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)

In rat liver cells isolated by perfusion in the perfusion in the presence of collagenase, the major portion of cytochrome P-450 is present in the oxidized, nonsubstrate-bound, low spin state. Drug addition to a suspension of liver cells results in the rapid formation of the cytochrome P-450 (Fe3+)-substrate complex which in turn is followed by the appearance of other species with different spectral characteristics before steady state drug monooxygenation is achieved. Cytochrome P-450-linked metabolism of various tested drugs and carcinogenic polycyclic hydrocarbons by isolated rat liver cells is as fast, or faster, as with rat liver microsomes supplemented with a NADPH generating system. Both experimental models respond similarily to phenobarbital or 3-methylcholanthrene pretreatment of the animals and to various of the wellknown inhibitors of drug metabolism. Except with liver cells isolated from fasted, phenobarbital-treated rats, generation of cytosolic NADPH seems sufficient to support optimal drug metabolism even in the absence of added substrates of intermediary metabolism. In isolated liver cells oxidized drug metabolites undergo subsequent metabolic conversion, most often to form the corresponding glucuronides and sulphates. These are readily excreted, whereas non-conjugated products, e.g. free phenols, tend to accumulate intracellularly. Cellular glucuronide formation is strongly inhibited by ethanol-presumably due to an unfavorable effect of the increased NADH/NAD+ ratio on the synthesis of uridine-5'-diphosphoglucuronic acid (UDPGA). In contrast, low concentrations of ethanol have no, or only a slight stimulatory effect on the cytochrome P-450-linked step of drug metabolism and there are indications that the oxidation of low concentrations of ethanol is in fact stimulated by a facilitated reoxidation of cytosolic NADH occuring during drug monooxygenation.
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PMID:Recent studies on cytochrome P-450-linked functions in isolated rat liver cells. 0 26

In vitro addition of rat insulin (200, 400 or 800 muU/ml) to collagenase-isolated pancreatic islets of adult rats diminished glucose (3 mg/ml)-induced insulin release which was correlated with a decrease of the ratio of total NADPH/NADP and inhibition of glucose oxidation via the pentose phosphate shunt (PPS). NADH and NAD levels were not affected. It is suggested that exogenous insulin diminishes the islet total NADPH/NADP ratio by a direct or indirect decrease in PPS activity. However, it is also conceivable that insulin decreases this ratio through another mechanism than PPS. It is possible that inhibition of insulin secretion by exogenous insulin is at least in part due to the decrease of the NADPH/NADP ratio.
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PMID:Pyridine nucleotides in pancreatic islets during inhibition of insulin release by exogenous insulin. 1 90

The influence of the mode of preparation upon some of the characteristics of white adipose tissue plasma membranes and microsomes has been reported. Plasma membrane fractions prepared from mitochondrial pellet were shown to have higher specific activities of (Mg2+ + Na+ + K+)-ATPase than plasma membranes originating in crude microsomes. Isolation of fat cells by collagenase treatment was found to result in a decrease in specific activity of the plasma membrane enzymes; in plasma membranes prepared from isolated fat cells, the specific activity values obtained for (Mg2+ + Na+ +k+)-ATPase and 5'-nucleotidase were only 42% and 6.3% respectively of those obtained in plasma membranes prepared from whole adipose tissue. Purification of whole adipose tissue crude microsomes by hypotonic treatment caused extensive solubilization of the endoplasmic reticulum marker enzymes, NADH oxidase and NADPH cytochrome c reductase. The lability of endoplasmic reticulum marker enzymes, however, was found to be greatly diminished in the preparations from isolated fat cells. The possibility that NADH oxidase and NADPH cytochrome c reductase activities found in the plasma membranes are microsomal enzymes adsorbed by the plasma membranes is discussed. The peptide patterns as well as the NADH oxidase and NADPH cytochrome c reductase activity patterns of plasma membranes and purified microsomes were compared by means of sodium dodecyl sulfate or Triton X-100 polyacrylamide gel electrophoresis.
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PMID:Comparison of plasma membranes and endoplasmic reticulum fractions obtained from whole white adipose tissue and isolated adipocytes. 12 89

