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:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Specific binding sites for cholecystokinin (CCK) have been identified and characterized in fundic glands isolated by collagenase treatment from guinea pig gastric mucosa using a biologically active 125I-labeled derivative of the C-terminal octapeptide of CCK (125IIE-CCK-8). The time course of binding to these glands was rapid, temperature dependent and saturable. At 24, 30 and 37 degrees C, half-maximal binding was reached at 5 min and full binding at 30 min. The addition of a large excess of CCK-8 after 15 and 30 min of binding at 24 degrees C caused a prompt and rapid decline in radioligand bound showing that the interaction was reversible. There was a progressive decline in the amount of 125IIE-CCK-8 bound to fundic glands with increasing concentrations of CCK-8 and other structurally related peptides. Gastrin II displaced 50% of the radioligand at 1.6nM, CCK-8 at 3.2nM, gastrin I at 16nM, and desulfated-CCK-8 and pentagastrin at 59nM. Secretin did not displace the radioligand from fundic glands at 1.0uM. The binding was also tissue specific as glands isolated from the antral mucosa did not contain specific binding sites for 125IIE-CCK-8. This data provides evidence for specific receptors for CCK on gastric fundic glands that may be involved in the control of acid and pepsinogen secretion.
...
PMID:Identification and characterization of a specific receptor for cholecystokinin on isolated fundic glands from guinea pig gastric mucosa using a biologically active 125I-CCK-8 probe. 632 10

Human gallbladders were used to investigate the mechanisms of the impaired contraction induced by cholecystokinin (CCK) associated with cholesterol stones. Single muscle cells were isolated enzymatically with collagenase. Inositol 1,4,5-trisphosphate was measured by high-performance liquid chromatography. Diacylglycerol was assayed by thin-layer chromatography. CCK stimulation showed decreased muscle contraction and production of inositol 1,4,5-trisphosphate and diacylglycerol in gallbladders with cholesterol stones compared with those with pigment stones. Exogenous calmodulin induced maximal contraction of 22.4 +/- 0.5 and 21.0 +/- 0.6% in gallbladders with cholesterol and pigment stones, respectively. Similar findings were observed with a synthetic diacylglycerol analogue. Two G protein activators, aluminum fluoride and guanosine 5'-O-(3-thiotriphosphate), evoked similar responses in these two types of gallbladders, with maximal contractions of 21.3 +/- 0.4 and 23.3 +/- 0.5%, respectively, in those with cholesterol stones and 20.9 +/- 0.8 and 22.6 +/- 0.4%, respectively, in those with pigment stones. These results suggest that receptor-dependent ligands like CCK cannot fully activate the intracellular pathways, which, however, can be fully stimulated by circumventing receptors with G protein activators or second messengers. After G protein activation, the pathways appear to be functionally intact. The defect might then reside in the receptor or in the interaction between receptors and G proteins.
...
PMID:Direct G protein activation reverses impaired CCK signaling in human gallbladders with cholesterol stones. 749 56

Epidermal growth factor (EGF) regulates pancreatic acinar enzyme secretion. The mechanism of action of EGF in pancreatic acinar cells is not clear. In the present study we investigated the role of heterotrimeric GTP-binding proteins (G proteins) in EGF receptor signal transduction. Pancreatic acini were isolated from rat pancreas by collagenase digestion and permeabilized by digitonin. Activation of phosphatidylinositol 4,5-bisphosphate-specific phospholipase C (PLC) was assessed using a radioreceptor assay specific for inositol 1,4,5-trisphosphate [IP3(1,4,5)]. For measurement of amylase secretion isolated pancreatic acini were incubated with secretagogues for 30 min at 37 degrees C. Amylase released into the medium was assessed by monitoring the hydrolysis rate of p-nitrophenyl-alpha,D-maltohepatoside. The weakly hydrolyzable GTP analogue guanosine 5'-[3-O-thio]triphosphate (GTP gamma S) and guanosine 5'-diphosphate (GDP) were used to activate and inhibit G protein-mediated signal transduction, respectively. EGF (90 nM) stimulated amylase release in isolated pancreatic acini. This effect was enhanced by guanosine 5'-[3-O-thio]triphosphate (0.1 mM), which stimulates G proteins. Guanosine 5'-diphosphate (1 mM), which inhibits the activity of heterotrimeric G proteins, had no effect on basal and EGF-induced amylase release. Lower EGF concentrations (20 nM) inhibited COOH-terminal cholecystokinin octapeptide (CCK8)-induced IP3(1,4,5) production and amylase release in pancreatic acini). However, in the presence of GDP, EGF had no significant effect on CCK8-stimulated amylase release. Furthermore, coincubation of the acini with CCK8, EGF, and GDP revealed that GDP reduces the inhibitory effect of EGF on CCK8-induced IP3(1,4,5) production.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Epidermal growth factor receptor signaling in rat pancreatic acinar cells. 754 69

