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)

Rat 1a fibroblasts transformed by the Gi2 oncogene, gip2, exhibit a constitutively elevated mitogen-activated protein (MAP) kinase activity that correlates with enhanced tyrosine phosphorylation of the p42 MAP kinase polypeptide. The MAP kinase activity in gip2 transformed cells is 50-60% of the pertussis toxin-sensitive, thrombin-stimulated activity observed in wild-type Rat 1a cells. A similar activation of MAP kinase is observed in src but not ras or raf transformed Rat 1a cells, indicating that the persistent MAP kinase activity results from the action of the specific oncoprotein and is not the consequence of cellular transformation. The enhanced transactivation function of c-Jun characteristic of the transformed phenotype, measured using a collagenase promoter-CAT reporter gene, is observed in gip2, src, ras, and raf transformed Rat 1a cells. The regulatory networks controlled by the four transforming oncogenes therefore alter the activity of specific transcription factors, but only gip2 and src constitutively activate MAP kinase. The findings demonstrate that the catalytic activity of growth factor-regulated cytoplasmic kinases are selectively and stably activated as a consequence of specific oncogene expression.
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PMID:MAP kinase is constitutively activated in gip2 and src transformed rat 1a fibroblasts. 131 14

The mechanisms involved in retinoic acid (RA)-mediated regulation of the collagenase gene in a rabbit synovial fibroblast cell line (HIG82) were investigated. When HIG82 cells are cotransfected with expression vectors containing cDNAs for retinoic acid receptor (RAR) alpha 1, beta 2, or gamma 1 and collagenase promoter-driven CAT reporter constructs, only RAR-gamma 1 represses basal CAT expression upon RA treatment, while RAR-alpha 1, beta 2, and gamma 1 all suppress phorbol-induced CAT expression. Thus, transcriptional regulation of collagenase by RA is mediated by RARs in an RAR-type specific manner. Using mutational and deletional analysis, we find that interaction between elements within 182 bp collagenase promoter plays an important role in this process. In addition, cotreatment with RA results in a decrease of phorbol-induced mRNA levels of fos and jun, and binding of nuclear proteins to an AP-1 oligonucleotide. Furthermore, RA-induced nuclear protein(s) specifically bind to a 22 bp sequence (-182 to -161) of the collagenase promoter. We propose that RA-mediated regulation of the collagenase gene depends on the availability and interaction of specific RARs with multiple DNA elements within the promoter and with transcription factors, including AP-1 related proteins.
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PMID:Differential regulation of collagenase gene expression by retinoic acid receptors--alpha, beta and gamma. 132 Feb 54

In this paper, the role of reactive oxygen species in photoaging is presented. Many photosensitizing agents are known to generate reactive oxygen species (singlet oxygen (1O2), superoxide anion (O2.-) and .OH radicals). Although photoaging (dermatoheliosis) of human skin is caused by UVB and UVA radiation, the hypothesis tested here in the pathogenesis of photoaging of human skin is the free radical theory involving the generation of reactive oxygen species by UVA (320-400 nm) radiation and their damaging oxidative effects on cutaneous collagen and other model proteins. The UVA-generated reactive oxygen species cause cross-linking of proteins (e.g. collagen), oxidation of sulfydryl groups causing disulfide cross-links, oxidative inactivation of certain enzymes causing functional impairment of cells (fibroblasts, keratinocytes, melanocytes, Langerhans cells) and liberation of proteases, collagenase and elastase. The skin-damaging effects of UVA appear to result from type II, oxygen-mediated photodynamic reactions in which UVA or near-UV radiation in the presence of certain photosensitizing chromophores (e.g., riboflavin, porphyrins, nicotinamide adenine dinucleotide phosphate (NADPH), etc.) leads to the formation of reactive oxygen species (1O2, O2.-, .OH). Four specific observations are presented to illustrate the concept: (1) the production of 1O2 and O2.- by UVB, UVA and UVA plus photosensitizing agents (such as riboflavin, porphyrin and 3-carbethoxypsoralens) as a function of UV exposure dose, the sensitizer concentration and the pH of the irradiated solution; (2) the formation of protein cross-links in collagen, catalase and superoxide dismutase by 1O2 and O2.- (.OH) and the resulting denaturation of proteins and enzyme activities as a function of UVA exposure dose; (3) the protective role of selective quenchers of 1O2 and O2.- (e.g. alpha-tocopherol acetate, beta-carotene, sodium azide, ascorbic acid, etc.) against the photoinactivation of enzymes and the prevention of the protein cross-linking reaction; (4) the possible usefulness of certain antioxidants or quenchers that interact with the UVA-induced generation of reactive oxygen species in the amelioration of the process of photoaging.
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PMID:Skin photosensitizing agents and the role of reactive oxygen species in photoaging. 133 86

