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)

Pump-1 cDNA has recently been isolated by screening a human tumor cDNA library with a transin (rat stromelysin) probe under low-stringency hybridization conditions. The cDNA codes for a potential protein with significant sequence similarity to the metalloproteinases collagenase and stromelysin, but which lacks the hemopexin-like domain characteristic of these enzymes. Expression of pump-1 cDNA in cos cells using an expression vector leads to secretion of a protein of Mr 28,000 with latent, organomercurial-activatable proteinase activity. Cos cells transfected with a partial pump-1 cDNA in the vector pPROTA secrete a fusion protein between the IgG-binding domains of staphylococcal protein A and pump-1. The fusion protein binds to IgG-Sepharose, and the bound fusion protein undergoes apparent autocleavage in the presence of 4-aminophenylmercuric acetate with elution of active pump-1 species of Mr 21,000 and 19,000. Active pump-1 degrades casein, gelatins of types I, III, IV, and V, and fibronectin and can activate collagenase. Active pump-1 is inhibited by EDTA, 1,10-phenanthroline, and the tissue inhibitor of metalloproteinases. These results show that, despite the absence of a hemopexin-like domain, pump-1 is a latent secreted metalloproteinase. Postpartum rat uteri contain elevated levels of rat pump-1 mRNA. On the basis of this observation, its size, and its substrate specificity, we suggest that pump-1 might correspond to a previously described uterine metalloproteinase, matrix metalloproteinase 7.
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PMID:Pump-1 cDNA codes for a protein with characteristics similar to those of classical collagenase family members. 255 50

The regeneration of connective tissue attachment is a major goal of clinical periodontics. Recent investigations on biochemically mediated periodontal regeneration have attempted to define the various biological response modifiers which may provide a mechanism for periodontal regeneration. Fibronectin and endothelial cell growth factor have been shown to selectively enhance periodontal ligament (PDL) cell adhesion, migration, and proliferation. In addition, dentin preconditioned with tetracycline HCl (TTC) or citric acid (CA) supports PDL cell adhesion, presumably by exposing collagen fibers. We have now extended these studies to include basic fibroblast growth factor (b-FGF) as a potential meditor of periodontal regeneration. Using AFSCM (assays for specific cell migration), b-FGF in concentrations as low as 10 ng per dentin block significantly stimulated PDL cell chemotaxis, while the antibody against b-FGF inhibited both the chemotactic and proliferative characteristics of the mitogen. We also found that 5 ng and above of b-FGF per dentin block significantly stimulated human endothelial cell migration and proliferation. Using 125I-b-FGF, we demonstrated that the factor binds to native dentin. This binding was increased when the dentin blocks were preconditioned by TTC or CA and reduced when the dentin was subsequently treated with collagenase. 125I-b-FGF also bound with moderate affinity to a type I collagen affinity column whereas the binding to a hydroxylapatite affinity column was negligible. The combination of FN and b-FGF was a marginally more potent chemo-attractant than b-FGF alone for PDL cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Repopulation of dentin surfaces by periodontal ligament cells and endothelial cells. Effect of basic fibroblast growth factor. 255 Jun 5

We have investigated the mechanism of inhibition of the serum-free monolayer growth of normal rat kidney (NRK) cells by transforming growth factor-beta (TGF-beta). NRK cells grown on fibronectin-coated dishes exhibited a biphasic response to TGF-beta. Monolayer growth was slightly stimulated by subpicomolar concentrations, while picomolar concentrations of TGF-beta inhibited NRK cell growth in the presence or absence of epidermal growth factor. NRK cells exhibited a similar biphasic growth response to exogenous type I collagen. TGF-beta induced a 3-5-fold increase in the deposition of type I collagen-like proteins into the extracellular matrix of NRK cells during serum-free growth. Type I collagen-like proteins were identified by their sensitivity to degradation by purified bacterial collagenase and by Western blot analysis. The TGF-beta dose-response curves for induction of extracellular matrix-localized collagen and inhibition of NRK cell growth were similar. Finally, the inclusion of a purified bacterial collagenase, which did not degrade TGF-beta or TGF-beta receptors, or alter control NRK growth, prevented exogenous collagen or TGF-beta from inhibiting the serum-free growth of NRK cells. Our results demonstrate that an increase in collagen secretion plays an important role in the inhibition of the growth of NRK cells by TGF-beta.
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PMID:Inhibition of normal rat kidney cell growth by transforming growth factor-beta is mediated by collagen. 255

