Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0432222 (SEM)
 47,337 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The aim of this study was to compare the effects of increased concentrations of MgADP, inorganic phosphate (Pi) and H+ ([MgADP], [Pi] and [H+], respectively) on the rate of relaxation in two different muscle types: skinned muscle fibres from the frog Rana temporaria and myofibrillar bundles from the giant Pacific acorn barnacle Balanus nubilus. Relaxation transients are produced by the photolysis of diazo-2 and are well fitted with a double exponential curve, giving two rate constants: k1 [5.6+/-0.1 s-1 for barnacle, n=30; 26.3+/-0.7 s-1 for frog, n=14 (mean+/-SEM)] and k2 [0.6+/-0.1 s-1 in barnacle, n=30; 10.4+/-1.0 s-1 in frog, n=14 (mean+/-SEM)], at 10 degrees C. Decreasing the pH by 0.5 pH units did not significantly affect k1 for barnacle relaxation [5.6+/-0.1 s-1 (mean+/-SEM), n=15] compared to the decrease in k1 of 40% seen in frog. Use of the Ca2+-sensitive fluorescent label acrylodan on barnacle wild-type troponin C demonstrated that decreasing the pH from 7.0 to 6.6 only alters the pCa50 value by 0.23 in the cuvette, while stopped-flow experiments with acrylodan revealed no significant change in koff from the labelled protein [322+/-32 s-1 at pH 7.0 and 381+/-24 s-1 (mean+/-SEM) at pH 6.6]. Increasing [MgADP] by 20 microM (50 microM added ADP) from control values of 50 microM in frog decreased k1 to 12.3+/-0.4 s-1 (mean+/-SEM, n=8), and at 400 microM MgADP, k1=9.6+/-0.1 s-1 (mean+/-SEM, n=12). In barnacle, 500 microM MgADP had a much smaller effect on k1 (4.0+/-0. 9 s-1, mean+/-SEM, n=8). Increasing the free [Pi] from the contaminant level of 0.36 mM to 1.9 mM slowed k1 by approximately 15% in barnacle [4.8+/-0.8 s-1, mean+/-SEM, n=7], compared to a approximately 30% reduction seen in frog. We conclude that the differences between barnacle and frog seen here are most probably due to different isomers of the contractile proteins, and that events underlying the crossbridge cycle are the same or similar. We interpret our results according to a model of crossbridge transitions during relaxation.
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PMID:A diazo-2 study of relaxation mechanisms in frog and barnacle muscle fibres: effects of pH, MgADP, and inorganic phosphate. 992 60

To determine if treatment with covalently bound heparin (Carmeda Bioactive Surface (CBAS)) to the synthetic surface of the extracorporeal circuit (ECC) would alter the stereotypic pattern of adverse platelet alterations, 450 ml of heparinized blood (lU/ml) was recirculated at a flow rate of twice the circulating volume (L/min) for 2 hrs at 37 degrees C through either untreated (CONT,n=7) or treated (CBAS,n=7) circuits constructed of identical components including a pediatric (0.8m 2) reversed hollow fiber membrane oxygenator. In CONT circuits, platelet count maintained 88+1% (x+/-SEM) of its initial level in the circuit prime sample, dropped to 36+/-6% after 5 min, and returned to 56+/-2% following 2 hrs of ECC. In CBAS circuits, platelet count in the circuit prime sample demonstrated 90+/-4%, decreased to 68+/-10% after 5 min (p less than 0.05) and declined further to 45+/-5% after 2 hrs (NS). Although platelets from both groups retained reactivity to ADP after priming the circuit, only at 5 min of recirculation did CBAS circuits significantly preserve this responsiveness. In CONT circuits, baseline plasma levels of platelet factor 4 rose from 24+/-3 to 581+/-82 ng/ml in the primed circuit and continued to rise to 2933+/-276 ng/ml by 2 hrs of ECC. In contrast, CBAS circuits markedly reduced this release after 2 hrs (577+/-165 ng/ml). Furthermore by 2 hrs of ECC, plasma levels of thromboxane B 2 in the CBAS circuits were significantly reduced when compared to CONT circuits (3035+/-1529 vs 29916+/-16293 pg/ml, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:The effects of heparin bound surface modification (Carmeda Bioactive Surface) on human platelet alterations during simulated extracorporeal circulation. 1014 74

