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Biomedical subjects

P Nguyen

Publications and source records attributed to P Nguyen.

At least 73 records · Page 4Linked to original sources
PubMed1999

The estrogen receptor enhances AP-1 activity by two distinct mechanisms with different requirements for receptor transactivation functions.

Estrogen receptors (ERs alpha and beta) enhance transcription in response to estrogens by binding to estrogen response elements (EREs) within target genes and utilizing transactivation functions (AF-1 and AF-2) to recruit p160 coactivator proteins. The ERs also enhance transcription in response to estrogens and antiestrogens by modulating the activity of the AP-1 protein complex. Here, we examine the role of AF-1 and AF-2 in ER action at AP-1 sites. Estrogen responses at AP-1 sites require the integrity of the ERalpha AF-1 and AF-2 activation surfaces and the complementary surfaces on the p160 coactivator GRIP1 (glucocorticoid receptor interacting protein 1), the NID/AF-1 region, and NR boxes. Thus, estrogen-liganded ERalpha utilizes the same protein-protein contacts to transactivate at EREs and AP-1 sites. In contrast, antiestrogen responses are strongly inhibited by ERalpha AF-1 and weakly inhibited by AF-2. Indeed, ERalpha truncations that lack AF-1 enhance AP-1 activity in the presence of antiestrogens, but not estrogens. This phenotype resembles ERbeta, which naturally lacks constitutive AF-1 activity. We conclude that the ERs enhance AP-1 responsive transcription by distinct mechanisms with different requirements for ER transactivation functions. We suggest that estrogen-liganded ER enhances AP-1 activity via interactions with p160s and speculate that antiestrogen-liganded ER enhances AP-1 activity via interactions with corepressors.

Binding Sites↗
PubMed1999

Heparin-induced thrombocytopenia in France, 1980-1998.

Due to the extensive use of unfractionated heparins in France, there is considerable experience with heparin-induced thrombocytopenia (HIT). It is recommended that platelet counts be performed twice a week for three weeks when patients are treated with any form of heparin. A drop in platelet counts can, however, occur not only in HIT patients but also for other unrelated reasons. For diagnosing HIT, all laboratories in France use platelet aggregometry inspite of poor sensitivity. Both false positive and false negative results are obtained. The serotonin release test is not used in France. The ELISA test for HIT does not always correlate with the platelet aggregation test and many patients with a positive ELISA test do not necessarily have other evidence for HIT. This is especially true in patients following cardiopulmonary bypass surgery. None of the available laboratory tests reliably identify patients with HIT. Patients with HIT should not be managed with low-molecular-weight heparins, but danaparoid, argatroban and ancrod are viable options. Also, recombinant hirudin has been employed. All have advantages and disadvantages. At present, the diagnosis and management of patients with HIT remains difficult and properly designed clinical studies are needed to obtain answers to several open questions.

Anticoagulants↗
PubMed1999

Free and total platelet glycoprotein IIb/IIIa measurement in whole blood by quantitative flow cytometry during and after infusion of c7E3 Fab in patients undergoing PTCA.

A quantitative flow cytometry assay was used to evaluate the ex vivo kinetics of c7E3 Fab platelet effect in 16 patients undergoing PTCA treated with abciximab and compared with aggregometry assay. Immunolabeling of platelets was directly assessed on whole blood, using in parallel two monoclonal antibodies (Mabs) raised against GPIIIa, Mab1, the binding of which is inhibited by c7E3 Fab, and Mab2, the binding of which is not affected by c7E3 Fab. We found a severe and sustained inhibition of both GPIIb/IIIa receptors and platelet functions. The inter-individual variation in response to abciximab was low. A significant transient increase at H24 and H48 in the binding of Mab2 was found as an unexpected result, and confirmed in vitro. Results demonstrate that flow cytometry is a reliable method in agreement with aggregation. In addition, our results show that it is a standardized tool and a time-saving technique.

Abciximab↗
PubMed1999

Inhibition of fMLP-triggered respiratory burst of human monocytes by adenosine: involvement of A3 adenosine receptor.

