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)

Experiments were performed to indentify the series elastic component (SEC) in intact dog carotid artery held at in situ length. The vessels were studied during excitation of the muscle with norepinephrine and after metabolic poisoning with potassium cyanide and sodium iodoacetate. Static circumferential stress-strain curves and stress-quick-release stiffness curves were examined to evaluate Maxwell and Voigt model elements. The vessels were studied at 33, 36, and 39 degrees C. Temperature variations altered active stress, but did not alter connective tissue properties or the Maxwell SEC stiffness. The Voigt model SEC stiffness was altered, but this was secondary to changes in active stress. Thus, most of the SEC is separate from the contractile apparatus. Other vessels were treated with elastase, collagenase, or hyaluronidase to digest the connective tissue components of the wall. Hyaluronidase had no effect on mechanics. Elastase and collagenase altered connective tissue properties, but only elastase unequivocally altered SEC stiffness. This analysis indicated 1) that the carotid artery wall is better represented by a Maxwell model than a Voigt model, and 2) that the SEC in intact carotid artery is primarily elastin.
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PMID:Identification of smooth muscle series elastic component in intact carotid artery. 19 Aug 98

Isolation of blood and intracellular forms of Trypanosoma cruzi was made mainly from rats (90-110 g) which had received 580 rad of whole-body gamma-irradiation not more than 24 h before subcutaneous inoculation with 10(7) trypomastigotes of the Sonya strain of T. cruzi. Unirradiated chinchillas (250-350 g) were, however, used for some experiments. Blood forms were isolated using a technique involving differential centrifugation to remove most of the erythrocytes and DEAE-cellulose chromatography to remove the remaining blood cells. Overall recoveries were usually in the range 30-70%. Parasites were mainly (approximately 98%) broad forms and were motile, metabolically active (as judged by respiratory and radio-tracer incorporation studies) and had lost none of their infectivity for mice. Intracellular forms were isolated from hind-limb muscle tissue. This was disrupted in an MSE tissue homogenizer and the homogenate incubated with DNase, collagenase and trypsin. Parasites, contaminated only by a few blood cells, were then obtained by differential centrifugation. For purer preparations, a terminal sucrose gradient step was used. Recoveries ranged between 40 and 70%. About 1-3% of the parasites isolated were epimastigotes and trypomastigotes; the remainder are probably best collectively termed 'amastigotes', though they were pointed and most had a short, free flagellum. They were undamaged as judged by light and electron microscopy and metabolically active as judged by respiratory and radio-tracer incorporation studies. However, the infectivity for mice of both these purified preparations and the initial cell homogenates could be accounted for by the epimastigotes and trypomastigotes present in them. Preliminary biochemical studies with isolated parasites have shown that blood, intracellular and culture forms of T. cruzi have a respiratory system which is in part sensitive to CN- and that all forms synthesize nucleic acids and proteins when incubated in vitro. There appears, however, to be a lack of DNA synthesis in blood stages, and thus it is not surprising that these forms do not divide.
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PMID:Isolation of blood and intracellular forms of Trypansoma cruzi from rats and other rodents and preliminary studies of their metabolism. 20 67

The concentrations of clindamycin were significantly higher in iris and choroid-retina or pigmented rabbits than in those of albino rabbits after subconjunctival injection. Equilibrium dialysis experiments showed no affinity of clindamycin for synthetic melanin or for collagenase digests of pigmented tissues. In contrast, strips of iris and choroid-retina took up clindamycin rapidly from solution, achieving concentrations substantially higher than those in the medium. Uptake by tissue strips was not influenced by temperature (4 C vs. 37 C), cyanide, or ouabain. However, N-ethylmaleimide, which reacts with sulfhydryl groups, decreased the tissue-to-medium ratios by about 50% for both albino and pigmented choroid-retina. Even in the presence of this inhibitor, the ratio for pigmented tissues remained higher than that for albino specimens. These findings suggest the existence of one, or possibly two, mechanisms of energy-independent accumulation of clindamycin by pigmented ocular tissues: one may relate to protein sulfhydryl bonds that are present in both breeds; the other may involve the pigmentation apparatus.
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PMID:Marked differences between pigmented and albino rabbits in the concentration of clindamycin in iris and choroid-retina. 43 32

