Signal transduction and activation of the NADPH oxidase in eosinophils

Activation of the eosinophil NADPH oxidase and the subsequent release of toxic oxygen radicals has been implicated in the mechanism of parasite killing and inflammation. At present, little is known of the signal transduction pathway that govern agonist-induced activation of the respiratory burst and is the subject of this review. In particular, we focus on the ability of leukotrine B4 to activate the NADPH oxidase in guinea-pig peritoneal eosinophils which can be obtained in sufficient number and purity for detailed biochemical experiments to be performed.

Expression and function of beta1 integrins on human eosinophils

Eosinophils preferentially accumulate at sites of chronic allergic diseases such as bronchial asthma. The mechanisms by which selective eosinophil migration occurs are not fully understood. However, interactions of cell-surface adhesion molecules on the eosinophil with molecular counterligands on endothelial and epithelial cells, and on extracellular matrix proteins, are likely to be critical during the recruitment process. One possible mechanism for selective eosinophil recruitment involves the alpha4beta 1 (VLA-4) integrin which is not expressed on neutrophils. Correlations have been found between infiltration of eosinophils and endothelial expression of VCAM-1, the ligand for VLA-4, in the lungs of asthmatic individuals as well as in late phase reactions in the lungs, nose and skin. Epithelial and endothelial cells respond to the Th2-type cytokines IL-4 and IL-13 with selective de novo expression of VCAM-1, consistent with the possible role of VCAM-1/VLA-4 interactions in eosinophil influx during allergic inflammation. Both beta 1 and beta 2 integrins on eosinophils exist in a state of partial activation. For example, eosinophils can be maximally activated for adhesion to VCAM-1 or fibronectin after exposure to beta 1 integrin-activating antibodies or divalent cations, conditions that do not necessarily affect the total cell surface expression of beta 1 integrins. In contrast, cytokines like IL-5 prevent beta 1 integrin activation while promoting beta 2 integrin function. Furthermore, ligation of integrins can regulate the effector functions of the cell. For example, eosinophil adhesion via beta 1 and/or beta 2 integrins has been shown to alter a variety of functional responses including degranulation and apoptosis. Thus, integrins appear to be important in mediating eosinophil migration and activation in allergic inflammation. Strategies that interfere with these processes may prove to be useful for treatment of allergic diseases.

Human eosinophil-lymphocyte interactions

While the eosinophil's effector functions clearly can contribute to the pathogenesis of allergic diseases, the evolutionary benefit to having eosinophils as a distinct class of leukocytes is not clear, especially if one must reconsider the nominally beneficial role of eosinophils in parasite host defense. Eosinophils are equipped to respond to lymphocytes and their cytokines (and not solely the eosinophil growth factor cytokines), but the functional consequences of such eosinophil responses need to be defined. Conversely, eosinophils, as antigen-presenting cells (APCs) or sources of lymphocyte-active cytokines, may stimulate and effect lymphocyte functioning. Eosinophils share with CD4+ lymphocytes expression of a number of receptors, including CD4 and IL-2R, and specific alpha4 integrins that may help in their common recruitment and activation. Further, elucidation of the interactions between lymphocytes and eosinophils will contribute to a broader understanding of the functioning of eosinophils in "normal" ongoing immune responses and in allergic disorders.

Divergent functions of three Candida albicans zinc-cluster transcription factors (CTA4, ASG1 and CTF1) complementing pleiotropic drug resistance in Saccharomyces cerevisiae.

