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 Wistar fat-storing cells were isolated by perfusion with collagenase and centrifugation with metrizamide density cushion technique and cultured in vitro. The fat-storing cells were confirmed by the presentation of the lipid drop in the cytoplasm and the visualization of dismin with anti-dismin antibody by using indirect immunofluorescence method. By the videotape recorder (VIR) and the imagine analysis system, we observed the wandering immigration, the abrupt contraction of fat-storing cells with spike, then becoming a ball-like shape in its division phase. After that the cell began to extend and the contraction of these cells can be induced by the presence of 10(-2) mmol/L endothelin-1, 1 mmol/L of substance P and 2 x 10(-5) mmol/L noradrenalin. After removal of these agents the contracted cells would become extended. All these findings indicate that the fat-storing cells have ability of contraction and movement.
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PMID:Observations on the contraction and movement of fat-storing cells in culture. 752 80

The interaction between endothelial cells and immune/inflammatory cells plays an important role in the pathogenesis of vascular diseases. Inflammatory cells also activate endothelial cells and release both proliferative and cytotoxic mediators. In order to examine the interaction between leukocytes and endothelial cells and the effect of various drugs, we established the methodology for isolating and culturing the endothelial cells from human umbilical vein. Endothelial cells were harvested by using 0.1% collagenase within 48 hr of collecting the cord. Cells were grown to confluency in 96-well plates in Medium 199 containing 20% fetal calf serum, endothelial cell growth supplement, heparin, and antibiotics. Using this method, we obtained a confluent layer of the cells in all the 96 wells within 48 hr. We then examined the effect of peptides, endothelin-1, substance P, and neurokinin-A on the adherence of human blood neutrophils (purity and viability > 98%) to endothelial monolayers. All the peptides enhanced (p < 0.05) the adherence of neutrophils to endothelial cells in a time-dependent manner. This method of endothelial cell culturing is reliable, reproducible, and effective in evaluating the role of various mediators and drugs on the adherence of various white blood cells to endothelium.
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PMID:Technique for assessment of leukocyte adherence to human umbilical vein endothelial cell monolayers. 753 69

The contractile response of cultured Ito cells to endothelin-1 and substance P was examined. Ito cells were obtained from rat liver by perfusion with collagenase, followed by separation through centrifugal elutriation, and were cultured for 24 hr. The area of the Ito cells was measured after treatment with endothelin-1 or substance P at various concentrations in the culture medium. The area of the cells decreased dose dependently after treatment with endothelin-1 or substance P. The area of Ito cells before addition of interleukin-1 or substance P was defined as 100%. The area of the cells after treatment with endothelin-1 or substance P medium was expressed as the percentage against the area before treatment with endothelin-1 or substance P. The percentage in area after treatment with 200 nmol/L endothelin-1 was as follows: 81% +/- 13% at 30 min, 77% +/- 15% at 60 min, 87% +/- 15% at 120 min and 99% +/- 18% at 180 min. The maximal decrease in area occurred at 60 min after treatment. The percentage values for 200 nmol/L substance P were as follows: 88% +/- 15% at 10 min, 95% +/- 17% at 30 min and 101% +/- 17% at 60 min. The maximal decrease in area was noted at 10 min. Thus Ito cells contracted in response to treatment with endothelin-1 or substance P. The mode of the extent and onset of the contraction was different for the two peptides. These findings suggest that Ito cells are involved in the regulation of the hepatic sinusoidal microcirculation.
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PMID:Ito cell contraction in response to endothelin-1 and substance P. 769 8