An improved method for the measurement of tissue metabolites associated with cellular energetic state by capillary electrophoresis is described. This method allows 17 compounds present in a mixture of standards to be determined simultaneously within 43 min with good reproducibility. ATP, ADP, AMP, UTP, IMP, inosine, hypoxanthine, creatine, phosphocreatine, UDP-galactose, NAD and NADH were detected in samples of either rat heart tissue or rat neonatal cardiomyocytes. This method can detect compounds at concentrations of 5 microm in samples. Recoveries for ATP and phosphocreatine added to cardiomyocyte samples were 99.4 +/- 2.1% and 103.1 +/- 3.3%, respectively (mean +/- SEM, n = 3). Our method has been comprehensively validated and is capable of measuring a wider range of tissue metabolites important in assessing cellular energy status than existing methods.
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PMID:An improved capillary electrophoresis method for measuring tissue metabolites associated with cellular energy state. 1021 91

Clopidogrel, a potent novel platelet ADP-receptor antagonist, induces a significant inhibition of ADP-induced platelet aggregation. Maximum inhibition of 40 to 50% is observed 2 to 5 hours after a single 400 mg dose. The same level of inhibition is achieved with 75 mg once daily at steady state, i.e., after 3 to 7 days of repeated dosing. Based on these data, two studies were undertaken to investigate whether a treatment regimen comprising a large initial dose (loading dose) of clopidogrel, followed by daily doses of 75 mg, might provide a sustained steady-state level of inhibition of platelet aggregation induced by 5 microM of ADP within hours after first dosing. In one study, 10 healthy male subjects received a 375 mg loading dose of clopidogrel on day 1, then daily doses of 75 mg from day 2 to day 10. Mean inhibition of platelet aggregation, already significant at 30 minutes, reached 55+/-8.2% (+/-SEM) at 60 minutes, and a maximum of 80+/-3.6% at 5 hours. No further significant change was observed between 5 hours and 24 hours, and from day 2 through day 10 with subsequent daily doses of 75 mg. In the second study, conducted according to a randomized, single-blind design, four parallel treatment groups of nine healthy male subjects received a loading dose of 75 mg, 150 mg, 225 mg, or 300 mg of clopidogrel on day 1, respectively, and 75 mg once daily from day 2 to day 5. Mean (+/-SD) inhibition of platelet aggregation over the 2 to 24 hours post-loading dose period was 22+/-14.5%, 21+/-13.4%, 35+/-20.6% and 31+/-13.3%, respectively. On day 5, it was 48+/-14.7%, 33 +/-14.1%, 51+/-15.7% and 40+/-10.9% for the 75, 150, 225 and 300 mg loading dose groups, respectively. The smallest day 1 to day 5 difference was observed for the 300 mg group and the largest for the 75 mg group, indicating that the development of the full inhibitory effect of clopidogrel was faster with the loading doses higher than with 75 mg, and fastest with the 300 mg loading dose. These data and those of previous studies indicate that a dose of 300 to 400 mg produces a rapid onset of the pharmacodynamic action of clopidogrel, with levels of inhibition close to steady-state reached within 2 hours.
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PMID:Clopidogrel loading dose regimens: kinetic profile of pharmacodynamic response in healthy subjects. 1044 Apr 17

