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

---

Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The three-dimensional structure of the extracellular substratum was found to regulate reversibly the morphology, proliferation and collagen synthesis of perisinusoidal stellate cells (lipocytes, i.e. fat-storing 'Ito' cells). On non-coated polystyrene and type I collagen-coated culture dishes, the cells spread well and extended their cellular processes. On the surface of type I collagen gels, the cells gathered and formed a mesh-like structure. However, in type I collagen gel where the cells were surrounded by type I collagen three-dimensionally, the cells extended their fine cellular processes and resembled the star-shaped stellate cells seen in vivo. The cell proliferation was more prominent in culture dishes coated with type I collagen or in polystyrene culture dishes than on or in type I collagen gels. The collagen synthesis was affected in the same manner. These data indicate that the nature and the three-dimensional structure of the extracellular matrix (ECM) can regulate morphology, proliferation and functions of the perisinusoidal stellate cells. In order to examine the reversibility of these regulations, we liberated cultured cells with trypsin or with purified bacterial collagenase and re-seeded them onto or into each substratum. The cells changed their shape, rate of proliferation and collagen synthesis according to each new substratum. These results indicate that the three-dimensional structure of ECM reversibly regulates the morphology, proliferation rate and functions of the perisinusoidal stellate cells.
...
PMID:Three-dimensional structure of extracellular matrix reversibly regulates morphology, proliferation and collagen metabolism of perisinusoidal stellate cells (vitamin A-storing cells). 893 17

An original and reliable technique to culture growth plate chondrocytes was developed to obtain an abundant amount of mature and functional chondrocytes. Growth plates were provided from the epiphysis of 3 week old rabbits. The isolation of the chondrocytes was optimized by the use of trypsin and collagenase. The culture was realized according to the following conditions: seeding at 20,000 or 30,000/cm2 on type I collagen substrate and in Ham F12 medium without a supplementation of glucose or growth factors. After 7 days of culture, the implantation was to be carried out. Different implantation substrates were evaluated in vivo. Agar turned out to be the only substrate to provide strong and healthy chondrocytes 21 days after the grafting. In an other experimentation, the culture was implanted into surgically created defects in the growth plate area. In this case, the culture did not avoid the occurrence of an epiphysiodesis. However, the 6 weeks post operative histologic examination, showed that the implant remained viable, continued to maintain a proteoglycan rich matrix, and began to organize in ordered columns of mature chondrocytes.
...
PMID:[In vitro culture of growth cartilage and in situ reimplantation]. 894 14

Mast cell activation is a characteristic feature of chronic inflammation, a condition that may lead to fibrosis as a result of increased collagen synthesis by fibroblasts. We have investigated the potential of tryptase, the major protease of human mast cells, to stimulate collagen synthesis in the human lung fibroblast cell line MRC-5. Tryptase was isolated from human lung tissue by ion-exchange and affinity chromatography. At concentrations of 18 and 36 mU/ml, tryptase stimulated both an increase in cell numbers, and a fivefold increase in DNA synthesis as determined by methyl-[3H]thymidine incorporation. Similar concentrations of tryptase resulted in a 2.5-fold increase in collagen synthesis as determined both by incorporation of [3H]proline into collagen, and by assay of hydroxyproline concentrations in the supernatants. There was also a twofold increase in collagenolytic activity in the culture medium after tryptase treatment, indicating that the increase in collagen synthesis was not a consequence of decreased collagenase production. All of these actions of tryptase were reduced in the presence of the protease inhibitors leupeptin and benzamidine hydrochloride, indicating a requirement for an active catalytic site. SDS-PAGE and autoradiographic analysis of the [3H]collagen produced by the cells revealed it to be predominantly type I collagen. Our findings suggest that the release of tryptase from activated mast cells may provide a signal for abnormal fibrosis in inflammatory disease.
...
PMID:Mast cell tryptase stimulates the synthesis of type I collagen in human lung fibroblasts. 907 41

The propeptide plus the catalytic domain of human fibroblast-type collagenase, stromelysin-1, and matrilysin were expressed in Escherichia coli to directly compare the properties of all three catalytic domains utilizing the same assays. Truncated fibroblast-type collagenase (mini-CL), truncated stromelysin-1 (mini-SL-1), and matrilysin, like their native counterparts, could be activated by organomercurials, trypsin, or SDS. The mini-CL and mini-SL-1 displayed catalytic properties similar to their native counterparts, except that the mini-CL could not cleave native type I collagen. The k(cat)/Km for matrilysin (355 microM(-1) h(-1)) on the synthetic Mca-peptide was much higher than that for mini-CL (69 microM(-1) h(-1)) or mini-SL-1 (23.6 microM(-1) h(-1)). Mini-SL-1 and matrilysin, but not mini-CL, were capable of superactivating collagenase thus increasing the rate of collagen cleavage. Mini-CL and mini-SL-1, but not matrilysin, were able to form SDS-stable complexes with TIMP-1 when co-incubated with an organomercurial and TIMP-1. The second-order rate constant (k(on)) for TIMP-1 inhibition of mini-CL and mini-SL-1 were similar, 0.635 x 10(5) M(-1) s(-1) and 1.52 x 10(5) M(-1) s(-1), respectively. The k(on) for TIMP-1 inhibition of matrilysin was lower (0.130 x 10(5) M(-1) s(-1)) supporting the observation that no SDS stable complexes were detected. This study demonstrates that these catalytic domains are distinct and play a major role in the specificity of these enzymes in regard to rate of catalysis, TIMP-1 binding, and superactivation of collagenase.
...
PMID:Catalytic domain comparisons of human fibroblast-type collagenase, stromelysin-1, and matrilysin. 910 22

