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)

Keloids are characterized by an overabundant deposition of collagen, and they recur frequently following excision. Fibroblasts isolated from keloid tissue and maintained in cell culture continue to express an increased capacity to produce collagen. In an effort to define the mechanisms responsible for keloid formation, the potential of exogenous transforming growth factor beta 1 (TGF-beta 1) to differentially affect DNA synthesis and collagen expression in cultured human fibroblasts derived from keloid or normal dermis was investigated. In this study, TGF-beta 1 at a concentration of 5.0 ng/ml was found to stimulate DNA synthesis of keloid-derived fibroblasts to a greater extent than fibroblasts derived from normal dermis. With a microassay to measure levels of collagenase-digestible radiolabeled proteins, TGF-beta 1 was found to elicit a greater increase in absolute collagen synthesis in keloid-derived fibroblasts compared with fibroblasts derived from normal dermis. Examination of tRNA(pro) pool-specific activities indicated that these observed differences in rates of collagen synthesis were not the result of unequal rates of proline transport or pool size. Likewise, TGF-beta 1 did not alter the uptake of vitamin C, an essential cofactor and mediator needed for maximal collagen expression. The increase in collagen synthesis by keloid-derived fibroblasts treated with TGF-beta 1 was accompanied by a corresponding increase in procollagen type I mRNA levels, indicating that the differential response of keloid and normal dermal fibroblasts to this growth factor is occurring primarily at a pretranslational level. These results suggest a unique sensitivity of keloid fibroblasts to TGF-beta 1 and thus a possible role for this mediator in keloid pathogenesis.
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PMID:The effect of TGF-beta on keloid fibroblast proliferation and collagen synthesis. 882 22

Type VI collagen is present in most connective tissues, where it is considered to play a crucial role in the attachment of cells to the extracellular matrix and/or in the three-dimensional organization of the collagen meshwork. Although some information is available on its formation, the mechanisms involved in its degradation are not understood. Here, we present evidence for lysosomal digestion of type VI collagen by fibroblasts of periosteal explants. In the lysosomal apparatus of these cells, broad-banded filamentous aggregates characterized by 100-nm periodicity were found, which proved to consist of type VI collagen as indicated by their stainability with anti-type VI collagen antibodies. By interfering with synthesis (ascorbate or alpha, alpha-dipyridyl), intracellular translocation of collagen-containing vesicles (colchicine) as well as phagocytosis (cytochalasin B), it was shown that the intracellular broad-banded type VI collagen represented phagocytosed material. In the presence of acidotropic agents (NH4Cl and methylamine) the amount of intracellular type VI collagen increased significantly (5- to 10-fold), suggesting that a rise of pH in the endosomal/lysosomal apparatus causes inhibition of its degradation. By using a variety of proteinase inhibitors, it was found that inhibition of collagenase (when used in combination with NH4Cl), or inhibition of cysteine proteinases (both with and without NH4Cl), resulted in an increased amount of intracellular type VI collagen, whereas inhibition of serine proteinases significantly lowered the level of intracellular type VI collagen. The data presented are the first to indicate a pathway by which type VI collagen degradation may occur: fibroblasts phagocytose type VI collagen and subsequently digest this collagen in their lysosomal apparatus. Degradation depends on the activity of several enzymes, among them collagenase and serine proteinases, probably exerting their activity in the extracellular space just before the actual internalization. After uptake, digestion involves pH-sensitive lysosomal enzymes, including those belonging to the class of cysteine proteinases.
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PMID:Type VI collagen is phagocytosed by fibroblasts and digested in the lysosomal apparatus: involvement of collagenase, serine proteinases and lysosomal enzymes. 905 16