Rat pancreatic acini were prepared with collagenase A and collagenase P and their secretory response to carbachol tested. The collagenase P gave the best acinar suspension, providing that the digestion was performed in the absence of added calcium and in the presence of a low concentration of albumin. More than ninety percent of the acini prepared with this crude collagenase were viable as judged by a coloration with eosine. Their basal amylase secretion was below 3% amylase released within 20 minutes. They responded to cholecystokinin (7-fold stimulation, EC50: 10 pM), to bombesin (7-fold stimulation, EC50: 1 nM), to carbachol (7-fold stimulation, EC50: 1 microM), to fluoroaluminate (4-fold stimulation, EC50: 3 mM) or to TPA (4-fold stimulation, EC50: 100 nM). Acini preloaded with fura2 and incubated in the presence of 0.5 mM extracellular calcium were able to maintain a large (5,000-fold) gradient of calcium across their plasma membrane. Carbachol, CCK-8 and bombesin increased the intracellular calcium concentration 6-fold, within 10 seconds. This level decreased for the next 20 seconds but remained higher than the basal value for the next 10 minutes. These acini could be permeabilized with a low concentration (0.1 U/ml) of streptolysin O without affecting the basal amylase secretion, an index of the integrity of zymogen granules. It is concluded that pancreatic acini which have retained their responsiveness to major pancreatic secretagogues can be prepared with crude collagenase and permeabilized with streptolysin O. They can thus be used as a model for the study of stimulus-secretion coupling in exocrine glands.
...
PMID:Isolation of rat pancreatic acini with crude collagenase and permeabilization of these acini with streptolysin O. 767 14

The binding of fluorescence-labelled lectins and a fluorescence-marked hormone to the cell surface of isolated rat pancreatic acinar cells was studied by light microscopy and flow cytometric measurements. The pancreatic acinar cells were prepared by collagenase digestion. The fluorescence of cells was studied after binding of FITC-labelled WGA or UEA I as well as of FITC-marked pancreocymin/cholecystokinin (CCK-FITC) in a fluorescence microscope or FACScan. The strong binding of lectins was inhibited by preabsorption with the specific sugars. In comparison to the lectins the binding of CCK-FITC was low. There were two populations of acinar cells with different CCK-FITC binding capacity as detected by flow cytometry. The CCK-FITC cell surface fluorescence was significantly decreased by preincubation with unmarked hormone as well as with the non-labelled lectins. The inhibition of CCK-FITC binding by lectins is discussed in respect to a possible competition of the lectins and CCK for the CCK receptor.
...
PMID:Fluorescence microscopic studies and flow cytometric measurements of lectin and hormone binding to isolated rat pancreatic acinar cells. 813 89

Lectin binding to the glycocalyx of isolated rat pancreatic acinar cells was studied by flowcytometric measurements. The pancreatic exocrine cells were prepared after collagenase digestion and cleaned in a density gradient. The fluorescence of cells was measured in a FACScan after binding of FITC marked WGA or UEA I. The binding of lectins was inhibited by preabsorption of WGA-FITC with N-acetyl-glucosamine, sialic acid or chitinous and by preabsorption of UEA-FITC with alpha-L-fucose, respectively. Furthermore, we were able to measure a decreased WGA-FITC and UEA-FITC binding after a short preincubation of isolated cells with the peptide hormone cholecystokinin and its agonists (caerulein, pentagastrin).
...
PMID:Lectin binding studies with FITC-marked WGA and UEA I and flowcytometric measurements on isolated rat pancreatic acinar cells. 827 34

The cause and effect relationship between membrane cholesterol and gallbladder muscle contractility was examined by altering membrane cholesterol to phospholipid mole ratio using cholesterol-rich or cholesterol-free liposomes. Gallbladder single muscle cells, from prairie dogs that were fed either a regular or high-cholesterol (1.2%) diet, were isolated enzymatically with collagenase. Plasma membranes of gallbladder muscle were purified in sucrose gradient. Cholesterol was measured using the cholesterol oxidase method. Phospholipids were measured with the method of G.R. Bartlett (J. Biol. Chem. 234: 466-468, 1959). The results of this experiment are 1) after high-cholesterol feeding, cholesterol contents and cholesterol/ phospholipid mole ratio in plasma membranes of gallbladder muscle increased 90%, and muscle cell contraction in response to cholecystokinin octapeptide decreased 58%; 2) similar changes were observed when normal gallbladder muscle cells were incubated with cholesterol-rich liposomes for 2 h; and 3) the changes induced either in vivo or in vitro were reversed when muscle cells were subsequently incubated with cholesterol-free liposomes for 2-6 h. We conclude that gallbladder muscle may incorporate excess cholesterol into its plasma membrane when exposed to a cholesterol-rich environment, that excess membrane cholesterol impairs muscle contractility, and that these changes appear to be reversible.
...
PMID:Membrane cholesterol alters gallbladder muscle contractility in prairie dogs. 876 Jan 7

