Redox regulatory mechanisms of cellular signal transduction.
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Animals↗
Biomedical subjects
K A Robinson
Publications and source records attributed to K A Robinson.
SURVEY AND SUMMARY: Saccharomyces cerevisiae basic helix-loop-helix proteins regulate diverse biological processes.
Basic helix-loop-helix (bHLH) proteins are among the most well studied and functionally important regulatory proteins in all eukaryotes. The HLH domain dictates dimerization to create homo- and heterodimers. Dimerization juxtaposes the basic regions of the two monomers to create a DNA interaction surface that recognizes the consensus sequence called the E-box, 5'-CANNTG-3'. Several bHLH proteins have been identified in the yeast Saccharomyces cerevisiae using traditional genetic methodologies. These proteins regulate diverse biological pathways. The completed sequence of the yeast genome, combined with novel methodologies allowing whole-genome expression studies, now offers a unique opportunity to study the function of these bHLH proteins. It is the purpose of this review to summarize the current knowledge of bHLH protein function in yeast.
Amino Acid Sequence↗
The effect of endovascular irradiation on platelet recruitment at sites of balloon angioplasty in pig coronary arteries.
BACKGROUND: Endovascular irradiation (EI) inhibits balloon-induced neointima formation in animals and is now in clinical trials for restenosis prevention. However, little is known of the effect of EI on vessel thrombogenicity due to delayed arterial healing. We investigated EI effects on platelet recruitment in pig coronary arteries. METHODS AND RESULTS: EI was performed using (90)Sr/Y at 0 Gray (Gy), 15Gy, or 30Gy at 2 mm after balloon overstretch injury. At 1 day, 1 week, and 1 month, platelet recruitment and thrombus formation were assessed using autologous (111)In-oxine-platelet labeling and light and scanning electron microscopy. In balloon-injured nonirradiated vessels, there was complete reendothelialization at 1 month, and platelet recruitment was similar to normal uninjured arteries. In irradiated vessels, scanning electron microscopy showed incomplete reendothelialization at 1 month, and these areas demonstrated attachment of activated platelets. Light microscopy of irradiated coronaries showed adherent partially organized thrombi and incomplete resolution of intramural hemorrhages. There was a significant increase in platelet recruitment at 1 month in arteries receiving EI at 15Gy (5.1+/-2. 8x10(6), P=0.02) or 30Gy (12.5+/-9.9x10(6), P=0.005) compared with nonirradiated controls (2.7+/-1.5x10(6)); 30Gy was also higher than 15Gy (P=0.05). Platelet recruitment was also increased for 30Gy compared with control at 1 day. CONCLUSIONS: Endovascular irradiation at 15Gy or 30Gy after balloon angioplasty results in incomplete endothelial recovery, impaired resolution of intramural hemorrhage, and a dose-dependent increase in platelet recruitment at 1 month.
Angioplasty, Balloon, Coronary↗
CPI-1189 inhibits interleukin 1beta-induced p38-mitogen-activated protein kinase phosphorylation: an explanation for its neuroprotective properties?
The p38 mitogen-activated protein kinase (p38-MAPK) is a central enzyme in one of the major protein kinase cascades that regulate proapoptotic and proinflammatory signal transduction. p38-MAPK is activated by receptor/ligand recognition events or by exposure to extracellular stressors, including oxidative stress. Activation of p38-MAPK is affected by dual phosphorylation on a specific inhibitory domain. Dual phosphorylation causes a structural change in the p38-MAPK enzyme which allows binding of ATP and target substrate. Agents which block ATP docking to phosphoactivated p38-MAPK are being investigated for treatment of inflammatory diseases and neurodegenerative pathologies. An alternative strategy for p38-MAPK antagonism would be the inhibition of p38-MAPK phosphoactivation. We now report potent inhibition of p38-MAPK phosphorylation by a synthetic benzamide (CPI-1189) which displays protective action against tumor necrosis factor-alpha (TNFalpha)-induced neurodegeneration. In primary astrocytes treated with interleukin 1beta (IL1beta), CPI-1189 inhibits p38-MAPK phosphorylation at concentrations of 10 nM or less. While the precise molecular target of CPI-1189 remains unknown, these findings suggest a novel mechanism for the neuroprotective properties of the compound. These findings also indicate that antagonism of the p38-MAPK may be achieved through pharmacological inhibition of p38-MAPK phosphorylation, a strategy that is conceptually distinct from direct inhibition of ATP binding to the active enzyme.
Animals↗
Doses of olanzapine, risperidone, and haloperidol used in clinical practice: results of a prospective pharmacoepidemiologic study.