Fifty-four strains of Peptostreptococcus magnus (11 were recovered from abdominal infections, 18 were from nonpuerperal breast abscesses, and 21 were from diabetic foot infections; the type strain and three other strains were from the American Type Culture Collection, Rockville, Md.) and the type strain of Peptostreptococcus micros were tested for their ability to produce various enzymes, including catalase, hippurate hydrolase, serine dehydratase, threonine dehydratase, collagenase, gelatinase, alkaline phosphatase, and esterase C4. The data were analyzed by cluster analysis. The results showed that all but one strain could be assigned to either of two distinct, valid clusters. The first cluster of 11 strains was composed of strains that were relatively inactive, having produced one or two of the eight strain-dependent enzymes. The second was a large cluster of strains (n = 43) that were considerably more active, all having produced at least three enzymes; the vast majority of strains (89%) produced four or more enzymes. The unclustered strain produced one enzyme that was different from that produced by the strains in the first cluster. The chi 2 test of homogeneity applied to the clustering solution indicated that greater enzyme activity was significantly associated with the site of infection (P less than 0.001). The more enzymatically active P. magnus strains were recovered significantly more often from nonpuerperal breast abscesses and diabetic foot infections than they were from abdominal infections. These results may provide insight into the nature of certain polymicrobial soft tissue infections and suggest that (i) P. magnus may participate more in nonpuerperal breast and diabetic foot infections than in abdominal infections and that (ii) peptostreptococcal production of proteolytic enzymes may have an important adjunctive effect on the pathogenesis of certain soft tissue infections.
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PMID:Enzymatically active Peptostreptococcus magnus: association with site of infection. 140 Sep 97

Hydrogen peroxide (H2O2) serves as a precursor for highly reactive oxygen intermediates. However, the respiratory function of myocytes is relatively resistant to exogenously administered H2O2. In this study, we examined whether or not the reduction of cellular defense increases the toxicity of H2O2. Rat heart myocytes were isolated by collagenase digestion. Respiratory rates of myocytes, suspended in a medium containing sucrose, 3-N-morpholino-propanesulfonic acid, EGTA and bovine serum albumin, were determined polarographically in the presence of pyruvate and malate with or without 2,4-dinitrophenol (DNP). Mitochondrial membrane potentials were measured by using [3H]triphenylmethylphosphonium+. Cellular defense was attenuated by i) inhibiting the catalase activity by 3-amino-1,2,4-triazole (AT), ii) reducing the glutathione concentration by diethyl maleate (DEM) or ethacrinic acid (EA), and iii) permeabilizing the sarcolemmal membrane by saponin. The dose-response relationship between H2O2 (0.1-5 mM) and mitochondrial membrane potential was not greatly affected by these experimental conditions. Myocyte respiration was inhibited by 5 mM H2O2, particularly that measured in the presence of DNP (48% of control). DEM treatment did not significantly affect the respiratory inhibition by H2O2, whereas the degree of inhibition was somewhat greater following EA or AT treatment. By contrast, the sensitivity of cellular respiration to H2O2 was potentiated approximately two orders of magnitude by the permeabilization of sarcolemmal membrane; thus, 100 microM H2O2 inhibited both DNP-stimulated and unstimulated respiration to 17% and 35% of control, respectively. The results indicate that factors existing in the sarcolemma and/or in the cytosol, which become ineffective and/or are diluted, respectively, following permeabilization with saponin, are important cellular defense mechanisms in alleviating the toxic effect of exogenous H2O2 on the respiration of mitochondria in situ in myocytes.
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PMID:Role of cellular defense against hydrogen peroxide-induced inhibition of myocyte respiration. 152 Feb 49