Rat hepatocytes were freshly prepared from adult animals using the collagenase-perfusion technique. The hepatic transport of thiamine was studied in isolated liver cells. The process was found to be saturable with an apparent Kt of 0.31 mM and a V max of 0.7 mumoles/ml intracellular fluid/5 minutes. However, at higher substrate concentrations, the process proceeded in a linear fashion. Both pyrithiamine and oxythiamine were inhibitory on the hepatic uptake of thiamine, the latter showed much weaker activity than the former. The system required the presence of sodium ions and was sensitive to ouabain. Anaerobic condition and metabolic inhibitors, e.g., 2,4-dinitrophenol, cyanide, and iodoacetate suppressed the uptake rate of thiamine. Addition of ethanol in the incubation medium also caused significant reduction of thiamine uptake. Efflux studies indicated that a portion of intracellular thiamine is readily available for exodus. Chromatographic analyses showed that thiamine was only slightly metabolically altered during the transport process. It is suggested that thiamine is transported into isolated hepatic cells by an active, sodium-dependent process.
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PMID:Active transport of thiamine by freshly isolated rat hepatocytes. 71 32

To determine the functioning rate of Na-K-ATPase in the rat nephron, a micromethod was developed to measure the rate of rubidium uptake in single nephron segments microdissected from collagenase-treated kidneys. Because the hydrolytic activity of Na-K-ATPase displayed the same apparent affinity for K and Rb ions, whereas the Vmax elicited by K was higher than that in the presence of Rb, experiments were performed in the presence of cold Rb plus 86Rb. Before the assay, tubules were preincubated for 10 min at 37 degrees C to restore the normal transmembrane cation gradients. 86Rb uptake was measured after washing out extracellular cations by rinsing the tubules in ice-cold choline chloride solution containing Ba2+. Rb uptake increased quasi-linearly as a function of incubation time up to 30 s in the thick ascending limb, 1 min in the proximal convoluted tubule, and 5 min in the collecting tubule, and reached an equilibrium after 5-30 min. The initial rates of Rb uptake increased in a saturable fashion as Rb concentration in the medium rose from 0.25 to 5 mM. In medullary thick ascending limb, the initial rate of Rb uptake was inhibited by greater than 90% by 2.5 mM ouabain and by 10(-5) M of the metabolic inhibitor carbonyl cyanide trifluoromethoxyphenylhydrazone. Correlation of Na-K-ATPase hydrolytic activity at Vmax and initial rates of ouabain-sensitive Rb uptake in the successive segments of nephron indicates that in intact cells the pump works at approximately 20-30% of its Vmax. Increasing intracellular Na concentration by tubule preincubation in a Rb- and K-free medium increased the initial rates of Rb intake up to the Vmax of the hydrolytic activity of the pump.
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PMID:Measurement of Na-K-ATPase-mediated rubidium influx in single segments of rat nephron. 216 56