The influence of clopidogrel 75 mg, given once daily for 10 days on hepatic P-450 mixed function oxidases, was examined by assessing its effect on the disposition of antipyrine, on urinary 6-betahydroxycortisol (6beta-OHC) and on the plasma activity of gamma-glutamyl transpeptidase. This double-blind, randomized, placebo-controlled study was conducted in two parallel groups of 10 healthy young volunteers. Subjects were required to fast for 12 hours before and for 4 hours after dosing. Antipyrine 10 mg/kg was administered in the morning, two days before treatment (day -2) and 24 hours after the last dose of clopidogrel or placebo. Plasma levels of antipyrine, and urinary excretion of antipyrine, 3-hydroxymethyl-antipyrine and nor-antipyrine were measured over 36 hours post-drug for pharmacokinetic determinations. Bleeding time and platelet aggregation induced by 5 microM of ADP were measured before treatment (baseline) and at regular intervals after dosing during treatment. Clopidogrel treatment had a marked effect on platelet aggregation and bleeding time. No significant change in the disposition of antipyrine was observed after the ingestion of clopidogrel over 10 days: mean AUC ratio (+/-SEM) for plasma antipyrine was 1.021+/-0.023 for the clopidogrel group versus 1.001+/-0.019 for the placebo group; mean day 10/day -2 t 1/2 ratios were 1.019+/-0.018 and 1.027+/-0.023, respectively. Urinary excretions of antipyrine and metabolites were unchanged by clopidogrel compared to placebo. The changes in plasma cortisol concentrations, 6beta-OHC excretion and serum gamma-glutamyl transpeptidase activities observed at the end of treatment were fully comparable between the two treatment groups. Thus, the different tests showed no evidence of hepatic enzyme induction by clopidogrel in a pharmacologically effective dose regimen.
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PMID:Clopidogrel and drug metabolism: absence of effect on hepatic enzymes in healthy volunteers. 1044 Apr 21

Platelet concentrate (PC) transfusions are useful for maintaining haemostasis in a variety of clinical situations. The function of transfused platelets is of critical importance, and changes on storage of buffy coat-prepared PC may influence their haemostatic potential. Total platelet adenine nucleotide content and platelet aggregation responses were studied, serially, in pooled buffy coat-derived PCs (n = 7), stored under UK recommended blood bank conditions, over the stipulated shelf-life of 5 days. Mean platelet volume (MPV), platelet counts and platelet distribution width (PDW) were also quantified. Total platelet ADP content decreased from 4.45+/-0.78 to 3.71+/-0.69 nmol/108 platelets (P<0.01, day 1 versus day 5, mean +/- SEM) over the shelf-life period. This was associated with reduced aggregatory responses: responses (expressed as percentage of maximum height) to 5 and 10 microM ADP decreased from 10.8+/-2.8% to 1.0+/-1.0% (P<0.005, 5 microM, day 1 versus day 5) and from 18.0+/-5.4% to 4.7+/-2.2% (P<0.02, 10 microM, day 1 versus day 5) while the decreased responsiveness was more pronounced for 4 microg/ml of collagen: 49.0+/-13.3% to 7.2+/-7.1% (P<0.01, day 1 versus day 4) and 49.9 +/-13.3% to 2.1+/-1.9% (P<0.001, day 1 versus day 5). These data indicate an acquired storage pool defect that is maximal by day 4 or 5 and accompanied by decreased platelet function, characterized by significant decreases in platelet aggregation responses. Addition of freeze-thawed plasma (autologous day 1) to PCs on days 2, 3, 4 and 5 did not alter the responses to ADP and collagen.
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PMID:Storage pool defect in pooled buffy coat platelet concentrates within the shelf-life period. 1076

