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)

The hamster histone H3.2 promoter contains an AP-1-like element (referred to as site X) that contains the sequence CGAGTCA. This site differs from the Jun/AP-1 consensus sequence by one base and is also similar to the cyclic AMP response element. Similar AP-1/cyclic AMP response element-like sites have been found in the promoters of other histone H3 genes and are known to bind proteins either in vivo or in vitro. Using site directed mutagenesis, we demonstrate that a 10-base pair region which encompasses site X is a positive control element that is necessary for the S phase dependent increase in H3.2 transcription in cells synchronized by serum stimulation or aphidicolin block. DNase I footprint analysis shows that mutating site X eliminates v-Jun and hamster cellular factor(s) binding. Further in vitro analysis with gel retardation assays reveals that the flanking sequence of this site is necessary for the formation of an H3.2 specific complex that can be distinguished from complexes formed with a collagenase or SV40 AP-1 element. Antibodies specific to the different members of the Jun and Fos family of transcription factors show that, in gel retardation assays, a Jun-like factor is a component of the H3.2 specific complex. However, the H3.2 specific complex exhibits different reactivity toward the Jun and Fos specific antibodies as compared to complexes formed with a collagenase AP-1 element. We hypothesize that a unique protein complex, containing a component related to the AP-1 family of transcription factors, binds to the AP-1-like motif of the hamster H3.2 promoter and may be involved in the S phase dependent regulation of transcription.
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PMID:Identification of a 10-base pair protein binding site in the promoter of the hamster H3.2 gene required for the S phase dependent increase in transcription and its interaction with a Jun-like nuclear factor. 147 72

To determine whether specific hormonal responses were involved in the production of cryoprotectant (glucose) by liver of the freeze tolerant wood frog, Rana sylvatica, metabolically active hepatocytes were isolated in reasonable yields (mean 20.1 +/- 1.30% SEM, n = 29) by in situ liver perfusion with collagenase. Freshly isolated cells from autumn-collected frogs contained large amounts of glycogen (650 mumol glucosyl units/g packed cells) and produced glucose from this endogenous reserve at a rate of 10 mumol g-1 hr-1 at 0 degrees. Glucose output from cells was highly responsive to the addition of hormones; rates of glucose release increased 2.1-, 1.7-, and 1.7-fold with the addition of 10(-7) M bovine glucagon, 10(-7) M epinephrine, and 5 x 10(-6) M dibutyryl-cyclic AMP, respectively. Norepinephrine, 5-hydroxytryptamine, and bovine insulin were without effect at 0.1 microM/l. Hormone stimulation of glucose release was correlated with an increase in both the total activity and the percentage a of glycogen phosphorylase in hepatocytes. However, none of the hormones tested affected the kinetic properties of hepatocyte pyruvate kinase, suggesting the absence of covalent modification control of the enzyme. The data indicate that the freezing-stimulated production of large quantities of glucose as a cryoprotectant by R. sylvatica liver does not involve qualitative differences in the hormonal control of liver glycogenolysis, compared with other lower vertebrates. However, quantitative differences were seen, such as the much greater phosphorylase activity, 4.38 +/- 0.33 mumol min-1 g-1 packed cells, in freshly isolated R. sylvatica hepatocytes compared with 0.36 +/- 0.06 mumol min-1 g-1 in Rana pipiens hepatocytes.
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PMID:Hormonal effects on glycogen metabolism in isolated hepatocytes of a freeze-tolerant frog. 162 97

The effect of leukoregulin, a 50-kD lymphokine with unique antitumor properties, was studied in vitro on several fibroblast functions. Leukoregulin did not inhibit fibroblast proliferation, as measured by cell enumeration and [3H]thymidine incorporation, and had no cytotoxic effect in terms of increased membrane permeability detected by trypan blue exclusion, two of the major leukoregulin actions on tumor cells. Leukoregulin induced a dose-dependent decrease in collagen synthesis, demonstrated by decreased [3H]proline incorporation into collagenase-digestible protein, as early as 6 h after the addition of the lymphokine to human fibroblasts. Leukoregulin inhibited the synthesis of both type I and type III collagen, as measured by SDS-PAGE and by specific radioimmunoassay. Neutralizing antibodies to interleukin-1 alpha, interleukin-1 beta, tumor necrosis factor-alpha, and interferon-gamma failed to alter the effect of leukoregulin on collagen synthesis, attesting that leukoregulin action was not due to contamination by these cytokines. Inhibition of collagen synthesis occurred concomitantly with increased secretion of prostaglandin E2 and a transient rise in intracellular cyclic AMP content, peaking at 6 h. However, blocking prostaglandin synthesis with indomethacin did not counteract inhibition of collagen synthesis by leukoregulin, demonstrating independence of this action of leukoregulin from cyclooxygenase metabolites. Leukoregulin also stimulated glycosaminoglycan production in a dose-dependent manner, affecting the synthesis of hyaluronic acid as the major fibroblast-derived extracellular glycosaminoglycan. In addition, secretion of neutral proteases (collagenase, elastase, caseinase) was increased. These observations indicate that leukoregulin is able to regulate synthesis of molecules critical to the deposition of the extracellular matrix by nontransformed nonmalignant fibroblasts.
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PMID:Modulation of human dermal fibroblast extracellular matrix metabolism by the lymphokine leukoregulin. 164 5