Testes of 29-35-day-old rats were separated into seminiferous tubules and interstitial tissue by collagenase treatment and examined by incubation studies with radioactive substrates. The activity of testosterone 5alpha-reductase/g protein in the interstitial tissue was 37 times greater than that in the tubules. The site of formation of 5alpha-reduced C21- and C19-steroids from progesterone was found to be primarily in the interstitial tissue. In the interstitial tissue, testosterone 5alpha-reductase activity, which was stimulated 300-fold by the addition of NADPH, was localized in the microsomal fraction (8,000-105,000 X g precipitate). NADPH was five times as effective a hydrogen donor as NADH when the washed microsomal fraction was the source of the enzyme. The formation of 5alpha-reduced C21- and C19-steroids from pregnenolone and progesterone was demonstrated in the microsomal fraction of interstitial tissue. These results indicate that in 29-35-day-old rat testes, 5alpha-reduction of C19-delta4-3-ketosteroids and the formation of 5alpha-reduced C21- and C19-steroids from pregnenolone take place largely in the microsomes of interstitial tissue.
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PMID:Localization of delta4-5alpha-reductase in immature rat testes. 74

There are two types of collagenases, products of two distinct genes, called MMP-1 (matrix metalloproteinase 1 or "fibroblast-type collagenase") and MMP-8 ("neutrophil collagenase"). In synovial fluid, MMP-8 is stored as latent proenzyme in polymorphonuclear neutrophils. MMP-8 is activated by hypochlorous acid produced by myeloperoxidase from hydrogen peroxide and chloride ion and by the hydroxyl radical produced in Haber Weiss reaction fed by superoxide produced by, eg, NADPH (reduced nicotinamide adenine dinucleotide) oxidase and xanthine oxidase. In addition to activation upon secretion, oxidatively modified MMP-8 is susceptible to a subsequent proteolytic attack and activation by cathepsin G. The authors suggest that activation of neutrophil-derived MMP-8 involves oxidative, nonproteolytic activation upon secretion and a more slowly progressive proteolytic activation by cathepsin G (or chymases and tryptases), and that these oxidative and proteolytic activation mechanisms act in concert. In contrast to MMP-8, MMP-1 is synthesized de novo and secreted immediately after synthesis by fibroblasts, macrophages, and some epithelial cells. Human rheumatoid synovial tissue contains mainly fibroblast-type MMP-1 collagenase as assessed by collagenase extracted from synovial tissue and by MMP-1 and MMP-8 immunostaining. It is suggested that in vivo, MMP-1 in synovitis tissue is activated by a plasminogen activator/plasminogen/prostromelysin (alternatively tryptases)/proMMP-1 cascade. In conclusion, MMP-8 and MMP-1 show type-specific compartmentalization and modes of activation in rheumatoid synovial fluid and tissue.
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PMID:Collagenase in synovitis of rheumatoid arthritis. 141 81

Cytochrome P450-dependent alkoxyphenoxazone dealkylase activity was measured in alveolar type II cells from control and beta-naphthoflavone (ip) treated-rats. Type II cells were isolated from collagenase/elastase-digested lung tissue and purified by centrifugal elutriation. The specificity of the cytochrome P450-dependent activity towards four alkoxyphenoxazones (methoxy-, ethoxy-, pentoxy-, and benzyloxyphenoxazone) was measured under conditions that minimized interference by cytosolic conjugating- and NADPH-dependent quinone reductase activities. Ethoxyphenoxazone dealkylase activity was induced 17-fold following beta-naphthoflavone treatment and was further characterized by its kinetic parameters and sensitivities toward in vitro inhibitors (Km(app) = 0.20 microM, Vmax = 1.74 pmoles resorufin min-1 (10(6) cells)-1 10(6) cells; I50 (alpha-naphthoflavone) = 0.025 microM, and I50 (metyrapone) = 72 microM). beta-Naphthoflavone pretreatment of the rats did not result in statistically significant changes in methoxy-, pentoxy-, or benzyloxyphenoxazone dealkylase activity of alveolar type II cells, although, a trend towards decrease activity was observed for benzyloxyphenoxazone. beta-Naphthoflavone pretreatment had no effect on oxygen consumption or trypan blue exclusion in alveolar type II cells and macrophage ethoxyphenoxazone dealkylase and benzyloxphenoxazone dealkylase activities were not affected by the beta-naththoflavone pretreatment. The results show that exposure to beta-naphthoflavone resulted in an increase in type II cell cytochrome P450-dependent ethoxyphenoxazone dealkylase activity but not in other alveolar type II cell or macrophage alkoxyphenoxazone dealkylase activities or in parameters that monitor viability and cell wall integrity.
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PMID:Cytochrome P450-dependent alkoxyphenoxazone dealkylase activity in rat alveolar type II cells: effect of pretreatment with beta-naphthoflavone. 247 71