Renin secretion can be stimulated by ATP via purinergic P2Y receptors. ATP is a cotransmitter with norepinephrine and is released from the cytosol during cell damage. Such release could account for the de novo renin expression seen in the proximal tubule in renal disease and in myocardial infarct borders. Whereas most P2Y purinoceptor subtypes utilize phosphoinositide signal-transduction pathways, the effector mechanisms of the subtype P2Y(11) also involve increases in cAMP, a well-known renin secretagogue and stimulus to renin production. The present study tested the effect of ATP on human renin gene (REN) promoter activity and the role of P2Y(11). By means of reverse transcriptase-polymerase chain reaction, we found that renin-expressing Calu-6 cells express P2Y(11) mRNA. Expression was also detected in the brain, kidney, testis, muscle, liver, and spleen. We made a novel cell line (Calu-6/P2Y11) in which P2Y(11) cDNA, under the control of a strong promoter, was stably integrated into genomic DNA. These cells produced P2Y(11) mRNA during culture. Treatment of Calu-6/P2Y11 cells with 1 mmol/L ATP caused a 3-fold increase in renin mRNA and protein over 36 hours. Transient transfection of Calu-6/P2Y11 cells with constructs containing 896 bp of human REN 5'-flanking DNA linked to the luciferase reporter gene led to a 5.8+/-0.6-fold increase (mean+/-SEM) in reporter activity in response to ATP (P=0.0015). In contrast, UTP produced only a 1.4+/-0.1-fold increase (P=0.016). For ADP, it was 1.7+/-0.1-fold (P=0.011). The response profile was ATP>ADP>AMP=adenosine=0, consistent with a P2Y(11) effect. Mutation of the cAMP response element (CRE) located at -222 in the REN promoter DNA abolished the effect of ATP. Furthermore, ATP induced a rapid, time-dependent increase in the phosphorylation of CRE binding protein (CREB) and activating transcription factor-1. These data implicate a cAMP pathway in mediation of the P2Y(11) effect. In conclusion, we have made a novel cell line that overexpresses the P2Y(11) purinoceptor. Stimulation of these cells by ATP activates a cAMP signal-transduction pathway that phosphorylates CREB and stimulates renin promoter activity via the CRE at -222. The data raise the possibility of a contribution of ATP/P2Y(11) effects to sympathetic stimulation of renin, as well as to responses in renin seen after tissue damage, such as in kidney disease and myocardial infarction.
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PMID:Capacity for purinergic control of renin promoter via P2Y(11) receptor and cAMP pathways. 1111 31

The adaptation of muscle oxidative function to 6 weeks of endurance cycle training was investigated in eight untrained subjects. Peak oxygen consumption (VO2peak) increased by 24% (2.69+/-0.21 versus 3.34+/-0.30 l O2 min(-1), mean +/-SEM, P<0.01) and lactate threshold intensity increased by 53% (121+/-13 versus 185+/-15 W, P<0.01) following the training period. Muscle biopsy samples were taken from vastus lateralis before and after training, and respiration in permeabilized muscle fibres was measured. Following training, non-ADP-stimulated respiration (VO) of skinned fibres increased by 35% (0.17+/-0.01 versus 0.23+/-0.01 mmol O2.min(-1).kg(-1) wet weight, P<0.05) and maximal ADP-stimulated respiration (VmaX) increased by 38% (1.17+/-0.07 versus 1.62+/-0.14 mmol O2.min(-1).kg(-1) wet weight, P<0.05). ADP sensitivity [i.e. the ratio between mitochondrial respiration (after correction for VO) at 0.1 mM ADP and Vmax] was reduced after training (0.40+/-0.05 versus 0.26+/-0.02; P<0.05). Mitochondrial resistance to oxidative stress was investigated by exposing skinned fibres to exogenous reactive oxygen species (ROS). ADP-stimulated respiration was reduced after ROS exposure and the relative decrease was similar before and after training. It is concluded that after endurance training: (1) the relative increase in maximal muscle fibre respiration exceeds that of whole-body oxygen uptake; (2) the sensitivity of mitochondrial respiration to ADP decreases; and (3) the impairment of oxidative function in skinned muscle fibres by ROS remains unchanged.
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PMID:Effect of endurance training on oxidative and antioxidative function in human permeabilized muscle fibres. 1148 74