Adenosine (Ado) is a potent anti-inflammatory agent acting on a variety of cell functions. However, its effects on human monocytes have been less well characterized. We investigated the effect of Ado and its receptor-specific analogs on NADPH oxidase activity with the use of luminol-enhanced chemiluminescence (CL). Adenosine inhibited fMLP-triggered NADPH oxidase activity with a maximal inhibition of 55+/-5%. IB-MECA, a selective A3 Ado receptor agonist reduced fMLP triggered NADPH oxidase activity more potently than the A2 receptor agonist CGS 2180 HCl (CGS) and the A1 Ado receptor agonist N-2-phenylethyl-adenosine (R-PIA). The inhibitory effect of Ado was reversed by neither the A1 Ado receptor antagonist 1,3-dipropyl-8(2-amino-4chlorophenyl)-xanthine (PACPX) nor the A2 Ado receptor antagonist 3,7-dimethyl-1-(2-propynyl)xanthine (DMPX). It was significantly reversed by the A1/A3 Ado receptor antagonist xanthine amine congener (XAC). Pretreatment of monocytes by cytochalasin B reversed the effect of Ado but not of dibutyryl cAMP (dBcAMP) on fMLP-CL response. KT 5720, a specific cAMP-dependent protein kinase inhibitor completely counteracted the inhibition of NADPH oxidase activity by dBcAMP but not by Ado. Using flow cytometry, we observed that Ado did not inhibit intracellular oxidative metabolism, whereas dBcAMP did. Furthermore, the inhibition of NADPH oxidase activity by Ado was not mediated by changes in cytosolic calcium. These results demonstrated that Ado inhibited NADPH oxidase activity via A3 Ado receptor independently of cAMP elevation or changes in calcium mobilization.

Adenosine↗
PubMed1998

Differential expression of Fas and Fas ligand in acute and chronic graft-versus-host disease: up-regulation of Fas and Fas ligand requires CD8+ T cell activation and IFN-gamma production.

The parent-into-F1 model of acute and chronic graft-vs-host disease (GVHD) was used as an example of in vivo cell-mediated or Ab-mediated responses, respectively, and the roles of Fas and Fas ligand (FasL) were investigated. Using both flow cytometry and PCR methodologies, we found that acute GVHD mice exhibited significant up-regulation of Fas and FasL, whereas Fas/FasL up-regulation in chronic GVHD mice was equal to or marginally greater than that in uninjected mice. Functional studies confirmed that Fas/FasL contributed to the anti-host CTL activity of splenocytes from acute GVHD mice, although a perforin-dependent pathway was also identified. Despite the presence of FasL on both donor CD4+ and CD8+ T cells in acute GVHD mice, depletion studies demonstrated that all the in vitro anti-host CTL activity resided in the CD8+ population. Furthermore, injection of CD8-depleted B6 spleen cells into F1 mice blocked Fas/FasL up-regulation and IFN-gamma production, resulting in chronic GVHD. Lastly, up-regulation of Fas/FasL in acute GVHD mice could be blocked by anti-IFN-gamma mAb in vivo. Thus, in this in vivo model of alloantigen immune responsiveness, Fas/FasL up-regulation is critically dependent on Ag-specific (donor) CD8+ T cell activation and IFN-gamma production. Donor CD4+ T cell activation in the absence of CD8+ T cell activation results in an autoantibody-mediated response, no significant Fas/FasL up-regulation, impaired elimination of autoreactive B cells, and persistent humoral autoimmunity.

Acute Disease↗
PubMed1998

Inhibition of the 3-hydroxy-3-methylglutaryl-coenzyme A reductase pathway induces p53-independent transcriptional regulation of p21(WAF1/CIP1) in human prostate carcinoma cells.

Progression through the cell cycle is controlled by the induction of cyclins and the activation of cognate cyclin-dependent kinases. The 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor lovastatin induces growth arrest and cell death in certain cancer cell types. We have pursued the mechanism of growth arrest in PC-3-M cells, a p53-null human prostate carcinoma cell line. Lovastatin treatment increased protein and mRNA levels of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1), increased binding of p21 with Cdk2, markedly inhibited cyclin E- and Cdk2-associated phosphorylation of histone H1 or GST-retinoblastoma protein, enhanced binding of the retinoblastoma protein to the transcription factor E2F-1 in vivo, and induced the activation of a p21 promoter reporter construct. By using p21 promoter deletion constructs, the lovastatin-responsive element was mapped to a region between -93 and -64 relative to the transcription start site. Promoter mutation analysis indicated that the lovastatin-responsive site coincided with the previously identified transforming growth factor-beta-responsive element. These data indicate that in human prostate carcinoma cells an inhibitor of the HMG-CoA reductase pathway can circumvent the loss of wild-type p53 function and induce critical downstream regulatory events leading to transcriptional activation of p21.

Apoptosis↗
PubMed1998

In vivo degradation of N-myc in neuroblastoma cells is mediated by the 26S proteasome.