The effects of endothelin-1 (ET-1) on protein synthesis and phosphoinositide (PI) hydrolysis were investigated in ventricular myocytes isolated by collagenase digestion of adult rat hearts. The maximum stimulation of protein synthesis by ET-1 was about 35% and the EC50 value was about 0.3 nM. The stimulation was exerted at the translational stage since it was insensitive to inhibition by actinomycin D. The maximum stimulation of PI hydrolysis by ET-1 as measured by the formation of [3H]inositol phosphates was about 11-fold and the EC50 value was about 0.7 nM. The ET-1 analogue sarafotoxin-6b stimulated protein synthesis by a maximum of 27% and stimulated PI hydrolysis about 8- to 9-fold. The EC50 values were 1.6 nM and 0.6 nM, respectively. Other endothelins stimulated protein synthesis and PI hydrolysis in the following order of potency: ET-1 approximately ET-2 > ET-3. This order of potency suggests that the stimulation of both protein synthesis and PI hydrolysis is mediated through the ETA receptor. Although both angiotensin II and [Arg]vasopressin stimulated PI hydrolysis significantly, the stimulation was less than 60%, i.e., much less than the stimulation by ET-1 and its analogues. Neither insulin nor substance P stimulated PI hydrolysis. Stimulation of protein synthesis by ET-1 and its analogues correlated strongly with the stimulation of PI hydrolysis and we suggest that the stimulation of protein synthesis may be dependent on the stimulation of PI hydrolysis. We hypothesize that the mechanism may involve a protein kinase C-mediated increase in intracellular pH.
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PMID:Stimulation of adult rat ventricular myocyte protein synthesis and phosphoinositide hydrolysis by the endothelins. 838 85

To examine the effects of endothelin-1(ET-1) on bile canalicular contractions, rat hepatocytes were isolated by a collagenase perfusion technique and cultured on p-n-p-vinylbenzenyl-D-lactonamide-coated dishes under serum-free conditions. The frequency of bile canalicular contractions was expressed by the percentage of the number of bile canaliculi(BC) that contracted compared to the total number of BC observed for 10 min. The treatment of the hepatocytes with ET-1 increased the frequency of contraction of the BC in a dose-dependent manner up to 2nM, control: 10.4%; 0.5nM ET-1:31.6%; 1nM ET-1:54.0%; 2nM ET-1:58.0%; 4nM ET-1:58.9%. The contraction, once it occurred, lasted for rest of the observation period. We also observed a transient increase in the concentration of intracellular Ca in cultured hepatocytes upon the addition of ET-1 to the medium. These results suggest that ET-1 plays an important role in the mortility of BC in vivo.
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PMID:Endothelin-1 induces contraction of bile canaliculi in isolated rat hepatocytes. 846 19

The purpose of this study was to compare coronary and interstitial endothelin-1 (ET-1) levels in perfused rat hearts under several experimental conditions, because the cardiac tissue concentration of ET-1 is not clear. Hearts were perfused in an upside-down position with a colloid-free buffer at a constant flow rate of 9 ml/min/g heart wet weight, and immunoreactive ET-1 was determined in timed collections of coronary effluent and interstitial transudate produced by the ventricles and appearing on their surface. Basal ET-1 release into effluent was 0.26 +/- 0.007 pg/min/g, and 0.005 +/- 0.0012 pg/min/g in transudate. Basal ET-1 concentration was 0.11 +/- 0.005 pg/ml (transudate) and 0.03 +/- 0.002 pg/ml (effluent), indicating four-fold higher transudate than effluent levels (P < 0.05). Following perfusion of hearts with collagenase to remove endothelial cells, ET-1 release into effluent was reduced to one-third and completely abolished in transudate, indicating that the peptide originated from the vascular endothelium. Perfusion of hearts with angiotensin II (0.1 mumol/l) or thrombin (5 U/ml) increased coronary perfusion pressure and ET-1 secretion, but little affected the transudate/effluent ET-1 concentration ratio (5.5 and 3.2, respectively). When coronary flow was reduced to ischaemic level (1 ml/min/g over several hours), ET-1 secretion rates into effluent were decreased by 55-65%, but increased three- to four-fold on reperfusion at normal flow (P < 0.05). The ET-1 concentrations in both fluids were still always below 1 pg/ml. No change in coronary perfusion pressure compared to time-matched normoxic controls was observed. In the presence of the ET-1 converting enzyme inhibitor, phosphoramidon (1.7 mumol/l), ischaemia-induced increases of ET-1 secretion were attenuated, and this was accompanied by a time-dependent rise in coronary perfusion pressure up to 60% (P < 0.05). These are the first measurements of endogenous cardiac tissue ET-1 levels; they do not support a vasoconstrictor (pro-ischaemic) action of endogenous ET-1 in rat hearts following ischaemia/reperfusion, but rather point to a possible vasodilator role of the peptide under these conditions.
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PMID:Tissue endothelin-1 levels in perfused rat heart following stimulation with agonists and in ischaemia and reperfusion. 852 55