Ultrasound has been applied therapeutically to accelerate connective tissue healing although there is little direct scientific evidence to support its use. This investigation was conducted to determine the effects of ultrasound on the rate of collagen synthesis and cell proliferation using cultured fibroblasts derived from Achilles tendons of neonatal rats. Ultrasound (intensity = 0.4 W.cm-2; frequency = 1 MHz) was applied to experimental cells growing as monolayers in culture flasks. Ultrasound had no effect on the rate of collagen synthesis by control fibroblasts over a period of 9 d. The addition of vitamin C to culture media stimulated collagen synthesis to the same extent in both control and ultrasound-treated cultures. Partial digestion of cell matrices with collagenase (used to simulate injury) resulted in an approximately 20% increase in the rate of collagen synthesis. Synthesis was further increased with ultrasound treatment (50-67%). For example, after a single ultrasound treatment, the rate of collagen synthesis was 3.0 +/- 0.4 pg.micrograms-1 DNA.h-1 in cultures treated with collagenase, compared with 1.8 +/- 0.3 pg.micrograms-1 DNA.h-1 in collagenase-treated cultures not treated with ultrasound and 1.4 +/- 0.3 pg.micrograms-1 DNA.h-1 in controls. Ultrasound applied to preconfluent cultures resulted in significant increases in the rate of thymidine incorporation and DNA content. Three daily ultrasound treatments caused a 100% increase in the rate of thymidine incorporation and a 28% increase in DNA content. The results indicate that ultrasound stimulates collagen synthesis in tendon fibroblasts in response to an injury of the connective tissue matrix and that ultrasound stimulates cell division during periods of rapid cell proliferation.
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PMID:The effect of ultrasound on collagen synthesis and fibroblast proliferation in vitro. 913 71

Human embryonic kidney cells (293-EBNA) have been transfected with the full-length human alpha1 chain of collagen V using an episomal vector. High yields (15 microgram/ml) of recombinant collagen were secreted in the culture medium. In presence of ascorbate, the alpha1(V) collagen is correctly folded into a stable triple helix as shown by electron microscopy and pepsin resistance. Circular dichroism data confirm the triple-helix conformation and indicate a melting temperature of 37.5 degrees C for the recombinant homotrimer. The major secreted form is a 250-kDa polypeptide (alpha1FL). N-terminal sequencing and collagenase digestion indicate that alpha1FL retains the complete N-propeptide but lacks the C-propeptide. However, alpha1FL might undergo a further N-terminal trimming into a form (alpha1TH) corresponding to the main triple-helix domain plus the major part of the NC2 domain. This processing is different from the one of the heterotrimeric (alpha1(V))2alpha2(V) and could have some physiological relevance. Analysis of cell homogenates indicates the presence of a 280-kDa polypeptide that is disulfide-linked through its C-terminal globular domain. This C-propeptide is rapidly cleaved after secretion in the medium, giving the first evidence of a C-terminal processing of recombinant fibrillar collagens. Rotary shadowing observations not only confirm the presence of a globular domain at the N-terminal end of the molecule but reveal the presence of a kink within the triple helix in a region poor in iminoacids. This region could represent a target for proteases. Together with the thermal stability data, these results might explain the low amount of (alpha1(V))3 recovered from tissues.
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PMID:Human recombinant alpha1(V) collagen chain. Homotrimeric assembly and subsequent processing. 937 85