Effects of intracellularly perfused ATP, and extracellularly applied cyanide and 2-deoxy-D-glucose, on fast and slow Ca2+ channel currents of isolated single vascular smooth muscle cells were investigated by a whole-cell voltage-clamp method combined with an intracellular perfusion technique. Single smooth muscle cells were prepared by collagenase treatment from guinea pig small mesenteric arteries (diameter of less than 300 micron). With Cs+-rich solution in the pipette and isotonic Ba2+ solution (100 mM) in the bath, depolarizing pulses evoked two types of the Ca2+ channel current. Depolarizing pulses from the holding potential of -80 mV to over -30 mV evoked a fast Ca2+ channel current. This fast component was inhibited by shifting the holding potential in a positive direction. With a holding potential of -40 mV, the fast component was almost inhibited. In contrast, the slow current was evoked by command potentials to above -10 mV, and its full amplitude was preserved at the holding potential of -40 mV. Without ATP in the pipette, the fast current was dominant. Increase in the ATP concentration in the pipette (0.3 to 5 mM) enhanced the slow current but did not affect the fast current. Maximum enhancement of the slow current was observed at 5 mM ATP. Increase in ATP concentration, however, did not modify the shape of the current trace and the steady state inactivation curve of the slow current. Maximum amplitudes of the fast current and slow current recorded with 5 mM ATP averaged 17.4 pA (SD of 10.4 pA, n = 30; observed at -10 mV to +10 mV) and 141.8 pA (SD of 27.1 pA, n = 30; observed at +30 mV to +40 mV), respectively. Presence of CN- and 2-deoxy-D-glucose (without glucose) in the bath, and absence of ATP in the pipette, abolished the slow current within 10 minutes; in contrast, it took more than 10 minutes to depress the fast current. The inhibitory effect of CN- and 2-deoxy-D-glucose on the slow current was reduced by intracellular application of ATP. In summary, the activation of the slow Ca2+ channel required physiological concentration of ATP, whereas the fast channel current was preserved, even under ATP-free conditions. These results indicate that only the slow current is a metabolically dependent Ca2+ channel current in these vascular smooth muscle cells.
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PMID:ATP regulation of the slow calcium channels in vascular smooth muscle cells of guinea pig mesenteric artery. 253 96

A suspension of renal tubule fragments from the rat was prepared by a method involving collagenase digestion of the excised renal cortex and dispersion of the digest by passage through a nylon mesh. Through the use of scanning electronmicroscopy it was confirmed that the tubular lumena were patent, thus ensuring access of medium to both the luminal and the contraluminal membranes of the tubular cells. The viability of the tubule fragments was ascertained by measuring the rate of formation of glucose from pyruvate as substrate and the uptake of [14C] L-lysine against a concentration gradient. The uptake of L-lysine was unimpaired in the presence of gentamicin (10(-3) M), suggesting that there is no competition between this basic amino acid and the polycationic aminoglycoside for transport into tubular cells. The uptake of [3H] gentamicin was studied and found to be reduced in the presence of 2,4-dinitrophenol, potassium cyanide, and ouabain. The reduced uptake in the presence of ouabain was interpreted to mean that a component of gentamicin uptake, which occurs at the luminal membrane, may be driven by the Na+ gradient created by Na+-K+ATPase activity at the contraluminal membrane. This renal tubule preparation offers advantages over the kidney-slice technique for studies into the mechanisms of aminoglycoside nephrotoxicity.
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PMID:The use of renal tubule fragments isolated from the rat to investigate aspects of gentamicin nephrotoxicity. 308 63

Several studies suggest that a portion of hepatocellular nonesterified fatty acid uptake may be carrier mediated. To further investigate this process, initial rates (Vo) of [14C]oleate uptake into rat hepatocytes, isolated by collagenase perfusion and incubated at 37 degrees C with oleate in the presence of bovine serum albumin, were studied as a function of the concentration of unbound [14C]oleate in the medium. Vo was saturable with increasing unbound oleate concentration (Km = 8.3 X 10(-8) M; Vmax = 197 pmol per min per 5 X 10(4) hepatocytes) and was not inhibited by up to 40 microM sulfobromophthalein, taurocholate, or cholic acid. Oleate uptake was sodium dependent. Vo was significantly diminished when Li+, K+, choline, or sucrose were substituted for Na+ in the incubation medium and was reduced 46% by 1 mM ouabain. Uptake was also markedly reduced after exposure of cells to metabolic inhibitors (e.g., 2,4-dinitrophenol, carbonyl cyanide m-chlorophenylhydrazone, antimycin, KCN). To evaluate the physiologic significance of the previously isolated rat liver plasma membrane fatty acid-binding protein, the effect of an antibody directed against this protein on hepatocellular [14C]oleate uptake was examined. Preincubation of hepatocytes with the IgG fraction of this antiserum inhibited Vo of [14C]oleate by up to 65% in dose-related fashion, without altering Vo for [35S]sulfobromophthalein, [14C]taurocholate, or [3H]cholate. These data indicate that at least a portion of hepatocellular oleate uptake is energy dependent, sodium linked, and mediated by a specific liver plasma membrane-fatty acid-binding protein.
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PMID:Hepatocellular uptake of oleate is energy dependent, sodium linked, and inhibited by an antibody to a hepatocyte plasma membrane fatty acid binding protein. 345 44