Murine Kupffer cells (KCs), which constitute one of the largest populations of tissue macrophages, differ from most other cells of the myelomonocytic lineage in lacking the capacity for a respiratory burst. A collagenase perfusion technique followed by adherence to plastic at low temperature yielded pure cultures of KCs uniformly expressing receptors for Fc and C3bi, and containing virtually no morphologically detectable intracytoplasmic debris. Such KCs took up and oxidized glucose via the hexose monophosphate shunt about the same as peritoneal macrophages (PCs). Respiratory burst stimuli failed to enhance the hexose monophosphate shunt in KCs, probably because no H2O2 was produced. Detergent-permeabilized KCs generated no O2- in the presence of 1 mM NADPH, in striking contrast to all PC populations studied. Yet, KCs contained at least one component of the O2(-)-producing oxidase, cytochrome b559, in the same quantities as PCs and neutrophils. Cytochrome b559 was demonstrated by a novel double-reduction spectral technique that eliminated interference from hemoglobin and mitochondrial cytochromes. Consistent with the presence of the oxidase, KCs acquired normal respiratory burst capacity after prolonged incubation in vitro. The defect in triggering the respiratory burst in KCs was selective for the reduction of O2 by NADPH, in that reduction of O2 by endogenous arachidonate was readily demonstrate in response to zymosan. The percent of arachidonate released, the percent oxygenated, and the suppression of prostacyclin and leukotriene C production, as well as the pattern of LFA-1 expression, all resembled the pattern reported with PCs several days after exposure to bacteria. Indeed, exposure of PCs to low numbers of zymosan particles led gradually to complete suppression of respiratory burst capacity and refractoriness to its enhancement by rIFN-gamma, as evident in KCs both before and after their explanation. Thus, the modulation of oxidative metabolism that characterizes KCs probably arises from frequent endocytic encounters. This phenomenon may permit macrophages to act as scavengers without oxidative damage to bystander cells.
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PMID:Analysis of the nonfunctional respiratory burst in murine Kupffer cells. 312 23

After the rat preputial gland was treated with collagenase and trypsin, five bands of cells were isolated by centrifugation in Ficoll gradients. Homogenates of the heavier cells (Bands IV and V) which contained less lipids, were more active than the homogenates of the lighter cells (Bands I, II and III) in transforming [1,2-3H]-dehydroepiandrosterone ([1,2-3H]-DHA) into [3H]-androstenedione and [3H]-testosterone and the latter into [3H]-dihydrotestosterone (DHT). In the presence of NAD, NADH and NADPH-generating system, [1,2-3H]-DHA was transformed into [3H]-DHT in 50-60% yield by homogenates of cells in Bands IV and V. DHT levels in the preputial gland were measured by radioimmunoassay. The levels in female rats reduced by 77% from 3.14 +/- 0.27 to 0.72 +/- 0.10 pg/mg tissue after adrenalectomy, and by 45% to 1.71 +/- 0.10 pg/mg tissue after ovariectomy. In male rats, the level reduced by 15% from 4.58 +/- 0.55 to 3.88 +/- 0.62 pg/mg tissue after adrenalectomy and by 40% to 2.74 +/- 0.21 pg/mg tissue after orchidectomy. These results demonstrated the transformation of DHA into DHT in the preputial gland of the rat, and that the adrenal is an important source of precursor steroid (DHA) for DHT formation in the preputial gland.
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PMID:Transformation of dehydroepiandrosterone into dihydrotestosterone by isolated cells from rat preputial gland. 622 75