N-myc is a short-lived transcription factor, frequently amplified in human neuroblastomas. The ubiquitin-proteasome system is involved in the degradation of many short-lived cellular proteins and previous studies have shown that ubiquitin-dependent proteolysis is implicated in the turn-over of N-myc in vitro. However, calpain has also been implicated in N-myc degradation in vitro. Here we report that, in vivo, N-myc is a sensitive substrate for the 26S proteasome in N-myc amplified neuroblastoma cells. We observed that inhibition of the 26S proteasome with two inhibitors, ALLnL and lactacystin, led to an elevation of the N-myc protein steady-state and increased N-myc protein polyubiquitination, as revealed by ubiquitin Western blotting. Pulse-chase experiments have shown that the increased N-myc levels resulted from stabilization of the protein. In contrast treatment with several calpain and cathepsin inhibitors failed to block N-myc degradation in vivo. Furthermore, fluorescence microscopy of ALLnL-treated cells localized N-myc exclusively to the nuclear compartment, suggesting the absence of a requirement for transport to the cytoplasm prior to degradation.

Acetylcysteine↗
PubMed1998

Role of glucose and glutamine synthesis in the differential recovery of 13CO2 from infused [2-13C] versus [1-13C] acetate.

Carbon exchange in the Krebs cycle may result in underestimation of substrate oxidation measured with 13C-labeled substrates, since carbon labeled in position 2 of acetyl-coenzyme A (CoA) could be incorporated into glucose (via gluconeogenesis) and glutamine. Five healthy volunteers were therefore infused with [1-13C] and [2-13C] acetate at a rate of 0.5 micromol x kg(-1) x min(-1) for 165 minutes on two different occasions in randomized order. Whole body acetate turnover did not differ between the two tracers: 7.9+/-0.3 and 7.5+/-0.6 micromol x kg(-1) x min(-1) (nonsignificant [NS]) for [1-13C] and [2-13C] acetate, respectively. Isotopic 13C enrichment was higher in expired CO2 (0.177+/-0.021 v 0.089+/-0.009 atom percent excess [APE], P < .01) and lower in glucose (0.074+/-0.017 v0.291+/-0.061 mole percent excess [MPE], P < .01) for [1-13C] acetate compared with [2-13C] acetate, respectively, at the end of the infusions. Glutamine isotopic enrichment was slightly but not significantly higher when infusing [1-13C] acetate versus [2-13C] acetate (0.348+/-0.038 v0.495+/-0.069 MPE, NS, respectively). At the end of the experiment, the recovery of 13CO2 from [1-13C] acetate was 44.8%+/-2.7%, and from [2-13C] acetate, 22.6%+/-1.3%. A significant correlation was observed between the differences in 13C enrichment of CO2 for the two tracers and glucose (deltaCO2=0.424 x deltaglucose + 0.001, R2=.9856, P=.0007) or glutamine (deltaCO2=0.621 x deltaglutamine + 0.004, R2=.9573, P=.0038) during the infusion. These results suggest that (1) although gluconeogenesis appears to be more responsible than glutamine for the differential recovery of [2-13C] versus [1-13C] acetate, other secondary pathways are probably also implicated; and (2) different recovery correction factors should be applied when measuring substrate oxidation with a stable isotope tracer depending on the expected position of 13C in acetyl-CoA.

Acetates↗
PubMed1998

Production rate of acetate during colonic fermentation of lactulose: a stable-isotope study in humans.

BACKGROUND: Breath tests are currently used to qualitatively assess colonic fermentation; no quantitative estimations are available for healthy subjects. OBJECTIVE: This study describes a stable-isotope-dilution method to measure acetate production quantitatively from colonic bacterial fermentation. DESIGN: Six volunteers received a primed, constant, intravenous infusion of [1-13C]acetate at a rate of 1.01 +/- 0.04 micromol x kg(-1) x min(-1) for 7 h. They ingested 20 g pure lactulose after 1 h of the tracer infusion. Expired air and arterialized venous blood were sampled every 15 min. RESULTS: Before lactulose intake, the breath-hydrogen concentration was 7 +/- 2 ppm and the plasma acetate concentration and isotopic enrichment were 141 +/- 14 micromol/L and 14.8 +/- 1.4 moles percent excess, respectively. Whole-body acetate turnover was 6.0 +/- 0.7 micromol x kg(-1) x min(-1). After lactulose ingestion, maximum breath hydrogen and acetate concentrations reached 63 +/- 15 ppm (P = 0.004) and 313 +/- 25 micromol/L (P = 0.002), respectively, whereas [13C]acetate enrichment decreased to 9.9 +/- 1.3 moles percent excess (P = 0.03). Whole-body acetate turnover increased to 9.8 +/- 1.5 micromol x kg(-1) x min(-1) and later decreased almost to baseline values. Colonic lactulose fermentation yielded 140 +/- 12 mmol acetate over 6 h, representing 86% of the production based on stoichiometric equations. CONCLUSION: This new method provides a quantitative estimate of colonic carbohydrate fermentation via evaluation of acetate production.