Recent studies indicate that vitamin D metabolites exert rapid effects on growth plate chondrocytes via changes in PG production and protein kinase C (PKC) activity. This suggests that these two products of vitamin D action may be interrelated. To test this hypothesis, we examined the effect of PGE2 on rat costochondral resting zone and growth zone cartilage cells and determined whether the effects of PGE2 are mediated by changes in the level of cAMP and/or PKC activity, whether there is a relationship between cAMP production and PKC activity, and whether cell maturation-specific effects are involved. Confluent, fourth passage resting zone and growth zone cartilage cell cultures were incubated in DMEM containing 10% FBS, 50 microg/ml vitamin C, and 1% antibiotics. The PGE2 concentration was varied from 0.007-15 ng/ml. Low concentrations of PGE2 caused a dose-dependent increase in cell number and [3H]thymidine incorporation and stimulated alkaline phosphatase specific activity. These effects were comparable in resting zone and growth zone cartilage cells at the same PGE2 concentrations. At higher concentrations, PGE2 caused a general increase in the synthesis of collagenase-digestible protein and noncollagenase-digestible protein in resting zone cartilage cells and of collagenase-digestible protein in growth zone cartilage cells, resulting in a net increase in the percent collagen synthesis for both cell types. cAMP production was increased over the entire range of chondrocyte response. Prevention of cAMP metabolism with the protein kinase A inhibitors H-8 and H-89 blocked the PGE2-dependent inhibition of PKC in resting zone cartilage cells in a dose-dependent manner. H-8 alone had no effect on PKC in resting zone cartilage cells, but stimulated PKC activity in growth zone cartilage cells; H-89 alone stimulated PKC activity in resting zone cartilage cells. These results suggest that low levels of PGE2 promote differentiation, whereas high doses promote an anabolic response; PGE2 increases cAMP production and PKC activity in a cell maturation-dependent manner; PGE2 exerts its effects via cAMP production and PKC activity; and regulation of PGE2-dependent PKC is via cAMP.
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PMID:The effect of prostaglandin E2 on costochondral chondrocyte differentiation is mediated by cyclic adenosine 3',5'-monophosphate and protein kinase C. 952 68

Cultured rat Schwann cells were stimulated to deposit fibrillar extracellular matrix by treatment with ascorbic acid in the absence of nerve cells. Immunofluoresence staining of the matrix showed that it contains collagens types I and IV, fibronectin and perlecan but not laminin. Collagen type IV, fibronectin and perlecan co-distributed completely in the matrix fibrils, whereas collagen type I was present in only a subset of these fibrils. Time course studies indicated that collagen type I fibrils appear at late stages of matrix formation. Digestion of Schwann cell extracellular matrix with collagenase effectively disrupted most of the matrix including fibronectin fibrils. This was in contrast with fibroblasts, where collagenase treatment removed collagen with no visible effect on fibronectin fibrils. alpha5 integrin was expressed on the cell surface of Schwann cells and partially codistributed with fibronectin-containing fibrils. This suggests that the inability of Schwann cells to deposit fibronectin-containing matrix through a conventional, collagen-independent mechanism was not due to the lack of fibronectin-binding integrins on their cell surface. Polyclonal anti-fibronectin antibodies inhibited the deposition of fibronectin into the matrix fibrils, whereas collagen type IV fibrils were generally unaffected. Growth of Schwann cells on collagen type IV-coated substrate in the absence of ascorbate induced deposition of fine fibronectin fibrils. These results suggest that Schwann cells use an apparently novel, collagen type IV-dependent mechanism for the deposition of fibronectin into their extracellular matrix.
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PMID:Schwann cells use a novel collagen-dependent mechanism for fibronectin fibril assembly. 971 69

During development, mRNA for matrix metalloproteinase-13 (MMP-13) is found associated with cartilage undergoing hypertrophy, suggesting that this collagenase plays a role in cell enlargement and/or cartilage calcification. Using chondrocytes from prehypertrophic cartilage of chick embryo sternae, we have examined the relationship between MMP-13 expression and the transition to hypertrophy. When hypertrophy was induced by serum-free culture with ascorbate and bone morphogenetic protein-2 (BMP-2), MMP-13 mRNA levels paralleled those for type X collagen. Chondrocytes from the caudal, nonhypertrophying portion of chick sternae expressed neither type X collagen nor MMP-13, confirming that MMP-13 mRNA is a marker for hypertrophy. Zymography with conditioned medium yielded a proteinase band at 59 kDa, which was absent in nonhypertrophic chondrocytes. A polyclonal antibody raised against chick MMP-13 reacted with the 59-kDa protein, confirming that it is MMP-13. Although mRNA for MMP-13 peaked at days 4-5 of culture, only low levels of MMP-13 activity were present, and the activity increased gradually in parallel with later increases in MMP-2. These results suggest that MMP-13 is activated by MMP-2 during chondrocyte maturation, and that the combination of both proteinases is required to prepare cartilage matrix for subsequent calcification, before endochondral ossification.
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PMID:MMP-13 is induced during chondrocyte hypertrophy. 1077 23