Angiotensin II (Ang II) has growth-stimulatory properties on different renal cell types. However, possible growth effects of this vasoactive peptide on endothelial cells isolated from the glomerular microvasculature have not been formally investigated. Therefore, we isolated and characterized primary cultures of rat glomerular endothelial cells. We used a simple technique in which collagenase-treated glomeruli were sparsely plated in several 96-well culture plates and microscopically screened for cobblestone-like outgrowth. After two limiting dilutions, homogeneous cultures were obtained. Cells were characterized by positive staining for the endothelial markers factor VIII, CD 31, endothelial leukocyte adhesion molecule-1, and the lectin Bandeiraea simplificifolia. Ang II stimulated the synthesis and release of endothelin-1 in culture supernatants. Moreover, in contrast to syngeneic mesangial cells, glomerular endothelial cells expressed angiotensin-converting enzyme. Ang II stimulated a mild but significant proliferation of quiescent cells, as measured by [3H]thymidine incorporation and direct cell counting. This mitogenesis was transduced by losartan-blockade angiotensin type 1 receptors. Moreover, Ang II mediated phosphorylation of mitogen-activated protein kinase 2 and induction of transcripts for the immediate early gene Egr-1. Our results indicate that Ang II is a moderate mitogen for primary cultures of rat glomerular endothelial cells and activation of these metabolically active cells may play a role in the pathophysiology of several types of glomerulonephritis. Moreover, remodeling of glomerular endothelial cells by Ang II may be important in the progression of structural renal damage during the course of hypertensive injury.
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PMID:Angiotensin II is mitogenic for cultured rat glomerular endothelial cells. 861 66

1. Endothelin-1 is a 21 amino acid peptide with potent inotropic and chronotropic actions in the heart. Relatively little is known about the underlying electrophysiological effects of the peptide. In this study, the effects of endothelin-1 (ET-1) on the acetylcholine-activated potassium current (IK(ACh) were investigated in the absence and presence of the receptor-selective antagonists, PD155080 (ETA receptor-selective) and RES-701 (ETB receptor-selective) in rabbit atrial cardiomyocytes. 2. Cells were obtained from New Zealand White rabbits (2.5-3 kg) by enzymatic dissociation with collagenase. Potassium currents were recorded, in the presence of nifedipine (5 microM), by use of the whole cell ruptured patch-clamp technique. Following stabilization, control recordings were made with standard pulse protocols, and drugs were applied by a gravity fed microperfusion system. 3. Endothelin-1 (10 nM) alone did not affect the "steady state' potassium current. Acetylcholine (1 microM) increased (P < 0.05) the potassium current to-1321 +/- 290 pA, from a control value of -955 +/- 191 pA, at a step potential of -100 mV. Acetylcholine also increased the holding current at -40 mV from +80 +/- 9 pA to +242 +/- 38 pA, and this effect was abolished (P < 0.05) in the presence of endothelin-1 (+44 +/- 13 pA). The responses to acetylcholine were attributed to activation of the atrial muscarinic-activated potassium current (IK(ACh)) as they were blocked by atropine (10 microM). Endothelin-1 (10 nM) in the presence of acetylcholine did not affect the "steady state' potassium current (-882 +/- 88 pA compared to a control value of -870 +/- 98 pA, at -100 mV). 4. The ETA receptor-selective antagonist, PD155080 (1 microM), prevented (P < 0.05) the ET-1 induced inhibition of IK(ACh) at all potentials. PD155080, in the presence of endothelin-1 and acetylcholine, increased the inward component of the "steady state' potassium current to -1030 +/- 210 pA from a control value of -804 +/- 224 pA at a step potential of -100 mV. Also the outward component was increased at a potential of -20 mV from +90 +/- 17 pA to +241 +/- 47 pA. 5. Unlike PD155080, the ETB receptor-selective antagonist, RES-701 (1 microM), only prevented (P < 0.05) the inhibitory effect of endothelin-1 on the inward component of the IK(ACh); at -100 mV, RES-701, in the presence of endothelin-1 and acetylcholine, increased the "steady state' potassium current to -913 +/- 137 pA from -733 +/- 116 pA. Furthermore, RES-701, in contrast to PD155080, failed to sustain this inhibitory effect as, in the presence of endothelin-1 and acetylcholine, the "steady state' potassium current returned to a value of -768 +/- 96 pA, at a step potential of -100 mV. 6. In conclusion, endothelin-1 clearly inhibits the effects of acetylcholine on IK(ACh) in rabbit atrial cardiomyocytes. This effect is primarily mediated by an ETA receptor-subtype, but is transiently and partially mediated by a RES-701-sensitive ETB receptor subtype. Inhibition of the IK(ACh) may account for the positive chronotropic properties of endothelin-1.
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PMID:Endothelin-1 mediated inhibition of the acetylcholine-activated potassium current from rabbit isolated atrial cardiomyocytes. 896 52