The lapine synovial cell line HIG-82 secretes factors that activate cultures of articular chondrocytes. We showed that these "chondrocyte-activating factors" (CAF) also activate quiescent cultures of HIG-82 cells in an autocrine fashion. After exposure to partially purified preparations of CAF, HIG-82 cells increased their synthesis of prostaglandin E2 (PGE2) and the neutral proteinases collagenase, gelatinase, and stromelysin. CAF also induced their own synthesis. Both PGE2 synthesis and endogenous production of CAF started to increase between 1 and 3 h after treatment of cells with exogenous CAF, but the neutral proteolytic activity of the conditioned medium took approximately 12 h to increase. Induction of neutral proteinases by CAF was inversely related to the degree of cell confluency, whereas their induction by phorbol myristate acetate (PMA) was independent of this parameter. Both CAF and PMA provoked morphologic changes in subconfluent cultures of HIG-82 cells. Although the intracellular concentration of free Ca2+ increased rapidly in response to CAF, the results of experiments with calcium channel blockers and ionophores failed to support a role for Ca2+ fluxes in induction of neutral proteinases. In similar types of experiments, no evidence could be found to implicate fluxes in cyclic AMP or cyclic GMP in the induction of collagenase, gelatinase, or stromelysin. Because PMA is such a strong inducer of these enzymes, protein kinase C may be involved in signal transduction, but further work is needed to determine whether this is so.
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PMID:Studies on the autocrine activation of a synovial cell line. 165 86

Exposure of rat epididymal fat pad to phorbol 12-myristate 13-acetate (TPA), an activator of protein kinase C, results in an 85% increase in isoproterenol-stimulated cyclic AMP (cAMP) accumulation, an effect which was antagonized by H7, a protein kinase C inhibitor. This promoting action of TPA appears to be related to (i) an increase in the catalytic activity of adenylate cyclase, (ii) an increase in the maximal response of adenylate cyclase to fluoride and guanylimidodiphosphate (GppNHp) with no change in the EC50 value for GppNHp, and (iii) a reduction of the isoproterenol-stimulated low-Km cAMP phosphodiesterase activity present in the 30,000 g pellet of fat pad homogenates. In contrast with fat pads, exposure of isolated rat fat cells to TPA failed to influence their adenylate cyclase response to GppNHp and their cAMP accumulation and lipolysis. However, the other alterations caused by TPA in fat pads were still observed in fat cells. These results suggest that (i) the major alteration responsible for the promoted isoproterenol-stimulated cAMP response observed in fat pads after exposure to TPA is an increased interaction between the alpha s subunit of Gs and the catalytic site of adenylate cyclase and (ii) this increased interaction is dependent on protein kinase C activation and is abolished by collagenase digestion.
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PMID:Differential modulation of the adenylate cyclase/cyclic AMP stimulatory pathway by protein kinase C activation in rat adipose tissue and isolated fat cells. Influence of collagenase digestion. 165 98

A method involving centrifugal elutriation followed by density gradient centrifugation and incubation with a macrophage monoclonal antibody has been investigated to separate and characterize Leydig cells and macrophages from adult rat testes. After dispersion of the testes with collagenase, the isolated interstitial cells were found to contain 18% Leydig cells and 12% macrophages. These cells were then separated by centrifugal elutriation into eight fractions (F1-F8) (9 to 74 ml/min at 386 g). Each of these fractions was then further purified by density gradient centrifugation on 0-90% Percoll gradients. After centrifugal elutriation, the macrophages were mainly eluted in the first three fractions (F1-F3), whereas the Leydig cell percentage increased in each fraction with increasing flow rate. After further purification of each fraction on Percoll gradients, high percentages of macrophages (11-20%) were found in fractions F1-F3 (average density 1.045 g/ml), containing 11-37% Leydig cells. Less than 3% of the cells in fraction F4-F8 (average density 1.075 g/ml) were macrophages and more than 95% were Leydig cells. Heterogeneity of Leydig cells with respect to sedimentation velocities and function was found. Leydig cells from elutriated- and Percoll-purified fractions F4-F8 were heterogeneous with respect to testosterone and cyclic AMP (cAMP) production but showed a similar binding capacity for 125I-labelled human chorionic gonadotrophin. Leydig cells with the highest sedimentation velocity (35.7 mm/h.g) from fractions F7 and F8 were approximately twofold more responsive to LH (3.3 nmol/l) with respect to testosterone and cAMP production compared with Leydig cells with the lowest sedimentation velocity (20.7 mm/h.g).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Separation and characterization of Leydig cells and macrophages from rat testes. 165 79