Two intralesional injections of interferon alfa-2b (1.5 million U per injection) into a progressively enlarging keloid resulted in a 41% reduction in its area. Fibroblasts cultured from the keloid before and 9 days after the initial injection were compared with fibroblasts cultured from the patient's normal skin with respect to proliferation and production of connective tissue matrix components and collagenase. There were no significant differences in the in vitro doubling times of keloidal fibroblasts before (p greater than 0.2) and after (p greater than 0.5) treatment with interferon alfa-2b, as well as of normal fibroblasts, in subconfluent cultures. Multiple passages of keloidal fibroblasts before interferon alfa-2b therapy assayed as confluent cultures produced more collagen (171%, 187%, and 204%), more glycosaminoglycans (153% and 141%), and less collagenase (26% and 31%) than the patient's own normal fibroblasts. In contrast, keloidal fibroblasts after interferon alfa-2b therapy persistently produced normal or subnormal amounts of collagen (107%, 73%, and 64%) and glycosaminoglycans (97% and 96%) and normalized levels of collagenase activity (96% and 86%). Normal amounts of fibronectin were produced by keloidal fibroblasts before and after treatment with interferon alfa-2b.
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PMID:Short-term keloid treatment in vivo with human interferon alfa-2b results in a selective and persistent normalization of keloidal fibroblast collagen, glycosaminoglycan, and collagenase production in vitro. 255 84

Endothelial cells were obtained from the aortas of Wistar rats by collagenase digestion. Cells were grown to confluence in medium 199 enriched with L-glutamine but without specific growth factors. Cells were subcultured into 35 mm dishes or 25 cm2 flasks coated with fibronectin. For cell growth studies, cells were seeded onto multiwell plates or 35 mm dishes. In two experiments the cells were grown in an hypoxic atmosphere of 5.3% O2 and in a third the level of oxygen was 2.5%. Control cultures for each experiment were grown in 5% CO2 and air. Cell populations were counted at 2-day intervals and at the termination of each experiment the cells were fixed and the area of each plate or flask occupied by sprouting cells was assessed by point counting. Endothelial cells grown in 5.3% O2 grew more rapidly and attained confluence earlier than in the controls. An atmosphere of 2.5% O2 did not accelerate growth but neither did it inhibit it, so after 9 days there were as many hypoxic cells as there were controls. Hypoxia also stimulated sprouting activity to occur earlier and to become much more extensive than in control cultures. Under the influence of hypoxia, sprouting consisted of complex anastomotic or arborizing patterns forming syncytium-like masses beneath the monolayer of oval cells. This process appeared to originate from foci of altered endothelial cells that had become retracted, smaller, elongated and migratory, and which displayed increased immunoreactivity for factor VIII antigen. It was concluded that a level of hypoxia, similar to that in systemic veins, stimulates growth of arterial endothelium and provokes enhanced sprouting activity.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effect of hypoxia upon the growth and sprouting activity of cultured aortic endothelium from the rat. 259 52

A high molecular weight extracellular protein has been purified from cell culture medium of Ewing's sarcoma cell lines, by high performance liquid chromatography and electroelution from SDS-PAGE electrophoresis. This protein has an apparent molecular mass of about 500,000 Da on SDS-PAGE. Immunoprecipitation studies with several extracellular matrix glycoproteins (laminin, fibronectin) specific antisera indicate it is a separate protein. Reduction of disulphide bonds with 2-ME or DTT fails to significantly alter its migration on SDS-PAGE gels, other than a slight apparent increase in molecular mass, indicating an apparent single polypeptide chain structure. The slightly greater mobility observed in unreduced gels suggests one or more regions of intrachain disulfide bonding. It is sensitive to pepsin and trypsin, but resistant to bacterial collagenase indicating that it does not contain collagenous domains. Metabolic labelling with 3H-proline, 3H-leucine, and 35S-methionine indicate that this protein is proline-poor, but leucine, and especially methionine, rich. Sodium 35S-sulfate incorporation is totally negative and treatment with glycosaminoglycan degrading enzymes has no effect on the mobility of the protein on gels, unlike typical proteoglycans. This protein appears by rotary shadowing electron microscopy as a long, thin, filamentous molecule at least 500 nm (0.5 um) in length. The tissue localization and function are unknown at this time, but are under active investigation.
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PMID:A novel 500,000 Da, linear, single chain extracellular protein synthesized by several childhood tumors. 263 60

Parenchymal organoidal structures that were obtained from collagenase digestion of reduction mammoplasty specimens of apparently normal human breasts have been grown in short-term primary cultures, either on plastic or on floating gels of polymerized rat-tail collagen. Three morphologically distinct major cell types are readily observed in both systems: cuboidal cells, which occupy apical positions on collagen gels; larger, epithelioid, or basal cells on gels; and elongated cells which penetrate into the gel. In addition, a fourth cell type, that of large, flat cell, is observed less readily by phase contrast microscopy on the surface of cultures grown on plastic. Immunofluorescent and immunocytochemical staining of cultures on plastic or histologic sections of cultures on gels have been undertaken with antisera and other histochemical reagents that stain the different parenchymal cell types in vivo. Thus antisera to epithelial membrane antigen(s), monoclonal antibodies (MABs) to the defatted mammary milk fat globule membrane, peanut lectin, and keratin MAB LE61, which preferentially stain the epithelial cells of ducts in vivo, also stain the cuboidal/apical cells in vitro. The large, flat cells are stained intensely by the first three reagents but not by the last one. Antisera to collagen IV, laminin, fibronectin, actin, keratin MAB LP34, MABs to the common acute lymphoblastic leukemia antigen, and MAB LICR-LON-23.10, which showed enhanced staining for the ductal myoepithelial cells in vivo, also stain the epithelioid/elongated cells in vitro. However, the effect of the last four reagents is reduced considerably in most elongated cells, and MAB LP34 stains the large, flat cells intensely. Heterogeneous cells of intermediate morphologies and staining patterns between the cuboidal cells and large, flat cells are related to mammary epithelial cells. whereas the large epithelioid/elongated cells have some characteristics of myoepithelial cells, and that intermediate forms may exist in culture between the two parenchymal cell types.
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PMID:Characterization of human mammary cell types in primary culture: immunofluorescent and immunocytochemical indicators of cellular heterogeneity. 264 83