The purpose of this study was to characterize time-dependent changes in pepsinogen (PG) synthesis of porcine gastric chief cells during long-term monolayer culture. Porcine chief cells were isolated by pronase/collagenase treatment of fundic mucosa and enriched by density gradient and counterflow centrifugation. PG isoenzymes were identified in [L-35S]methionine-labelled cultured chief cells by native polyacrylamide gel electrophoresis followed by phosphor imager analysis, protease detection and immunoblots with specific PG A and C antibodies. The obtained results suggest that porcine chief cell cultures, after an initial settling period, reached an approximate steady state in total protein content and synthesis as well as in PG content and isoenzyme pattern from days 3 to 9 of culture. The latter was characterized by the presence of at least two PG A and two PG C isoenzymes. During the supposed steady-state total PG synthesis averaged out at 34 +/- 2% of total protein synthesis, as detected by [L-35S]methionine incorporation, due to the synthesis of, mainly, PG A2 and, to a much lesser extent, PG C and A1. In line with an active secretion, PG A2 proportion was on average significantly higher in released (44 +/- 3%) than in intracellular labelled proteins (19 +/- 2%). In addition, PG release from chief cells cultured for 6 and 9 days could be stimulated by cholecystokinin-octapeptide. These data suggest that porcine chief cells in monolayer culture are a model well suited for the quantitative and qualitative characterization of PG isoenzyme synthesis and release during long-term investigations, for which an establishment of a culture steady state appears to be a useful prerequisite.
...
PMID:Pepsinogen synthesis during long-term culture of porcine chief cells. 939 83

This study was designed to elucidate the mechanism of action of progesterone on gallbladder smooth muscle in guinea pigs. Adult male guinea pigs were treated with either progesterone (2 mg.kg-1.day-1) or saline for 7 days. Gallbladder muscle cells were isolated by enzymatic digestion with collagenase. Contractile responses to agonists were expressed as percent shortening from control cell length. [35S]guanosine 5'-O-(3-thiotriphosphate) ([35S]GTP gamma S)-binding properties of G proteins were assessed in crude membranes of gallbladder muscle with or without cholecystokinin octapeptide (CCK-8) stimulation. Gallbladder muscle cells from progesterone-treated guinea pigs exhibited an impaired contractile response to CCK-8, GTP gamma S, or aluminum fluoride but a normal response to potassium chloride or D-myo-inositol 1,4,5-trisphosphate compared with controls. Western blot analysis of gallbladder muscle revealed the presence of Gi1-2, Gi3, Gq/11, and Gs proteins. The maximal contraction induced by CCK-8 was blocked by pertussis toxin and Gi alpha 3-specific antibodies, but not by Gi alpha 1-2 or Gq/11 alpha antibodies. CCK-8 caused a significant increase in [35S]GTP gamma S binding to Gi alpha 3, but not to Gq/11 alpha or Gi alpha 1-2. The stimulation of Gi alpha 3 binding, however, was significantly reduced in gallbladder muscle membranes from progesterone-treated guinea pigs compared with that in control animals. In conclusion, progesterone might cause gallbladder hypomotility by downregulating Gi3 proteins.
...
PMID:Impaired G protein function in gallbladder muscle from progesterone-treated guinea pigs. 948 81

The effects of vanadate were examined by monitoring intracellular free calcium concentration ([Ca2+]i) and amylase secretion in collagenase-dispersed rat pancreatic acinar cells. Vanadate increased [Ca2+]i by mobilizing calcium from agonist-releasable intracellular calcium stores, since this increase was observed in the absence of extracellular calcium and vanadate failed to increase [Ca2+]i after treatment with thapsigargin in calcium-free medium. Moreover, pretreatment of acinar cells with vanadate prevented the cholecystokinin octapeptide (CCK-8)-induced signal of [Ca2+]i, whereas co-incubation with CCK-8 potentiated the plateau phase of calcium response to CCK-8 without modifying the transient calcium spike. The effects of vanadate on calcium mobilization were reversed by the presence of the sulfhydryl reducing agent dithiothreitol. Vanadate also activated the calcium influx, since an additional enhancement of calcium influx induced by thapsigargin-evoked intracellular store depletion was observed and vanadate reversed the inhibitory effect of lanthanum (an inhibitor of calcium entry) into acinar cells. In addition, vanadate evoked a concentration-dependent release of amylase from pancreatic acinar cells and moreover, reduced the secretory response to CCK-8. We conclude that, in pancreatic acinar cells, vanadate releases calcium from the agonist-releasable intracellular calcium pool and consequently induces amylase secretion. These effects are likely due to the oxidizing effects of this compound.
...
PMID:Oxidizing effects of vanadate on calcium mobilization and amylase release in rat pancreatic acinar cells. 1040 21


<< Previous 1 2 3 4 Next >>

---