Isolation and culture techniques for hepatocytes from whole livers of the cynomolgus monkey, Macaca fascicularis, are described. Hepatocytes were isolated by two-step perfusion of livers, using collagenase with hyaluronidase; fructose and trypsin inhibitor were included to reduce cell loss. Yields from a single liver average 4 X 10(9) cells with viabilities of 90.8 +/- 5.7%. Cells, plated on collagen substrates, were assessed for changes in morphology and various marker enzyme activities over a period of 7 d in culture. Cells exhibited a morphology similar to that observed for this species in vivo; little change in attached and spread cells was observed over the length of time monitored. Enzyme activities for catalase, succinate dehydrogenase, and tyrosine aminotransferase were observed to decrease significantly (though considerable activity remained), whereas acid phosphatase and 5'-nucleotide phosphodiesterase remained unchanged. Activity of cytochrome P-450 reductase was observed to increase slightly for the first 2 d, then decrease to about 60% of initial levels. Activity of alpha-mannosidase was stable for 4 d but was observed to be increased at Day 7. Cells were observed to retain metabolic responsiveness, demonstrated by glucose production by both gluconeogenesis and glycogenolysis in response to glucagon stimulation. The monkey hepatocytes obtained by methods described here thus retain hepatocellular morphology and activity through at least 1 wk in culture without medium or culture modification.
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PMID:Isolation and culture of hepatocytes from the cynomolgus monkey (Macaca fascicularis). 197 77

Ursodeoxycholate (UDC) and tauroursodeoxycholate (TUDC) have been reported to be protective against liver injury induced by other bile salts. UDC also has been shown to be effective against refluxed bile-induced gastritis after gastric surgery. However the mechanism of the therapeutic effect of UDC on gastric mucosa has not been known. In the present study, cytoprotective actions of UDC and TUDC against chenodeoxycholate (CDC)-induced gastric injury were investigated using rabbit gastric cell cultures without systemic factors. Rabbit gastric mucosal cells were cultured after the isolation of rabbit gastric cells with collagenase and ethylenediaminetetraacetic acid. Cytotoxicity was quantified by measuring 51Cr release from prelabeled cells and MTT assay. Prostaglandin (PG) E2 was assayed by radioimmunoassay. Concentrations of CDC greater than 0.5 mM or UDC greater than 5 mM caused cellular damage and increased 51Cr release in a dose-dependent and time-dependent fashion, while TUDC up to 10 mM did not. TUDC, but not UDC, showed a significant decrease of CDC (1.5 mM)-induced 51Cr release dose dependently. The protective effect of TUDC against CDC-induced damage was confirmed by MTT assay. On phase-contrast microscopy, disruption of monolayers induced by CDC (1.5 mM) was clearly protected by TUDC (10 mM). Free radical scavengers (500 units/ml of superoxide dismutase, 300 units/ml of catalase, and 100 mM of dimethyl sulfoxide) or a calcium blocker (10(-7)-10(-5) M verapamil) did not show significant protection against CDC-induced damage. Deprivation of Ca2+ in the media did not affect CDC-induced damage. Thus free radicals or Ca2+ might not be involved in the cell toxicity of CDC.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Protective effect of tauroursodeoxycholate against chenodeoxycholate-induced damage to cultured rabbit gastric cells. 200 57

Free radical damage has the potential to significantly affect the behavior of cells in culture. In this study the effects of antioxidants (superoxide dismutase, catalase, and vitamin E) and lowered oxygen tension (1% oxygen) on primary culture of rat mammary epithelial cells were examined. Rat mammary epithelial cells were dissociated in collagenase with or without the addition of antioxidants and low oxygen tension, then cultured for 10 d in rat-tail collagen gel matrix and fed with Dulbecco's modified Eagle's F12 medium supplemented with various hormones and growth factors. Growth potential of the mammary cells was enhanced when antioxidants and low oxygen tension were used, alone or in combination, during the cell dissociation period. Using antioxidants and low oxygen tension during the culture period failed to improve growth potential regardless whether cells were dissociated in standard conditions or with antioxidants and low oxygen tension. The use of antioxidants and low oxygen tension during the cell dissociation period also reduced the degree of keratinization of the cells after 10 d of culture. Using antioxidants and low oxygen tension during the cell culture period did not further reduce keratinization if antioxidants and low oxygen tension were used during the dissociation period, but were effective in reducing keratinization if cells were dissociated in standard condition. In this system, antioxidants and low oxygen tension reduced lipid peroxidation during the cell dissociation period. An iron chelator, desferal, can also reduce lipid peroxidation and enhance growth when used during cell dissociation, suggesting the enhanced growth potential by the addition of antioxidants and low oxygen to be due to the reduction of lipid peroxidation.
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PMID:Effects of antioxidants and reduced oxygen tension on rat mammary epithelial cells in culture. 203 19