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Antipsychotic Agents↗
Inhibition of neointima formation by tranilast in pig coronary arteries after balloon angioplasty and stent implantation.
OBJECTIVES: We evaluated the effect of orally administered tranilast, N-(3,4-dimethoxycinnamoyl) anthranilic acid, on histologic and histomorphometric changes after angioplasty or stent implantation in pig coronary arteries. BACKGROUND: Tranilast, which has antikeloid and antiallergic properties and therefore may modulate the fibrotic and inflammatory tissue responses to angioplasty and stenting, has been shown to inhibit angiographic restenosis in small clinical trials. However, its effect on histomorphometric changes in coronary arteries after angioplasty and stenting is unknown. METHODS: Following initial pharmacokinetic studies in two pigs to determine desirable plasma levels of orally administered tranilast, 36 crossbred juvenile pigs were randomized to placebo or tranilast before undergoing balloon angioplasty in both the left anterior descending and left circumflex plus stent implantation in the right coronary artery. Oral tranilast was administered at 3 g/day starting 3 days before coronary injury and continued for 28 days until euthanasia. Injured vessels were harvested and sections analyzed by computer-assisted microscopic planimetry. RESULTS: In balloon-injured vessels, tranilast was associated with a 37% reduction in neointimal area normalized to fracture length (0.47 +/- 0.01 vs. 0.74 +/- 0.03 mm; p < 0.001) and a 23% reduction in adventitial area normalized to vessel size (0.43 +/- 0.02 vs. 0.56 +/- 0.03; p = 0.003). In stented arteries, neointimal area normalized to injury score was 32% lower in the tranilast-treated group compared to control (1.94 +/- 0.17 vs. 2.86 +/- 0.29; p = 0.01). CONCLUSIONS: In pig coronary arteries, tranilast was associated with a reduction in neointima formation and adventitial reaction after balloon injury. In stented vessels, tranilast was associated with a reduction in neointima formation normalized to injury score.
Administration, Oral↗
Prevention of thromboembolism in atrial fibrillation. A meta-analysis of trials of anticoagulants and antiplatelet drugs.
OBJECTIVE: Appropriate use of drugs to prevent thromboembolism in patients with atrial fibrillation (AF) involves comparing the patient's risk of stroke and risk of hemorrhage. This review summarizes the evidence regarding the efficacy of these medications. METHODS: We conducted a meta-analysis of randomized controlled trials of drugs used to prevent thromboembolism in adults with nonpostoperative AF. Articles were identified through the Cochrane Collaboration's CENTRAL database and MEDLINE until May 1998. MAIN RESULTS: Eleven articles met criteria for inclusion in this review. Warfarin was more efficacious than placebo for primary stroke prevention (aggregate odds ratio [OR] of stroke = 0.30, 95% confidence interval [CI] 0.19, 0.48), with moderate evidence of more major bleeding (OR 1.90; 95% CI 0.89, 4.04). Aspirin was inconclusively more efficacious than placebo for stroke prevention (OR 0.56, 95% CI 0.19, 1.65), with inconclusive evidence regarding more major bleeds (OR 0.81, 95% CI 0.37, 1.77). For primary prevention, assuming a baseline risk of 45 strokes per 1,000 patient-years, warfarin could prevent 30 strokes at the expense of only 6 additional major bleeds. Aspirin could prevent 17 strokes, without increasing major hemorrhage. In direct comparison, there was evidence suggesting fewer strokes among patients on warfarin than among patients on aspirin (aggregate OR 0.64, 95% CI 0.43, 0.96), with only suggestive evidence for more major hemorrhage (OR 1.60, 95% CI 0.77,3.35). However, in younger patients, with a mean age of 65 years, the absolute reduction in stroke rate with warfarin compared with aspirin was low (5.5 per 1,000 person-years) compared with an older group (15 per 1,000 person-years). CONCLUSION: In general, the evidence strongly supports warfarin for patients with AF at average or greater risk of stroke. Aspirin may prove to be useful in subgroups with a low risk of stroke, although this is not definitively supported by the evidence.
Anticoagulants↗
The promoter of the yeast INO4 regulatory gene: a model of the simplest yeast promoter.