The metabolism of saturated nitriles, including acetonitrile, has been assumed to occur by a cytochrome P-450-dependent oxidation at the alpha-carbon, yielding a cyanohydrin intermediate which may spontaneously degrade to hydrogen cyanide and an aldehyde. However, results of studies in our laboratory suggest that formaldehyde is not a metabolite of acetonitrile. Since acetonitrile is structurally similar to iodomethane, a substrate for glutathione (GSH) S-transferases, we hypothesized that the metabolism of acetonitrile to cyanide might also occur by a nucleophilic substitution reaction involving GSH. The present studies were conducted to investigate these hypotheses and to further our study of the effects of acetone on acetonitrile metabolism. Female Sprague-Dawley rats were pretreated with buthionine sulfoximine BSO (4 mmol/kg ip, at -4 and -2 hr), cobalt heme (90 mumol/kg sc, at -48 hr), acetone (1960 mg/kg po, at -24 hr), or vehicle, and hepatocytes were isolated after collagenase perfusion of the liver. BSO reduced the cellular GSH content by greater than 80%, but did not appear to affect the metabolism of acetonitrile: the liberation of cyanide correlated with cytochrome P-450, and not GSH, concentrations. Cobalt heme depleted hepatocellular cytochrome P-450 (-45%) content, decreased cell yield and viability, and resulted in a marked reduction in the metabolism of acetonitrile to cyanide. Cobalt heme did not affect the recovery of sodium cyanide from hepatocyte suspensions. Pretreatment of rats with acetone resulted in a twofold increase in the metabolism of acetonitrile to cyanide. Addition of acetone in vitro inhibited acetonitrile metabolism, with an IC50 of 319 microM.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The metabolism of acetonitrile to cyanide by isolated rat hepatocytes. 355 37

Previous studies have shown that the administration of cadmium causes extensive necrosis of the testes and, eventually, a high incidence of interstitial cell tumors. However, the interactions of cadmium with interstitial cells of the testes have not been well defined. Therefore, this study was designed to assess the uptake of cadmium into this potential target cell of cadmium carcinogenesis. Interstitial cells were prepared by collagenase dispersion of decapsulated Wistar rat testes and separated from seminiferous tubules by unit gravity sedimentation. Such preparations showed a high exclusion rate of trypan blue. The interstitial cell preparations were incubated at 33 degrees with various concentrations of cadmium (1.0 to 100 microM) for periods ranging from 0.5 to 60 min. At the end of the incubation, cellular cadmium was separated from cadmium in the media by centrifugation through an oil layer. Initial experiments showed three distinct phases of cadmium influx into interstitial cells, a primary rapid velocity phase (V0; 0 to 1.5 min), a second intermediate velocity phase (V1; 3 to 12 min), and a third low velocity phase (V2; 15 to 60 min). V2 appeared to have both influx and efflux components, as efflux experiments indicated an approximate 20% loss of cadmium from 15 to 60 min. The initial phase was found to be nonsaturable and was not decreased by inclusion of potassium cyanide (1.0 mM), N-ethylmaleimide (1.0 mM), or zinc (100 microM) in the incubation mixture. However, V1 was found to be saturable between 50 and 100 microM cadmium and was substantially decreased by the inclusion of potassium cyanide, N-ethylmaleimide or zinc during incubation. These data suggest that cadmium is taken up into interstitial cells by a transport system that may normally function in zinc uptake and may possibly constitute carrier mediated or active transport.
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PMID:Interactions of cadmium with interstitial tissue of the rat testes. Uptake of cadmium by isolated interstitial cells. 401 91


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