Pz-peptidase was purified from rabbit muscle by acid precipitation of tissue homogenate followed by cation- and anion-exchange chromatography, gel chromatography, and immunoadsorption. In analytical gel chromatography, one single peak of protein with corresponding Pz-peptidase activity was obtained. The enzyme had an apparent Mr of 74,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and was eluted at pH 4.8 in chromatofocusing. No metals were detectable in the protein by neutron activation analysis. Purified Pz-peptidase hydrolyzed Dnp-Pro-Leu-Gly-Pro-Trp-D-Lys (Km 7.2 microM) most effectively in the presence of 5 mM 2-mercaptoethanol and 10 mM CaCl2. No inhibition was observed with inhibitors of serine proteinases, aspartic proteinases, or metalloproteinases, apart from some nonspecific reversible inhibition by 1,10-phenanthroline. The activation by Ca2+ was reversed by EDTA. The enzyme was not inhibited by E-64, cystatin, or leupeptin, but was irreversibly inactivated by iodoacetate, iodoacetamide, and N-ethylmaleimide. It was therefore concluded that rabbit muscle Pz-peptidase is not a typical member of any of the four recognized catalytic classes of proteinases, but may be an atypical cysteine endopeptidase. The peptidase was not bound by alpha 2-macroglobulin. No hydrolysis of gelatin or fibronectin by the enzyme was detected, nor was there any activation of latent collagenase.
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PMID:Purification and characterization of Pz-peptidase from rabbit muscle. 267 41

A procedure for extraction and quantification of fibronectin in human aortic tissue is described in this paper. Dried, defatted samples of human aortic tissue were subjected to sequential extraction with (i) 0.89% NaCl, 10 mmol/l Tris/HCl, pH 7.4, (ii) 5 mg/ml heparin, 2 mol/l urea and (iii) collagenase digestion. More than 75% of hexosamine-containing molecules were solubilized by this procedure. Immunoblotting of extracted proteins separated by SDS-PAGE showed that extracted fibronectin had a mobility in the same range as that of plasma fibronectin. Fibronectin ELISA performed on these extracts gave dilution curves parallel to the standard curve, the sensitivity was 2.7 micrograms/l. Recoveries of a fibronectin standard added to the NaCl, heparin/urea and collagenase solutions during extraction were 97%, 90% and 84% respectively. Normal aortic tissue from 31 patients was subjected to the sequential extraction scheme and fibronectin quantification in the various extracts demonstrated that 4.52 +/- 1.79 micrograms was dissolved in the NaCl extracts, 5.41 +/- 2.28 in the heparin/urea extract and 1.08 +/- 0.43 in the collagenase digest, respectively. (Values are expressed as micrograms fibronectin/10 mg dry, defatted tissue (mean +/- SD]. Our results indicate that the ELISA method can be applied for the measurement of fibronectin in extracts of human aortic tissue. This might be useful in the study of diseases where alterations in arterial fibronectin content may be expected.
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PMID:Quantification of fibronectin in extracts of human aortae by an ELISA. 274 Aug 16

To obtain data concerning the pathology of diabetic arteries, aortas from 23 patients with diabetes mellitus [9 with insulin-dependent diabetes mellitus (IDDM) and 14 with non-insulin-dependent diabetes mellitus (NIDDM)] were collected at autopsy together with aortas from sex- and age-matched nondiabetic persons. A histomorphometric study was performed blindly on antifibronectin PAP-stained sections to determine the distribution of fibronectin-containing space in the vessels. In both IDDM and nonIDDM groups a statistically significant increase of approximately 45% was seen in the amount of stainable material in the tunica media. The increase was not influenced by the presence or absence of overlying plaque. No differences were seen between diabetic and nondiabetic vessels in the tunica intima. The content of extractable fibronectin in intima-media preparations was measured. The samples were extracted sequentially with buffered saline, a heparin-urea solution, and finally collagenase digestion. Fibronectin measured in these extracts showed that statistically significantly more of this glycoprotein was found in vessels from diabetic persons compared with nondiabetic persons, when comparing areas of the vessels without macroscopical visible plaque. However, only among IDDM patients increased amounts were apparent in plaque areas. These results indicate that diabetic patients develop structural alterations in the connective tissue of their arteries, consistent with a hypothesis of a diabetic macroangiopathy.
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PMID:Accumulation of fibronectin in aortas from diabetic patients. A quantitative immunohistochemical and biochemical study. 279 91


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