In Saccharomyces cerevisiae, the phospholipid biosynthetic genes are transcriptionally regulated in response to inositol and choline. This regulation requires the transcriptional activator proteins Ino4p and Ino2p, which form a heterodimer that binds to the UAS(INO) element. We have previously shown that the promoters of the INO4 and INO2 genes are among the weakest promoters characterized in yeast. Because little is known about the promoters of weakly expressed yeast genes, we report here the analysis of the constitutive INO4 promoter. Promoter deletion constructs scanning 1,000 bp upstream of the INO4 gene identified a small region (-58 to -46) that is absolutely required for expression. S1 nuclease mapping shows that this region contains the transcription start sites for the INO4 gene. An additional element (-114 to -86) modestly enhances INO4 promoter activity (fivefold). Thus, the region required for INO4 transcription is limited to 68 bp. These studies also found that INO4 gene expression is not autoregulated by Ino2p and Ino4p, despite the presence of a putative UAS(INO) element in the INO4 promoter. We further report that the INO4 steady-state transcript levels and Ino4p levels are regulated twofold in response to inositol and choline, suggesting a posttranscriptional mechanism of regulation.
Basic Helix-Loop-Helix Proteins↗
Lower calorie intake enhances muscle insulin action and reduces hexosamine levels.
Previous studies have demonstrated enhanced insulin sensitivity in calorie-restricted [CR, fed 60% ad libitum (AL) one time daily] compared with AL-fed rats. To evaluate the effects of reduced food intake, independent of temporal differences in consumption, we studied AL (unlimited food access)-fed and CR (fed one time daily) rats along with groups temporally matched for feeding [fed 3 meals (M) daily]: MAL and MCR, eating 100 and 60% of AL intake, respectively. Insulin-stimulated glucose transport by isolated muscle was increased in MCR and CR vs. AL and MAL; there was no significant difference for MCR vs. CR or MAL vs. AL. Intramuscular triglyceride concentration, which is inversely related to insulin sensitivity in some conditions, did not differ among groups. Muscle concentration of UDP-N-acetylhexosamines [end products of the hexosamine biosynthetic pathway (HBP)] was lower in MCR vs. MAL despite unaltered glutamine-fructose-6-phosphate aminotransferase activity (rate-limiting enzyme for HBP). These results indicate that the CR-induced increase in insulin-stimulated glucose transport in muscle is attributable to an altered amount, not timing, of food intake and is independent of lower triglyceride concentration. They further suggest that enhanced insulin action might involve changes in HBP.
3-O-Methylglucose↗
Reduced thrombus formation by hyaluronic acid coating of endovascular devices.
Biocompatible stent coatings may alleviate problems of increased (sub)acute thrombosis after stent implantation. Hyaluronic acid (HA), a ubiquitous, nonsulfated glycosaminoglycan, inhibits platelet adhesion and aggregation and prolongs bleeding when administered systemically. However, the effects of immobilized HA for reducing stent platelet deposition in vivo are unknown. We therefore quantified the antithrombotic effects of coating stainless steel stents and tubes with HA using an established baboon thrombosis model under physiologically relevant blood flow conditions. HA-coated and uncoated (control) stents (3.5 mm in diameter, n=32) and stainless steel tubes (4.0 mm in diameter, n=18) were deployed into exteriorized arteriovenous shunts of conscious, nonanticoagulated baboons. Accumulation of (111)In-radiolabeled platelets was quantified by continuous gamma-camera imaging during a 2-hour blood exposure period. HA coating resulted in a significant reduction in platelet deposition in long (4 cm) tubes (0.24+/-0.15 x 10(9) versus 6.12+/-0.49 x 10(9) platelets; P<0.03), short (2 cm) stainless steel tubes (0.18+/-0.06 x 10(9) versus 3.03+/-0.56 x 10(9) platelets; P<0.008), and stents (0.82+/-0.20 x 10(9) versus 1.83+/-0. 23 x 10(9) platelets; P<0.02) compared with uncoated control devices. Thus, HA coating reduces platelet thrombus formation on stainless steel stents and tubes in primate thrombosis models. These results indicate that immobilized HA may represent an attractive strategy for improving the thromboresistance of endovascular devices.
Animals↗
High glucose and glucosamine induce insulin resistance via different mechanisms in 3T3-L1 adipocytes.
