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)

Different procedures of enzymatic digestion of rat prostatic tissue and unique sets of mitogenic factors made it possible to culture practically pure populations of epithelial and stromal cells without previous separation of the two types of cells. Keratin-positive epithelial cells dissociated by trypsin and collagenase from adult rat ventral prostate proliferated in medium WAJC 404 supplemented with epidermal growth factor, insulin, cholera toxin, and bovine pituitary extract. Proliferation of epithelial cells was completely inhibited by dexamethasone as low as 30 nM. On the other hand, fibroblast-like stromal cells released by trypsin digestion required a plastic substratum coated with calf serum or fibronectin, and proliferated in Eagle's minimum essential medium supplemented with cholera toxin, bovine pituitary extract, dexamethasone, and bovine serum albumin. Epidermal growth factor and insulin had negligible effect on proliferation of stromal cells. Physiological concentrations of dihydrotestosterone and estradiol showed no effect on proliferation of both types of cells.
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PMID:Differences in growth requirements between epithelial and stromal cells derived from rat ventral prostate in serum-free primary culture. 223 29

Human rheumatoid synovial cells in culture secrete at least three related metalloproteinases that digest extracellular matrix macromolecules. One of them, termed matrix metalloproteinase 2 (MMP-2), has been purified as an inactive zymogen (proMMP-2). The final product is homogeneous on SDS/PAGE with Mr = 72,000 under reducing conditions. The NH2-terminal sequence of proMMP-2 is Ala-Pro-Ser-Pro-Ile-Ile-Lys-Phe-Pro-Gly-Asp-Val-Ala-Pro-Lys-Thr, which is identical to that of the so-called '72-kDa type IV collagenase/gelatinase'. The zymogen can be rapidly activated by 4-aminophenylmercuric acetate to an active form of MMP-2 with Mr = 67,000, and the new NH2-terminal generated is Tyr-Asn-Phe-Phe-Pro-Arg-Lys-Pro-Lys-Trp-Asp-Lys-Asn-Gln-Ile. However, following 4-aminophenylmercuric acetate activation, MMP-2 is gradually inactivated by autolysis. Nine endopeptidases (trypsin, chymotrypsin, plasmin, plasma kallikrein, thrombin, neutrophil elastase, cathepsin G, matrix metalloproteinase 3, and thermolysin) were tested for their abilities to activate proMMP-2, but none had this ability. This contrasts with the proteolytic activation of proMMP-1 (procollagenase) and proMMP-3 (prostromelysin). The optimal activity of MMP-2 against azocoll is around pH 8.5, but about 50% of activity is retained at pH 6.5. Enzymic activity is inhibited by EDTA, 1,10-phenanthroline or tissue inhibitor of metalloproteinases, but not by inhibitors of serine, cysteine or aspartic proteinases. MMP-2 digests gelatin, fibronectin, laminin, and collagen type V, and to a lesser extent type IV collagen, cartilage proteoglycan and elastin. Comparative studies on digestion of collagen types IV and V by MMP-2 and MMP-3 (stromelysin) indicate that MMP-3 degrades type IV collagen more readily than MMP-2, while MMP-2 digests type V collagen effectively. Biosynthetic studies of MMPs using cultured human rheumatoid synovial fibroblasts indicated that the production of both proMMP-1 and proMMP-3 is negligible but it is greatly enhanced by the treatment with rabbit-macrophage-conditioned medium, whereas the synthesis of proMMP-2 is constitutively expressed by these cells and is not significantly affected by the treatment. This suggests that the physiological and/or pathological role of MMP-2 and its site of action may be different from those of MMP-1 and MMP-3.
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PMID:Matrix metalloproteinase 2 from human rheumatoid synovial fibroblasts. Purification and activation of the precursor and enzymic properties. 226 96