Progressive interstitial fibrosis accompanied by loss of renal tubules and interstitial capillaries typifies all progressive renal diseases. Dynamic and complex, the process evidently overlaps with matrix remodeling; it may even be reversible. The interstitial fibrous tissue comprises several normal and novel matrix proteins, proteoglycans, and glycoproteins. Interstitial myofibroblasts are a major site of matrix protein overproduction, although resident fibroblasts, tubular cells, and inflammatory cells may contribute. Inadequate matrix degradation also appears to contribute to the fibrogenic process. Two protease cascades, the metalloproteinases and the plasminogen activator/ plasmin family of serine proteases, are implicated in the turnover of interstitial matrix proteins; upregulated expression of protease inhibitors has been observed in each. Increased tissue inhibitor of metalloproteinase-1 and plasminogen activator inhibitor-1 levels suggest that the intrinsic renal activity of the metalloproteinases and serine proteases are inhibited while matrix proteins accumulate in the interstitium. Several signals that may direct the interstitial fibrogenic process have been identified, but not yet proved to cause it. Upregulated expression of transforming growth factor beta-1, the proteotypic fibrogenic cytokine, has been observed in experimental and human models; it probably does not act alone. There may be supportive roles for platelet-derived growth factor, interleukin-1, basic fibroblast growth factor, angiotensin II, and endothelin-1. Although it is not known why interstitial fibrosis compromises renal function, atrophy of renal tubules may be pivotal. Ischemic necrosis and/or apoptosis may generate nonfunctioning atubular and sclerotic glomeruli. Future studies must delineate the molecular basis of the differences between renal repair and renal destruction by fibrosis, two processes that share many common features.
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PMID:Molecular insights into renal interstitial fibrosis. 898 27

Ets-1 is a transcription factor that activates expression of matrix-degrading proteinases such as collagenase and stromelysin. To study the control of ets-1 gene expression in rat vascular smooth muscle cells (VSMC), cells were exposed to factors known to regulate VSMC migration and proliferation. Platelet-derived growth factor-BB (PDGF-BB), endothelin-1 (ET-1), and phorbol 12-myristate 13-acetate (PMA) induced a dose-dependent expression of ets-1 mRNA. These effects were abrogated by inhibition of protein kinase C (PKC) by H-7 or chronic PMA treatment. Ets-1 mRNA was superinduced by PDGF-BB and ET-1 in the presence of cycloheximide. The chelation of intracellular Ca2+ by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester and the depletion of endoplasmic reticulum intracellular Ca2+ concentration ([Ca2+]i) by thapsigargin inhibited PDGF-BB- and ET-1-induced ets-1 mRNA, whereas ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid had no effect. However, [Ca2+]i release alone was not sufficient to increase ets-1 mRNA. Forskolin blocked ET-1-, PDGF-BB-, and PMA-induced ets-1 mRNA, as well as inositol phosphate formation, consistent with an effect through impairment of PKC activation. Inhibitors of ets-1 gene expression, such as H-7 and herbimycin A, inhibited the ET-1 induction of collagenase I mRNA. We propose that ets-1 may be an important element in the orchestration of matrix proteinase expression and of vascular remodeling after arterial injury.
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PMID:Ets-1 is an early response gene activated by ET-1 and PDGF-BB in vascular smooth muscle cells. 948 38


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