 2.5 h. Mature alpha 1(I) and alpha 2(I) collagen components were first detected at 10 h and continued to increase in both cell layer and culture medium for up to 72 h. Ascorbic acid also increased the rate of procollagen secretion from cell layers to culture medium. The secretion of another matrix protein, fibronectin, was only slightly affected. Alkaline phosphatase or its mRNA was first detected 2-3 days after ascorbic acid addition, but osteocalcin mRNA was not seen until day 6. Two inhibitors of collagen triple-helix formation, ethyl-3,4-dihydroxybenzoate and 3,4-dehydroproline, inhibited procollagen hydroxylation and alkaline phosphatase induction. 3,4-Dehydroproline also inhibited the induction of alkaline phosphatase and osteocalcin mRNAs. Surprisingly, induction was not blocked if cells were exposed to ascorbic acid before inhibitor addition. Alkaline phosphatase was also partially inhibited if cells were grown in the presence of purified bacterial collagenase. These results indicate that the induction of osteoblast markers by ascorbic acid does not require the continuous hydroxylation and processing of procollagens and suggest that a stable, possibly matrix-associated signal is generated at early times after ascorbic acid addition that allows subsequent induction of osteoblast-related genes.
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PMID:Effects of ascorbic acid on collagen matrix formation and osteoblast differentiation in murine MC3T3-E1 cells. 807 60

Chicken liver is lack of ascorbic acid biosynthesis system, different from mammals and highly evoluted birds. Chicken hepatocytes cultured without ascorbate was expected to have lower ascorbate amounts than physiological levels. Intracellular was decreased as compared with intact liver by cell preparation performed with in situ collagenase perfusion. We added ascorbate to a primary culture of chicken hepatocytes in order to restore the amount of ascorbate. Serum-free Leivobitz's L-15 medium which do not contain ascorbate was used for control medium. Cells were cultured with several concentrations of ascorbate for 24 or 48 h. After ascorbate supplementation for 24 to 48 h, cellular ascorbate concentration increased depending on the dose of medium ascorbate. Medium lactate dehydrogenase activity derived from hepatocytes, an index of cell injury, decreased upon 5-100 mg/l of ascorbate supplementation for 48 h. Tyrosine aminotransferase activity, an index of liver function, increased following culture with 50 and 100 mg/l ascorbate for 48 h. The activities, however, decreased by supplementation with 1000 mg/l of ascorbate. In conclusion hepatocytes lost intracellular ascorbate during preparation by in situ collagenase perfusion. Supplementation of ascorbate restored cellular ascorbate concentration, lowered cell injury and raised tyrosine aminotransferase activitv in primary cultured chicken hepatocytes. Ascorbate treatment for 48 h at 50 mg/l was the best combination in this study for primary culture of chicken hepatpcyte with non-serum L-15 medium
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PMID:Ascorbic acid supplementation to primary culture of chicken hepatocytes with non-serum medium. 1108 76

Ascorbic acid (AA) enhances osteoblastic differentiation by increasing collagen accumulation, which in turn, results in increased alkaline phosphatase (AP) expression in some osteogenic cells. However, in other cells, including human periodontal ligament (PDL) cells, additional osteoinductive agents are required for this response. To understand the potential basis for the maintenance of the AP phenotype of PDL cells exposed to AA, we examined the modulation of the tissue-degrading matrix metalloproteinases (MMPs) and their inhibitors by AA in short-term cell cultures. Early passage PDL cells in serum-free medium were exposed to AA for 5 days. The samples were analyzed for MMPs and their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs), AP, collagen I(alpha1), and osteocalcin. We found that AA dose-dependently increased the expression of collagenase-1, and minimally TIMP-1, but not stromelysin-1 or TIMP-2. Additionally, AA caused substantial increases in levels of type I collagen. AA was unable to increase AP activity or osteocalcin messenger RNA in PDL cells. However, the cells retained the ability to show a significantly greater AP expression in high- versus low-density cultures, and increased osteocalcin as well as AP levels when cultured in the presence of dexamethasone. Moreover, in cells exposed to dexamethasone, increases in AP and osteocalcin were accompanied by a repression of collagenase-1 expression. In contrast to PDL cells, AA did not induce collagenase but produced a significant increase in AP expression in MC3T3-E1 cells. These findings provide the first evidence that AA, by modulating both collagen and collagenase-1 expression in PDL cells, most likely contributes to a net matrix remodeling response in these cells. Furthermore, the relationship between changes in collagenase expression and alterations in AP activity in PDL and MC3T3-E1 cells suggests a potential role for collagenase in modulating the AP phenotype of cells with osteoblastic potential.
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PMID:Ascorbic acid induces collagenase-1 in human periodontal ligament cells but not in MC3T3-E1 osteoblast-like cells: potential association between collagenase expression and changes in alkaline phosphatase phenotype. 1251 Aug 7

Cancer is associated with increased cell growth, and expression of matrix metalloproteinases (MMPs) and transforming growth factor-beta (TGF-beta). The dose-dependent effects of ascorbate (Vitamin C) on cancer cell growth, and expression of MMPs and TGF-beta were examined. Renal-adenocarcinoma, melanoma and mammary cancer cells were dosed with 0-100mM ascorbate and examined for cell survival or proliferation, and expression of MMP-1, MMP-2 and TGF-beta at protein and/or mRNA levels. The lower concentrations of ascorbate significantly inhibited cancer cell viability while stimulating MMPs and TGF-beta expression, indicating elimination of cancer cells with damage to the extracellular matrix (ECM). Conversely, ascorbate at higher concentrations dramatically stimulated cell proliferation and inhibited MMPs and TGF-beta expression, implicating growth and ECM advantage.
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PMID:Reciprocal effects of ascorbate on cancer cell growth and the expression of matrix metalloproteinases and transforming growth factor-beta. 1760 32

Ascorbate has dose-dependent inverse effects on cancer cells growth and expression of matrixmetalloproteinases (MMP) and transforming growth factor-beta (TGF-beta), which regulate extracellular matrix (ECM) remodeling. We examined melanoma cell viability and ECM remodeling mechanisms of ascorbate and its modulation by an extract from Polypodium leucotomos (PL) (a fern) via the regulation of apoptosis, heat-shock proteins (HSPs), MMP-1 or tissue inhibitors of matrix metalloproteinase-1 (TIMP-1) that inhibits MMP-1. The dose-dependent regulation of cell viability/proliferation by ascorbate was associated with inverse regulation of apoptosis and stimulation of HSPs at growth-inhibitory concentrations. PL antagonized the stimulation of MMP-1, TGF-beta and HSPs by a growth-inhibitory ascorbate dose and stimulated the expression of TIMP-1, while maintaining growth inhibition. We infer that a combination of ascorbate with PL is beneficial to cancer management via the simultaneous inhibition of cell growth and expression of MMP-1, TGF-beta and HSPs, and furthermore, the stimulation of TIMP-1.
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PMID:Cancer cell growth and extracellular matrix remodeling mechanism of ascorbate; beneficial modulation by P. leucotomos. 1966 40


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