1. Low molecular weight fractions (mol. wt. 3500-10 000) prepared from cytosols of luteinized rat ovaries inhibited succinate-supported cholesterol side chain cleavage by intact ovarian mitochondria utilizing endogenous or exogenous sterol as substrate. 2. The low molecular weight fractions inhibited steroid secretion by collagenase-dispersed ovarian cells stimulated with lutropin or dibutyryl cyclic AMP. 3. Steroidogenesis by intact mitochondria incubated with NADPH was enhanced by the low molecular weight ovarian fraction, but cholesterol side chain cleavage carried out by sonicated mitochondria incubated with NADPH was unaffected. 4. Succinate-supported mitochondrial respiration was stimulated by the low molecular weight factor, apparently by uncoupling of oxidative phosphorylation. The uncoupling seems to be the mechanism by which steroid synthesis is inhibited. 5. The low molecular weight factor was heat-labile and not extracted by activated charcoal. Similar heat-labile material capable of inhibiting succinate-supported mitochondrial steroid synthesis was not found in low molecular weight fractions prepared from rat kidney, liver, spleen, brain, plasma and bovine corpus luteum. 6. Treatment of rats with cycloheximide 1 h before killing resulted in a reduction of inhibitory activity in ovarian low molecular weight cytosolic fractions. 7. We conclude that ovarian cytosols contain a low molecular weight factor, presumably a protein, which inhibits mitochondrial cholesterol side chain cleavage by uncoupling oxidative phosphorylation. The physiological function of this factor remains to be determined.
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PMID:Presence of a low molecular weight inhibitor of succinate-supported cholesterol side chain cleavage in rat ovaries. 624 96

We have previously shown that inositol-1,4,5-trisphosphate (IP3) releases Ca2+ from an intracellular calcium store in permeabilized acinar cells of rat pancreas (H. Streb et al., 1983, Nature (London) 306:67-69). This observation suggests that IP3 might provide the missing link between activation of the muscarinic receptor and Ca2+ release from intracellular stores during stimulation. In order to localize the intracellular IP3-sensitive calcium pool, IP3-induced Ca2+ release was measured in isolated subcellular fractions. A total homogenate was prepared from acinar cells which had been isolated by a collagenase digestion method. Endoplasmic reticulum was separated from mitochondria, zymogen granules and nuclei by differential centrifugation. Plasma membranes and endoplasmic reticulum were separated by centrifugation on a sucrose step gradient or by precipitation with high concentrations of MgCl2. IP3-induced Ca2+ release per mg protein in the total homogenate was the same as in leaky cells and was sufficiently stable to make short separation procedures possible. In fractions obtained by either differential centrifugation at 7000 X g, sucrose-density centrifugation, or MgCl2 precipitation there was a close correlation of Ip3-induced Ca2+ release with the endoplasmic reticulum markers ribonucleic acid (r = 0.96, 1.00, 0.91, respectively) and NADPH cytochrome c reductase (r = 0.63, 0.98, 0.90, respectively). In contrast, there was a clear negative correlation with the mitochondrial markers cytochrome c oxidase (r = -0.64) and glutamate dehydrogenase (r = -0.75) and with the plasma membrane markers (Na+ + K+)-ATPase (r = -0.81) and alkaline phosphatase (r = -0.77) in all fractions analyzed. IP3-induced Ca2+ release was distributed independently of zymogen granule or nuclei content of the fractions as assessed by electron microscopy. The data suggest that inositol-1,4,5-trisphosphate releases Ca2+ from endoplasmic reticulum in pancreatic acinar cells.
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PMID:Effect of inositol-1,4,5-trisphosphate on isolated subcellular fractions of rat pancreas. 633 62


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