Sustained hyperglycemia induces insulin resistance, but the mechanism is still incompletely understood. Glucosamine (GlcN) has been extensively used to model the role of the hexosamine synthesis pathway (HSP) in glucose-induced insulin resistance. 3T3-L1 adipocytes were preincubated for 18 h in media +/- 0.6 nmol/l insulin containing either low glucose (5 mmol/l), low glucose plus GlcN (0.1-2.5 mmol/l), or high glucose (25 mmol/l). Basal and acute insulin-stimulated (100 nmol/l) glucose transport was measured after re-equilibration in serum and insulin-free media. Preincubation with high glucose or GlcN (1-2.5 mmol/l) inhibited basal and acute insulin-stimulated glucose transport only if insulin was present during preincubation. However, only preincubation with GlcN plus insulin inhibited insulin-stimulated GLUT4 translocation. GLUT4 and GLUT1 protein expression were not affected. GlcN (2.5 mmol/l) increased cellular UDP-N-acetylhexosamines (UDP-HexNAc) by 400 and 900% without or with insulin, respectively. High glucose plus insulin increased UDP-HexNAc by 30%. GlcN depleted UDP-hexoses, whereas high glucose plus insulin increased them. Preincubation with 0.5 mmol/l GlcN plus insulin maximally increased UDP-HexNAc without affecting insulin-stimulated or basal glucose transport. GlcN plus insulin (but not high glucose plus insulin) caused marked GlcN dose-dependent accumulation of GlcN-6-phosphate, which correlated with insulin resistance of glucose transport (r = 0.935). GlcN plus insulin (but not high glucose plus insulin) decreased ATP (10-30%) and UTP (>50%). GTP was not measured, but GDP increased. Neither high glucose plus insulin nor GlcN plus insulin prevented acute insulin stimulation (approximately 20-fold) of insulin receptor substrate 1-associated phosphatidylinositol (PI)-3 kinase. We have come to the following conclusions. 1) Chronic exposure to high glucose or GlcN in the presence of low insulin caused insulin resistance of glucose transport by different mechanisms. 2) GlcN inhibited GLUT4 translocation, whereas high glucose impaired GLUT4 "intrinsic activity" or membrane intercalation. 3) Both agents may act distally to PI-3 kinase. 4) GlcN has metabolic effects not shared by high glucose. GlcN may not model HSP appropriately, at least in 3T3-L1 adipocytes.
3T3 Cells↗
Anesthesia management during cataract surgery.
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Anesthesia, Conduction↗
Efficacy of agents for pharmacologic conversion of atrial fibrillation and subsequent maintenance of sinus rhythm: a meta-analysis of clinical trials.
CONTEXT: Physicians have little evidentiary guidance for pharmacologic agent selection for atrial fibrillation (AF). OBJECTIVE: To assess antiarrhythmic agent efficacy for AF conversion and subsequent maintenance of sinus rhythm (MSR). DATA SOURCE: We searched the clinical trial database of the Cochrane Collaboration and MEDLINE encompassing literature from 1948 to May 1998. STUDY SELECTION: We selected 36 (28%) articles eligible as randomized trials of nonpostoperative AF conversion or MSR in adults. DATA EXTRACTION: Study quality; rates of conversion, MSR, and adverse events were extracted. DATA SYNTHESIS: Compared with control treatment (placebo, verapamil, diltiazem, or digoxin), the odds ratio (OR) for conversion was greatest for ibutilide/dofetilide (OR=29.1; 95% confidence interval [CI], 9.8-86.1) and flecainide (OR=24.7; 95% CI, 9.0-68.3). Less strong but conclusive evidence existed for propafenone (OR=4.6; 95% CI, 2.6-8.2). Quinidine (OR=2.9; 95% CI, 1.2-7.0) had moderate evidence of efficacy for conversion. Disopyramide (OR=7.0; 95% CI, 0.3-153.0) and amiodarone (OR=5.7; 95% CI, 1.0-33.4) had suggestive evidence of efficacy. Sotalol (OR=0.4; 95% CI, 0.0-3.0) had suggestive evidence of negative efficacy. For MSR, strong evidence of efficacy existed for quinidine (OR=4.1; 95% CI, 2.5-6.7), disopyramide (OR=3.4; CI, 1.6-7.1), flecainide (OR=3.1; 95% CI, 1.5-6.2), propafenone (OR=3.7; 95% CI, 2.4-5.7), and sotalol (OR=7.1; 95% CI, 3.8-13.4). The only amiodarone data, from comparison with disopyramide, provided moderate evidence of efficacy for MSR. No trial evaluated procainamide. Direct agent comparisons and adverse event data were limited. CONCLUSIONS: Although multiple antiarrhythmic agents had strong evidence of efficacy compared with control treatment for MSR, ibutilide/dofetilide and flecainide had particularly strong evidence of efficacy compared with control treatment for AF conversion. There is sparse and inconclusive evidence on direct agent comparisons and adverse event rates. Obtaining information regarding these relative efficacies should be a research priority.
Aged↗
Management of new onset atrial fibrillation.
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Adult↗
Fibrocellular tissue responses to endovascular and external beam irradiation in the porcine model of restenosis.