A method for large-scale production of hepatocytes on microcarriers have been developed for the purpose of bioartificial liver support system. Hepatocytes obtained by collagenase treatment of rat liver were efficiently attached and spread on a microcarrier surface in the presence of O2-saturated perfluorodecalin. In order to compare the metabolic activities of hepatocytes under long-term cultivation on microcarriers with those of cells under conventional monolayer culture, some liver-specific functions were investigated. Microcarrier-attached hepatocytes cultured in the absence of serum for 8 days synthesized and secreted albumin and fibronectin. Moreover, hepatocytes on microcarriers retained the ability to conjugate bilirubin for 4-5 days. With respect to these specific metabolic properties, microcarrier-attached hepatocytes were comparable to those from routine dish culture. These results suggest that this method developed for large-scale production of hepatocytes on microcarriers will allow one to obtain metabolically active cells suitable for extracorporeal liver support systems.
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PMID:Large-scale production and cultivation of hepatocytes on Biosilon microcarriers. 228 91

Basement membrane collagen from bovine anterior lens capsules (ALC) was isolated by a non-degradative procedure and characterized. The material was obtained by extracting the ALC with 0.5 M acetic acid, passing the extract through a DEAE-cellulose column, collecting and concentrating the unbound fraction. Amino acid and carbohydrate analysis, polyacrylamide gel electrophoresis and rotary shadowing electron microscopy showed the purified extract to have the characteristics of basement membrane procollagen. Examination by rotary shadowing revealed the material to consist of type IV procollagen-like tetramers in various degrees of aggregation. When this purified procollagen was injected into rabbits, antibody of high titer and specificity was obtained. The competitive ELISA was then used to demonstrate lack of reactivity of the antibody with collagen types I, II, III and V and fibronectin. The electroimmunoblot technique was used to demonstrate lack of reactivity with laminin. Competition with collagenase resistant fragments of type IV procollagen representing the carboxyl terminal domain (NCI) and the 7-S domain, as well as with a pepsin-resistant alpha 1 (IV)125K chain was markedly weaker as compared to the native type IV procollagen molecule.
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PMID:Immunochemical characterization of type IV procollagen from anterior lens capsule. 241 54

Elastin was isolated from the human aorta by three different extraction methods. Immunization was carried out in sheep. The presence of antibody against each elastin preparation in the sheep sera was confirmed by immunofluorescence. However, antiserum against elastin isolated by collagenase/guanidine dithioerythritol showed the most specific reactions in the cryostat sections. No cross-reactivity to type I, III and IV collagen, fibronectin, laminin and proteoglycan BM 1 was observed.
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PMID:Preparation of a specific antiserum against elastin of the human aorta. 242 14

This study elucidates the biochemical response of rabbit corneal keratocytes (fibroblasts) to retinol and retinoic acid in their production of collagen, fibronectin, sulfated glycosaminoglycans, collagenase, and [3H]thymidine incorporation. The morphologic appearance of cultured keratocytes was not altered by retinoid treatment. Collagen production and [3H]thymidine incorporation demonstrated a parallel decline in response to retinoids. Collagen type was unaffected as was collagenase activity. Retinoids had minimal effect on cell layer-associated 35S-labeled glycosaminoglycans, however medium-soluble 35S-glycosaminoglycans were increased. The most dramatic effect was in fibronectin synthesis which was increased 2-3-fold. These data demonstrate that rabbit keratocytes alter their biosynthesis of extracellular matrices in response to retinoids. This may be significant in corneal pathology, since the delicate balance of these extracellular macromolecules is responsible for corneal integrity and stability.
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PMID:Modulation of rabbit keratocyte production of collagen, sulfated glycosaminoglycans and fibronectin by retinol and retinoic acid. 243 Jun 25

In order to study human bile duct cells in vitro, cystic ducts were obtained during cholecystectomy and treated with collagenase and mechanical abrasion to isolate biliary epithelial cells. The culture medium was supplemented with 50% of a bovine bile duct conditioned medium obtained by incubating minced bovine extrahepatic bile ducts for 24 hr in Dulbecco's modified Eagle's medium. Cells grew in monolayer and showed contact inhibition at confluency. The epithelial origin of primary cultures was verified by their growth pattern, ultrastructure, and indirect immunofluorescence for cytokeratin. The cultures showed specific immunofluorescence for lysozyme, collagen types I, III, and IV, fibronectin, and laminin, but were negative for collagen type V and factor VIII-associated antigen. Thus, these cultures provide an experimental model for the in vitro study of biliary atresia and other bile duct diseases.
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PMID:Characterization of human extrahepatic biliary duct epithelial cells in culture. 245 76