Acetates↗
PubMed1998

Whole-body, peripheral and intestinal endogenous acetate turnover in dogs using stable isotopes.

Acetate metabolism supplies about 10% of energy requirements in food-deprived nonruminant animals. This study used a stable isotope dilution method to investigate the fate of acetate in 24-h food-deprived dogs free of colonic fermentation. Three dogs received intravenous bolus injections of 40 or 70 micromol/kg of [1-13C] acetate, and carotid blood was then sampled during a 15-min period to estimate the acetate distribution volume. Ten dogs received intravenous [1-13C] acetate infusions of 1.05 +/- 0.02 or 2.10 +/- 0. 10 micromol/(kg.min) for 120 or 200 min after a prime of 200 or 70 micromol/kg, respectively. Cephalic venous and carotid arterial blood were sampled for all dogs, and portal blood for five. Acetate distribution volume was 0.27 +/- 0.16 L/kg (mean +/- SEM). The concentrations of acetate in arterial (144 +/- 17 micromol/L), venous (155 +/- 20 micromol/L) and portal plasma (131 +/- 16 micromol/L) were not significantly different during infusion, whereas isotopic enrichments [mole percent excess (MPE): labeled acetate/all acetate molecules] in portal (1.2 +/- 0.2 MPE) and venous plasma (1.7 +/- 0.3 and 2.6 +/- 0.7 MPE) were lower than in arterial plasma for both infusion rates (4.9 +/- 0.6 and 7.6 +/- 0.8 MPE, respectively, P < 0.005). Whole-body acetate turnover was 24.4 +/- 2.4 micromol/(kg.min). Fractional acetate extractions for forelimb and intestine were 62 +/- 7 and 72 +/- 6%, respectively, and the production for each organ was 0.3 and 1.1 micromol/(kg.min) respectively, similar to that of utilization (P > 0.05). It is concluded that the forelimb and intestine produce and utilize acetate as an energy source in 24-h food-deprived dogs free of colonic fermentation.

Acetates↗
PubMed1998

Estrogen receptor activation function 1 works by binding p160 coactivator proteins.

Estrogen receptor-alpha contains two transactivation functions, a weak constitutive activation function (AF-1) and a hormone-dependent activation function (AF-2). AF-2 works by recruiting a large coactivator complex, composed of one or more p160s, CREB-binding protein (CBP)/p300, and P/CAF (p300 and CBP-associated factor), via direct contacts with the p160s. We report here that independent AF-1 activity also requires p160 contacts. Unlike AF-2, which binds signature NR boxes in the center of the p160 molecule, AF-1 binds to sequences near the p160 C terminus. We propose that the ability of AF-1 and AF-2 to interact with separate surfaces of the same coactivator is important for the ability of these transactivation functions to synergize.

Acetyltransferases↗
PubMed1998

[Modulators of leukocytic functions].

Polymorphonuclear neutrophils (PMN) and monocytes play a role in vascular diseases. Animal experimental models, using deleukocytation or injection of anti-CD11b-anti-CD18 monoclonal antibodies (inhibition of leukocytic adhesion and of interaction with the endothelial cell) have confirmed this role in the ischemia-reperfusion syndrome and in myocardial infarction. In man, increased production of oxygen radicals, PMN release of elastase, increased monocyte formation of tissue factor (TF) and integrins have been noted in coronary artery disease, in chronic arteriopathy of the lower limbs and in association with vascular risk factors such as diabetes. Pharmacological alteration of leukocyte hyperactivity therefore seems to be justified. Pentoxifylline, used with good effect in arteriopathy of the lower limbs, affects numerous leukocytic functions: diminution in adherence and in PMN production of free radicals, diminution in the formation of TF and cytokines (TNF). Gingkolides reduce leukocytic interactions and platelet activation through an anti-PAF (Platelet Activation Factor) action. Aspirin may interfere with free radicals and inhibit TF formation. alpha-tocopherol blocks the activation, by free radicals, of the transcription factor NF k B. By altering the TNF and IL-1 cytokines, leukocytic activation can be controlled. Other cytokines (IL-4, IL-10) have an immunosuppressive action and reduce the formation of TF. The pharmacological targets are therefore multiple. Their use in vascular diseases is only at a very preliminary stage.

Animals↗