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

The aim of the study was to determine whether collagenase inhibitors reduce corneal haze after photorefractive keratectomy (PRK). Inhibition of the initial removal phase of healing may limit the subsequent repair and replacement phases responsible for haze and regression. Thirty rabbits received -6.00D 5 mm right PRK. They were randomized to five treatment groups: G. cysteine, G. ethylene diamine tetra-acetic acid (EDTA), G. ascorbate, Oc. tetracycline or no drops. Dichlorotriazinyl aminofluorescein (DTAF) was applied to the wound immediately after surgery in two rabbits of each group, to delineate newly-synthesized from original tissue. Corneal haze was assessed by a video-linked frame grabber with computerized grey scale analysis. Corneas were taken for histology at 1 or 3 months post-operatively. Corneal haze was not significantly different between the treatment groups and controls. The severity of the histological changes varied between individuals. Within the ablation zone the epithelium was on average 10% thicker (3--4 micro m) than outside, and in some rabbits there were irregularities of the epithelial--stromal junction. The new subepithelial tissue had a mean depth of 7.8 micro m, and the superficial stroma was disorganized to a mean depth of 49 micro m. No particular treatment demonstrated significant benefits over controls; but of the treatments used, cysteine tended to produce the best results. Eyes treated with EDTA fared worst in most respects. The collagenase inhibitors used did not improve the outcome of PRK in rabbits. It remains to be determined whether firstly, the new more potent agents would have an effect, and secondly, whether collagenase inhibitors are of benefit in humans.
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PMID:Effect of collagenase inhibitors on corneal haze after PRK. 1118 Sep 74

The photoprotective potential of the dietary antioxidants vitamin C, vitamin E, lycopene, beta-carotene, and the rosemary polyphenol, carnosic acid, was tested in human dermal fibroblasts exposed to ultraviolet-A (UVA) light. The carotenoids were prepared in special nanoparticle formulations together with vitamin C and/or vitamin E. Nanoparticle formulations, in contrast to dimethylsulphoxide, stablized lycopene in the cell culture medium and allowed efficient cellular uptake. The presence of vitamin E in the formulation further increased the stability and cellular uptake of lycopene. UVA irradiation of the human skin fibroblasts led to a 10-15-fold rise in metalloproteinase 1 (MMP-1) mRNA. This rise was suppressed in the presence of low microM concentrations of vitamin E, vitamin C, or carnosic acid but not with beta-carotene or lycopene. Indeed, in the presence of 0.5-1.0 microM beta-carotene or lycopene, the UVA-induced MMP-1 mRNA was further increased by 1.5-2-fold. This increase was totally suppressed when vitamin E was included in the nanoparticle formulation. Heme-oxygenase 1 (HO-1) mRNA expression was strongly induced by UVA irradiation but none of the antioxidants inhibited this effect at the concentrations used in this study. Indeed, beta-carotene or lycopene (0.5-1.0 microM) led to a further 1.5-fold rise in the UVA-induced HO-1 mRNA levels. In conclusion, vitamin C, vitamin E, and carnosic acid showed photoprotective potential. Lycopene and beta-carotene did not protect on their own but in the presence of vitamin E, their stability in culture was improved and the rise in MMP-1 mRNA expression was suppressed, suggesting a requirement for antioxidant protection of the carotenoids against formation of oxidative derivatives that can influence the cellular and molecular responses.
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PMID:Photoprotective potential of lycopene, beta-carotene, vitamin E, vitamin C and carnosic acid in UVA-irradiated human skin fibroblasts. 1205 67


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