PURPOSE: Endovascular radiation has reduced postangioplasty restenosis in preclinical and early clinical studies. External radiation treatment may have advantages over endovascular therapy. We examined vascular and perivascular tissue responses to endovascular and external irradiation in pig coronary arteries. METHODS AND MATERIALS: Ninety-one animals received endovascular or external radiation following balloon injury and were sacrificed at 14, 30, or 180 days. Injured segments of coronary vessels including perivascular and myocardial tissues were evaluated with histochemistry. RESULTS: Endovascular radiation was associated with delayed arterial wound healing as late as 6 months, evidenced by paucity of smooth muscle alpha-actin in neointimal cells compared to control. External treatment was associated with increased collagen in neointima and adventitia, and focal interstitial necrosis in adjacent myocardium. CONCLUSIONS: These investigations showed whole-heart 14 Gy external radiation treatment following coronary injury exacerbated certain aspects of arterial healing. In addition focal myocardial necrosis and fibrosis was observed following external but not endovascular irradiation. Endovascular radiation has some advantages over external irradiation; however the persistence of a synthetic smooth muscle cell phenotype in the neointima at 6 months suggests ionizing radiation in general may have profound effects on vessel architecture over the long term.
Actins↗
Basal protein phosphorylation is decreased and phosphatase activity increased by an antioxidant and a free radical trap in primary rat glia.
Reversible protein phosphorylation regulates a wide array of cellular functions. Cells respond to cytokines and various stressors via phosphorylation and thus activation of one or more of the mitogen-activated protein kinase (MAPK) pathways. Involvement of these signal transduction pathways has been implicated in numerous pathologies, including inflammation. Using a primary glia cell culture, we show here that the antioxidant N-acetylcysteine (NAC) and the nitrone-based free radical trap, alpha-phenyl-N-tert-butyl nitrone (PBN), reduce total basal protein phosphorylation in a concentration-dependent manner as assessed by phosphotyrosine analysis and by [gamma-32P]ATP transfer radioassay. In addition we show that NAC inhibits H2O2-induced phosphatase inactivation in glia cell lysate. The PBN- and NAC-induced reduction in protein phosphorylation is accompanied by an increase in phosphatase activity, suggesting that PBN and NAC reduce protein phosphorylation by globally augmenting oxidant-sensitive phosphatase activities. These results partly explain why certain antioxidants also possess anti-inflammatory actions.
Acetylcysteine↗
Phenyl-N-tert-butylnitrone demonstrates broad-spectrum inhibition of apoptosis-associated gene expression in endotoxin-treated rats.
Systemic exposure to gram-negative bacterial substances such as lipopolysaccharide (LPS, or endotoxin) induces an uncontrolled, massive inflammatory reaction which culminates in multiple system organ failure and death. Septic shock often does not respond to corticosteroids; however, certain low-molecular-weight antioxidant compounds have been discovered to possess potent anti-inflammatory action, and some of these novel compounds can rescue animals from experimentally induced septic shock. Phenyl-N-tert-butylnitrone (PBN) is the archetype of the nitrone class of antioxidants which we have previously shown to suppress LPS-induced cytokine biosynthesis in vivo. Using a multiprobe ribonuclease protection assay, we now demonstrate the ability of PBN to suppress proapoptotic gene expression in the LPS-induced model of endotoxic shock. The broad-spectrum gene-suppressive affects of PBN are discussed in the context of inflammatory signal transduction and models are proposed to explain why certain antioxidants may also possess anti-inflammatory and antiapoptotic properties.
Animals↗
Redox-sensitive protein phosphatase activity regulates the phosphorylation state of p38 protein kinase in primary astrocyte culture.
Reactive oxygen species (ROS) have been implicated as second messengers that activate protein kinase cascades, although the means by which ROS regulate signal transduction remains unclear. In the present study, we show that interleukin 1beta (IL1beta), H2O2, and sorbitol-induced hyperosmolarity mediate a 5- to 10-fold increase in phosphorylation (activation) of the p38 protein kinase in rat primary glial cells as measured by analyses of Western blots using an antibody directed against the dually phosphorylated (active) p38. Additionally, IL1beta was found to elicit H2O2 synthesis in these cells. Concurrent with p38 phosphorylation, all three stimulation paradigms caused an inhibition of protein phosphatase activity. Phenyl-tert-butyl nitrone (PBN), a nitrone-based free radical trap and N-acetyl-cysteine (NAC), a thiol reducing agent, were examined for their effects on the phosphorylation of p38 as well as phosphatase activity. Pretreatment of cells with either PBN or NAC at 1.0 mM suppressed IL1beta H2O2, and sorbitol-mediated activation of p38 and significantly increased phosphatase activity. These data suggest that ROS, particularly H2O2, are used as second messenger substances that activate p38 in part via the transient inactivation of regulatory protein phosphatases.
Acetylcysteine↗