Prolidase (EC 3.4.13.9) is a ubiquitously distributed imidodipeptidase that catalyzes the hydrolysis of C-terminal proline or hydroxyproline containing dipeptides. The enzyme plays an important role in the recycling of proline for collagen synthesis and cell growth. An increase in enzyme activity is correlated with increased rates of collagen turnover indicative of extracellular matrix (ECM) remodeling, but the mechanism linking prolidase activity and ECM is poorly understood. Thus, the effect of ECM-cell interaction on intracellular prolidase activity is of special interest. In cultured human skin fibroblasts, the interaction with ECM and, more specifically, type I collagen mediated by the beta 1 integrin receptor regulates cellular prolidase activity. Supporting evidence comes from the following observations: 1) in sparse cells with a low amount of ECM collagen or in confluent cells in which ECM collagen was removed by collagenase (but not by trypsin or elastase) treatment, prolidase activity was decreased; 2) this effect was reversed by the addition of type I collagen or beta 1 integrin antibody (agonist for beta 1 integrin receptor); 3) sparse cells (with typically low prolidase activity) showed increased prolidase activity when grown on plates coated with type I collagen or on type IV collagen and laminin, constituents of basement membrane; 4) the relative differences in prolidase activity due to collagenase treatment and subsequent recovery of the activity by beta 1 integrin antibody or type I collagen treatment were accompanied by parallel differences in the amount of the enzyme protein recovered from these cells, as shown by Western immunoblot analysis. Thus, we conclude that prolidase activity responded to ECM metabolism (tissue remodeling) through signals mediated by the integrin receptor.
...
PMID:Prolidase activity in fibroblasts is regulated by interaction of extracellular matrix with cell surface integrin receptors. 932 22

A useful gelatinolytic enzyme assay for fibroblasts, utilizing a novel sample preparation method for collagenase with dithiothreitol (DTT) treatment to inactivate endogenous collagenase inhibitors, was developed using soluble fluorescein isothiocyanate (FITC)-labeled gelatin. The substrate, gelatin was prepared by heating commercially available FITC-labeled type I collagen. The denatured collagen was cleaved with purified trypsin and partially purified fibroblast gelatinase, and the digested FITC-fragments were measured fluorometrically. The intensity of the fluorescence was in proportion to the reaction time and enzyme concentration. Both enzyme activities were measurable within the nanogram range of enzyme preparations. The enzyme activity was detected after 4-aminophenylmercuric acetate (APMA) treatment which was completely inhibited by metalloproteinase inhibitors, but not by serine- and cysteine-proteinases' inhibitors. Conditioned media of human periodontal ligament fibroblasts (PLF) and gingival fibroblasts (GF) were separately treated with DTT prior to the enzyme assay, and then the assay was performed in the presence of APMA. The enzyme activities of PLF and GF were 106- and 55-fold higher than those of the conventional gelatinase assay which was carried out without DTT treatment. This assay method allowed the measurement of gelatinolytic enzyme activity when tissue inhibitors of metalloproteinases were present in the fibroblast culture medium.
...
PMID:An improved assay method for fibroblast gelatinolytic enzyme. 947 31