The majority of the biological effects of pertussis toxin (PT) are the result of a toxin-catalyzed transfer of an adenosine diphosphate-ribose (ADP-ribose) moiety from NAD(+)to the alpha-subunits of a subset of signal-transducing guanine-nucleotide-binding proteins (G-proteins). This generally leads to an uncoupling of the modified G-protein from the corresponding receptor and the loss of effector regulation. This assay is based on the PT S1 subunit enzymatic transfer of ADP-ribose from NAD to the cysteine moiety of a fluorescent tagged synthetic peptide homologous to the 20 amino acid residue carboxyl-terminal sequence of the alpha-subunit of the G(i3)protein. The tagged peptide and the ADP-ribosylated product were characterized by HPLC/MS and MS/MS for structure confirmation. Quantitation of this characterized ADP-ribosylated fluorescently tagged peptide was by HPLC fluorescence using Standard Addition methodology. The assay was linear over a five hr incubation period at 20 degrees C at PT concentrations between 0.0625 and 4.0 microg/ml and the sensitivity of the assay could be increased several fold by increasing the incubation time to 24 h. Purified S1 subunit of PT exhibited 68.1+/-10.1% of the activity of the intact toxin on a molar basis, whereas the pertussis toxin B oligomer, the genetically engineered toxoid, (PT-9K/129G), and several of the other components of the Bordetella pertussis organism possessed little (<0.6%) or no detectable ribosylation activity. Commonly used pertussis vaccine reference materials, US PV Lot #11, BRP PV 66/303, and BRP PV 88/522, were assayed by this method against Bordetella pertussis Toxin Standard 90/518 and demonstrated to contain, respectively, 0.323+/-0.007, 0.682+/-0.045, and 0.757+/-0.006 microg PT/ml (Mean+/-SEM) or in terms of microg/vial: 3.63, 4.09 and 4.54, respectively. A survey of several multivalent pertussis vaccine products formulated with both whole cell as well as acellular components indicated that products possessed a wide range of ribosylation activities. The pertussis toxin S1 subunit catalyzed ADP- ribosylation of the FAC-Galpha(i3)C20 peptide substrate and its subsequent quantitation by HPLC was demonstrated to be a sensitive and quantitative method for measuring intrinsic pertussis toxin activity. This methodology not only has the potential to be an alternative physicochemical method to replace existing bioassay methodology, but has the added advantage of being a universal method applicable to the assay of pertussis toxin in both whole cell and acellular vaccines as well as bulk and final formulated vaccine products. Acceptance of this method by regulatory agencies and industry as a credible alternative to existing methods would, however, require validation in an international collaborative study against the widely accepted bioassay methods.
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PMID:A quantitative analysis for the ADP-ribosylation activity of pertussis toxin: an enzymatic-HPLC coupled assay applicable to formulated whole cell and acellular pertussis vaccine products. 1158 Feb 13

Platelets, a major constituent of thrombus, play a crucial role in the pathogenesis of acute ischemic coronary syndromes. The effect of ultraviolet laser emission on platelets within thrombi is unknown. The effects of increasing levels of laser energy on platelets in whole blood were investigated. Blood samples were obtained by aseptic venipuncture and anticoagulated with 3.8% sodium citrate. Samples were exposed to increased levels (0, 30, 45, 60 mJ/mm2; 25 Hz) of ultraviolet excimer laser fluence (308 nm wave-length) and then tested for ADP and collagen induced platelet aggregation, platelet concentration, and for platelet contractile force (PCF) development. Scanning electron microscopy was used to detect laser induced morphologic changes of platelets and by flow cytometric analysis to detect changes in expression of platelet surface antigens p-selectin (CD 62) and glycoprotein IIb/IIIa (CD 43). Exposure to excimer laser energy produced dose dependent suppression of platelet aggregation and force development ("stunned platelets"). ADP aggregation decreased from 8.0+/-1.1 Ohms (mean+/-SEM) to 3.7+/-0.8 Ohms (p<0.001) to 2.7+/-0.6 Ohms (p <0.001) and to 1.8+/-0.5 Ohms (p <0.001) as the laser energy increased from 0 to 30 to 45 to 60 mJ/mm2, respectively. Collagen induced aggregation decreased from 21.4+/-1.4 Ohms to 15.7+/-1.2 Ohms (p <0.001) to 11.7+/-1.1 Ohms (p <0.001) and to 9.9+/-1.0 Ohms (p <0.001), in response to the same incremental range of laser energy. Platelet contractile forces declined from 34,500+/-3700 to 27.800+/-2700 dynes as laser energy increased from 0 to 60 mJ/mm2 (p <0.03). Platelet concentration did not change with increasing laser energy. The expression of platelet surface antigen p-selectin (CD 62) remained stable through increasing levels of laser energy exposures while the percentage of CD 43 positive platelets significantly increased with exposure to laser energy, yet the level of expression did not exceed 0.5% of cells. Thus, aggregation kinetics are altered in platelets exposed to ultraviolet laser energy as manifested by decreased platelet aggregation and reduction in platelet force development capability. The response is dose dependent and most pronounced at higher energy levels such as 60 mJ/mm2.
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PMID:Alterations of platelet aggregation kinetics with ultraviolet laser emission: the "stunned platelet" phenomenon. 1168 28


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