We have established mutant SaOS-2 cell lines that express a cyclic AMP (cAMP)-resistant phenotype to investigate the regulation and functional importance of orthophosphoric-monoester phosphohydrolase alkaline optimum (ALPase) in the action of parathyroid hormone (PTH). Cells were stably transfected with a plasmid that directs the synthesis of a mutant form of the type I regulatory subunit of protein kinase A (PKA) under the control of the metallothionein promotor. There was no significant difference between parental SaOS-2 cells and the mutant lines in the affinity or number of receptors for 125I-Nle8,18Tyr34bPTH1-34NH2, either in the absence or presence of Zn2+. When cAMP-dependent gene transcription was examined using transient transfection with a somatostatin promoter-chloramphenicol acetyl transferase (CAT) reporter plasmid, CAT activity stimulated by human PTH and dibutyryl cAMP (DBcAMP) was inhibited by greater than 90% in the presence of Zn2+ in the mutant cell lines. In contrast, activation by a phorbol ester of a pentameric collagenase promoter/CAT construct containing five tandem copies of the AP-1 response element (5x-TRE-CAT) was unaffected in Zn(2+)-treated mutant cells. The inhibitory actions of PTH and DBcAMP on ALPase release were blunted by up to 80-90% in the mutant cell lines in the presence of Zn2+; there were no significant differences in the magnitude of inhibitory effects between these agonists. We conclude that the inhibitory action of PTH on ALPase release in SaOS-2 cells is mediated via activation of PKA. These cAMP-resistant cell lines will be especially useful in elucidating signal transduction mechanism(s) for PTH in human osteoblastic cells.
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PMID:Protein kinase A-dependent inhibition of alkaline phosphatase release by SaOS-2 human osteoblastic cells: studies in new mutant cell lines that express a cyclic AMP-resistant phenotype. 166 91

The present study examines the mechanism(s) of action of anethole trithione compared to the sialogogue pilocarpine. This was done by comparing the acute effects of these drugs on autonomic receptor binding (homogenates) together with parallel tests evaluating the biological activities of the receptor systems in collagenase-isolated rat parotid acini. The responses were measured as receptor-activated changes in cyclic nucleotide formation and acinar oxygen consumption. The results revealed that anethole trithione was unable to bind to muscarinic acetylcholine receptors and unable to stimulate the dynamic processes directly. It did, however, inhibit part (about 50%) of the carbachol-induced cyclic guanosine 3',5'-monophosphate (cyclic GMP) formation and O2 uptake. Furthermore, anethole trithione (greater than 1 microM) displaced [3H]prazosin (but not [3H]dihydroalprenolol ([3H]DHA] binding, without any effect upon the adrenaline-induced cyclic adenosine 3',5'-monophosphate (cyclic AMP) formation and O2 uptake. In conclusion, this study has shown that anethole trithione is not to be considered as a simple cholinergic agonist like pilocarpine, and further elucidation of the mechanism(s) of action of this agent would be useful.
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PMID:Acute effects of a possible sialogogue, anethole trithione, in rat parotid glands. 166 56

Human gastric mucosal cells were isolated by digestion of fundic biopsies with pronase and collagenase. The mean value of gastric cells per milligram biopsy specimen +/- SEM was 59,000 +/- 4,300 (n = 31) with a viability of 90 +/- 5%. With the cell yield of 1 patient a series of approximately 70-80 cyclic AMP measurements was possible. Histamine stimulated intracellular cyclic AMP production with an EC50 value of 35 +/- 25 mumol/l (SEM; n = 4). In the presence of 100 mumol/l histamine the Ki values (mumol/l) for the histamine H2 receptor antagonists averaged 1.45 (cimetidine), 0.10 (ranitidine), and 0.02 (famotidine). No significant inhibition of histamine-induced cyclic AMP production was obtained with the histamine H1 receptor antagonist triprolidine. With the new method histamine-induced cyclic AMP production can be measured in intact human gastric mucosal cells from fundic biopsy samples.
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PMID:A method to assess histamine-induced cyclic AMP production in isolated gastric mucosal cells from human biopsies. 168 55

When mechanical stress is applied, osteoblasts have shown to produce bone turnover stimulating hormones and enzymes like prostaglandin E2 (PGE2), cyclic AMP, alkaline phosphatase, and collagenase. Osteocalcin (bone Gla protein) is also a protein produced by osteoblasts to control bone metabolism. Thus, its production may also be stimulated by mechanical stress. The purpose of this investigation was to test if mechanical stress stimulates osteoblast-like cells to produce osteocalcin in vitro. The results suggest that osteocalcin production is stimulated at the initial stage of the culture by cyclic tension and relaxation force, and secretion may decrease with time.
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PMID:Mechanical stress as a stimulant to the production of osteocalcin in osteoblast-like cells. 181 33


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