Dermatosparaxis is a recessive disorder of animals (including man) which is caused by mutations in the gene for the enzyme procollagen N-proteinase and is characterised by extreme skin fragility. Partial loss of enzyme activity results in accumulation of pNcollagen (collagen with N-propeptides) and abnormal collagen fibrils in the fragile skin. How the N-propeptides persist in the tissue and how abnormal fibril morphology results in fragile skin is poorly understood. Using biochemical and quantitative mass mapping electron microscopy we showed that the collagen fibrils in the skin of a dermatosparactic calf contained 57% type I pNcollagen and 43% type I collagen and the fibrils were irregularly arranged in bundles and hieroglyphic in cross-section. Image analysis of the fibril cross-sections suggested that the deviation from circularity of dermatosparactic fibrils was caused by N-propeptides of pNcollagen being located at the fibril surface. Comparison of experimental and theoretical axial mass distributions of the fibrils showed that the N-propeptides were located to the overlap zone of the fibril D-period (where D=67 nm, the characteristic axial periodicity of collagen fibrils). Treatment of the dermatosparactic fibrils with N-proteinase did not remove the N-propeptides from the fibrils, although the N-propeptides were efficiently removed by trypsin and chymotrypsin. However, the N-propeptides were efficiently cleaved by the N-proteinase when the pNcollagen molecules were extracted from the fibrils. These results are consistent with close packing of N-propeptides at the fibril surface which prevented cleavage by the N-proteinase. Long-range axial mass determination along the fibril length showed gross non-uniformity with multiple mass bulges. Of note is the skin fragility in dermatosparaxis, and also the appearance of mass bulges along the fibril long axis symptomatic of the fragile skin of mice which lack decorin. Western blot analysis showed that the dermatosparactic fibrils bound elevated levels of the proteoglycan, compared with normal skin fibrils. The results showed that N-propeptides can distort the morphology of fibrils, that they do not inhibit binding of gap-associated macromolecules (such as decorin) and that the normal mechanical properties of skin are strongly dependent on the close association of near-cylindrical fibrils, thereby enabling maximal fibril-fibril interactions.
...
PMID:Surface located procollagen N-propeptides on dermatosparactic collagen fibrils are not cleaved by procollagen N-proteinase and do not inhibit binding of decorin to the fibril surface. 957 Oct 43

An original and reliable technique to culture growth plate chondrocytes was developed to obtain an abundant amount of mature and functional chondrocytes. Growth plates were provided from the epiphysis of 3-week-old rabbits. Isolation of the chondrocytes was optimized by the use of trypsin and collagenase. The culture was realized according to the following conditions: seeding at 20,000 or 30,000/cm2 on type I collagen substrate and in Ham F-12 medium without a supplementation of glucose or growth factors. After 7 days of culture, the implantation was to be carried out. Different implantation substrates were evaluated in vivo. Agar turned out to be the only substrate to provide strong and healthy chondrocytes 21 days after the grafting. Then implantation was tested on large iliac resections in rabbits to check whether an enchondral ossification occurred with the culture. Poor results were obtained because of an early disappearance of the cultured chondrocytes. In an other experimentation, the culture was implanted into surgically created defects in the growth plate area. In this case, the culture did produce an epiphysiodesis. However, the 6-week postoperative histological examination showed that the implant remained viable, continued to maintain a proteoglycanrich matrix, and began to organize in ordered columns of mature chondrocytes.
...
PMID:Reimplantation of growth plate chondrocyte cultures in central growth plate defects: Part II. Surgical experimentation in rabbits. 959 98

Matrix proteinases play a critical role in extracellular matrix remodeling, which is particularly involved in cancer invasion and metastasis. We have previously characterized and purified a new tetrameric serine proteinase (SP220K) from human kidney clear cell carcinoma plasma membranes. Here, we report that SP220K exhibits gelatinase activity as assessed both in solution and by zymography. Optimum gelatinase activity ranges between pH 7.5 to pH 9.0. Fibronectin and type I collagen were hydrolyzed by SP220K, at variance with laminin and type IV collagen. Like other trypsin-like fibronectin degrading proteinases, SP220K released the 29-kDa N-terminal heparin-binding domain of fibronectin. By using a panel of proteinase inhibitors, we found that the inhibition profile of SP220K was different from that of other known serine proteinases such as thrombin, trypsin, plasmin, plasminogen activators and tryptase. Altogether, our results indicate that SP220K corresponds to a novel matrix proteinase that exhibits a marked specificity for fibronectin and type I collagen.
...
PMID:SP220K is a novel matrix serine proteinase. 965 May 63

Type I and II collagen (native-type) fibrils, positively stained with uranyl acetate, present typical periodic (D = 67 nm) cross-striation patterns. Although the two patterns are similar, the distributions of charged amino acids along the type I and II collagen molecules are different. After optical diffraction analysis or computer image processing of electron micrographs, different Fourier transforms were obtained from type I and II collagen fibrils, either as native fibrils or after in vitro reconstitution from purified molecules. With tissues such as tendon and cartilage, better results were obtained after mild trypsin treatment, which allowed better isolation and staining of the collagen fibrils. The main difference observed in the Fourier transforms was the presence in type II collagen fibrils of a strong tenth-order peak (corresponding to the tenth harmonic of the fundamental frequency). In order to discriminate between the two collagens, we measured the ratio (R) of the areas under the ninth- and tenth-order peaks. In trypsin treated tissues, the distributions of these ratios were clearly separated: below 1.0 for type II collagen fibrils and above 1.5 for type I collagen fibrils. This method appears to be suitable for rapid typing of type I and II collagen fibrils and might be useful for determining the exact composition of fibrils in tissues, such as intervertebral discs, that contain these both types of collagen.
...
PMID:Fourier analysis of electron micrographs of positively stained collagen fibrils: application to type I and II collagen typing. 977 8


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---