Baby hamster kidney cells were seeded onto Western blots of fetal serum proteins which had been extracted from several foreign surfaces. This revealed that the major cell adhesive proteins adsorbed onto these surfaces from fetal serum were (1) fibronectin of Mr 220,000 Da and (2) vitronectin of Mr 65,000 and 78,000 Da. Two minor bands of cell attachment were observed at Mr 153,000 and Mr 134,000 Da in the fetal serum proteins extracted from heparin-agarose and serotonin-agarose. However, by exposing the Western blots of separated proteins to a second round of serum proteins, prior to cell blotting, very strong cell adhesive bands were revealed at Mr 153,000, 134,000, and 120,000 Da. By (i) modifying the composition of the serum proteins used to treat the Western blots, (ii) using specific antibodies to fibronectin, and (iii) using radiolabeled fibronectin, it was conclusively demonstrated that the new cell adhesive bands owed their increased cell attachment activity to secondary binding of fibronectin. The new bands were shown (i) to be trypsin sensitive and collagenase sensitive and therefore to be collagen-like proteins and (ii) to react negatively in immunoblots using anti-fibronectin, anti-vitronectin, anti-fibrinogen, anti-fetuin or anti-thrombospondin. In SDS-PAGE (i) the Mr 120,000-Da protein comigrated with the alpha 2-chain of Type I collagen, (ii) the Mr 134,000-Da protein comigrated with the alpha 1-chain of Type I collagen, and (iii) the Mr 153,000-Da protein comigrated with the pN-alpha 1-chain of Type III collagen. Since the novel collagen-like proteins acted as strong sites of cell attachment on nitrocellulose blots by binding fibronectin, they might well promote cell attachment on the foreign surfaces from which they were extracted.
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PMID:Adsorption from fetal calf serum of collagen-like proteins which bind fibronectin and promote cell attachment. 245 51

In order to determine whether interferons (IFNs) play a universal role in terminating the fibrotic response by inhibiting other fibroblast functions in addition to growth and collagen production, we investigated the effect of human recombinant (hu-r) IFN-alpha, -beta, and -gamma on the glycosaminoglycan, fibronectin, and collagenase production of cultured human dermal fibroblasts. Our results show that short-term (48 h) treatment of confluent fibroblast cultures with hu-r-IFN-alpha 2 and hu-r-IFN-beta-ser17 causes a concentration (1 to 1 x 10(5) U/ml)-dependent inhibition of glycosaminoglycan production, has no effect on fibronectin production, and markedly increases collagenase production. In contrast, hu-r-IFN-gamma not only causes a concentration-dependent increase in collagenase production but also increases both glycosaminoglycan and fibronectin production. These results demonstrate that IFNs differently regulate fibroblast functions rather than universally inhibit all functions, and show that IFN-alpha and -beta exhibit a broader antifibrotic spectrum that IFN-gamma.
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PMID:Differential regulation of glycosaminoglycan, fibronectin, and collagenase production in cultured human dermal fibroblasts by interferon-alpha, -beta, and -gamma. 247 67

Fat-storing cells were isolated and purified from livers of normal adult rats and maintained in primary culture. By light and electron microscopy it was established that they underwent phenotypic changes into cells with the ultrastructural characteristics of myofibroblasts, between the third and sixth day in culture. These morphological changes were accompanied by a 2-fold increase of L-[3H]proline incorporation into secretory proteins and an 11-fold increase into secreted collagenase-sensitive proteins. In contrast, incorporation into cell layer-associated proteins and into cell layer-associated collagenase-sensitive proteins was not significantly elevated. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in combination with fluorography, demonstrated that the main collagen type secreted by the myofibroblast-like cells was collagen type I. Collagen types III and IV, and fibronectin were present in lesser amounts. The similarity between the well known in vivo alterations of fat-storing cells under pathological conditions and the spontaneous in vitro differentiation described in this study, makes primary cultures of fat-storing cells a valuable tool for studying their role in chronic liver disease.
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PMID:In vitro differentiation of fat-storing cells parallels marked increase of collagen synthesis and secretion. 250 9


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