Hepatocytes harvested by collagenase perfusion of rat liver were attached to collagen-coated microcarriers and injected intraperitoneally into congeneic or allogeneic bilirubin-UDP-glucuronosyltransferase (EC 2.4.1.17)-deficient (Gunn) rats or allogeneic analbuminemic (NAR) rats. Five days later, the microcarriers were observed to have formed conglomerates chiefly on the anterior surface of the pancreas. Scanning electron microscopy showed hepatocytes attached to the granular collagen-coated surface of the microcarriers and newly formed connective tissue. Light microscopy revealed that the microcarriers formed a lattice with the collagen tissue; hepatocytes were seen within this lattice or on the surface of the microcarriers. Hepatocyte plasma membranes were nucleoside-diphosphatase (NDPase)-positive. Newly formed blood islands, blood vessels containing erythrocytes and leukocytes and NDPase-positive endothelium were observed in close proximity to the hepatocytes and fibroblasts. Transmission electron microscopic examination showed hepatocytes with microvilli and nucleoid-containing peroxisomes with catalase activity. Hepatocytes were present for up to 2 months in congeneic recipients, the longest period of observation after transplantation. After normal microcarrier-attached hepatocytes were transplanted into allogeneic Gunn rats, bilirubin glucuronides were present in bile for 6 days. When congeneic Gunn rat recipients were used, bilirubin glucuronides were present in bile throughout the study (28 days); this was accompanied by reduction of serum bilirubin concentrations to nearly normal levels. After injection of normal hepatocytes into allogeneic NAR rats, plasma albumin concentration progressively increased for 6 days and then declined. In NAR recipients which were immunosuppressed with cyclosporin A, peak plasma albumin levels were reached in 14 days and persisted nearly at that level throughout the study (28 days).
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PMID:Survival, organization, and function of microcarrier-attached hepatocytes transplanted in rats. 242 7

The influence of the endothelium on pulmonary venular responses to reduced oxygen tension has not been defined. To examine this question, endothelial injury was induced in small guinea pig pulmonary artery and venule segments (effective lumen radius, 174 +/- 5 and 122 +/- 2 microns, respectively) by perfusion with either a mixture of hypoxanthine (5 mM) and xanthine oxidase (0.05 U/ml) (HX/XO) or collagenase (2 mg/ml). HX/XO significantly (p less than 0.05) reduced the relaxation of precontracted pulmonary arteries by acetylcholine (ACH), bradykinin (BK), and A-23187, and the relaxations were restored by including superoxide dismutase (40 micrograms/ml) in the HX/XO solution. However, neither HX/XO nor collagenase affected vasodilation induced by ACH, BK, and A-23187 in precontracted pulmonary venules. In contrast, HX/XO significantly (p less than 0.05) augmented the sustained contraction of pulmonary venules to hypoxia (HX/XO, 3.2 +/- 1.0 mg/mm; control, 1.0 +/- 0.5 mg/mm) and anoxia (HX/XO, 35.1 +/- 6.6 mg/mm; control, 20.3 +/- 4.0 mg/mm). Collagenase also significantly (p less than 0.05) enhanced the anoxic contractions (collagenase, 36.0 +/- 3.7 mg/mm; control, 20.9 +/- 6.8 mg/mm). Superoxide dismutase (40 micrograms/ml) and catalase (323 micrograms/ml) abolished HX-XO-induced augmentation of the hypoxic and anoxic contractions of pulmonary venules. Collagenase removed 54 +/- 8% of the venular endothelium (control, 5 +/- 1%), whereas HX/XO-exposed endothelial cells contained numerous craters. Neither gossypol (5 microM) nor methylene blue (10 microM) affected pulmonary venular contractions to reduced PO2. Endothelial damage augments the PO2-dependent contractions of the pulmonary venule, and this augmentation does not appear to be due to decreased release of endothelium-derived relaxing factor.
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PMID:Effect of endothelial injury on the responses of isolated guinea pig pulmonary venules to reduced oxygen tension. 254 70


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