One of the mediators of pleiotropic drug resistance in Saccharomyces cerevisiae is the ABC-transporter gene PDR5. This gene is regulated by at least two transcription factors with Zn(2)-Cys(6) finger DNA-binding motifs, Pdr1p and Pdr3p. In this work, we searched for functional homologues of these transcription factors in Candida albicans. A C. albicans gene library was screened in a S. cerevisiae mutant lacking PDR1 and PDR3 and clones resistant to azole antifungals were isolated. From these clones, three genes responsible for azole resistance were identified. These genes (CTA4, ASG1 and CTF1) encode proteins with Zn(2)-Cys(6)-type zinc finger motifs in their N-terminal domains. The C. albicans genes expressed in S. cerevisiae could activate the transcription of a PDR5-lacZ reporter system and this reporter activity was PDRE-dependent. They could also confer resistance to azoles in a S. cerevisiae strain lacking PDR1, PDR3 and PDR5, suggesting that CTA4-, ASG1- and CTF1-dependent azole resistance can be caused by genes other than PDR5 in S. cerevisiae. Deletion of CTA4, ASG1 and CTF1 in C. albicans had no effect on fluconazole susceptibility and did not alter the expression of the ABC-transporter genes CDR1 and CDR2 or the major facilitator gene MDR1, which encode multidrug transporters known as mediators of azole resistance in C. albicans. However, additional phenotypic screening tests on the C. albicans mutants revealed that the presence of ASG1 was necessary to sustain growth on non-fermentative carbon sources (sodium acetate, acetic acid, ethanol). In conclusion, C. albicans possesses functional homologues of the S. cerevisiae Pdr1p and Pdr3p transcription factors; however, their properties in C. albicans have been rewired to other functions.

Lufaxin, a novel factor Xa inhibitor from the salivary gland of the sand fly Lutzomyia longipalpis blocks protease-activated receptor 2 activation and inhibits inflammation and thrombosis in vivo.

OBJECTIVE: Blood-sucking arthropods' salivary glands contain a remarkable diversity of antihemostatics. The aim of the present study was to identify the unique salivary anticoagulant of the sand fly Lutzomyia longipalpis, which remained elusive for decades. METHODS AND RESULTS: Several L. longipalpis salivary proteins were expressed in human embryonic kidney 293 cells and screened for inhibition of blood coagulation. A novel 32.4-kDa molecule, named Lufaxin, was identified as a slow, tight, noncompetitive, and reversible inhibitor of factor Xa (FXa). Notably, Lufaxin's primary sequence does not share similarity to any physiological or salivary inhibitors of coagulation reported to date. Lufaxin is specific for FXa and does not interact with FX, Dansyl-Glu-Gly-Arg-FXa, or 15 other enzymes. In addition, Lufaxin blocks prothrombinase and increases both prothrombin time and activated partial thromboplastin time. Surface plasmon resonance experiments revealed that FXa binds Lufaxin with an equilibrium constant ≈3 nM, and isothermal titration calorimetry determined a stoichiometry of 1:1. Lufaxin also prevents protease-activated receptor 2 activation by FXa in the MDA-MB-231 cell line and abrogates edema formation triggered by injection of FXa in the paw of mice. Moreover, Lufaxin prevents FeCl(3)-induced carotid artery thrombus formation and prolongs activated partial thromboplastin time ex vivo, implying that it works as an anticoagulant in vivo. Finally, salivary gland of sand flies was found to inhibit FXa and to interact with the enzyme. CONCLUSIONS: Lufaxin belongs to a novel family of slow-tight FXa inhibitors, which display antithrombotic and anti-inflammatory activities. It is a useful tool to understand FXa structural features and its role in prohemostatic and proinflammatory events.

Inhibition of glial cell proinflammatory activities by peroxisome proliferator-activated receptor gamma agonist confers partial protection during antimyelin oligodendrocyte glycoprotein demyelination in vitro.

Peroxisome proliferator-activated receptor gamma (PPAR-gamma) is a member of the nuclear hormone superfamily originally characterized as a regulator of adipocyte differentiation and lipid metabolism. In addition, PPAR-gamma has important immunomodulatory functions. If the effect of PPAR-gamma's activation in T-cell-mediated demyelination has been recently demonstrated, nothing is known about the role of PPAR-gamma in antibody-induced demyelination in the absence of T-cell interactions and monocyte/macrophage activation. Therefore, we investigated PPAR-gamma's involvement by using an in vitro model of inflammatory demyelination in three-dimensional aggregating rat brain cell cultures. We found that PPAR-gamma was not constitutively expressed in these cultures but was strongly up-regulated following demyelination mediated by antibodies directed against myelin oligodendrocyte glycoprotein (MOG) in the presence of complement. Pioglitazone, a selective PPAR-gamma agonist, partially protected aggregates from anti-MOG demyelination. Heat shock responses and the expression of the proinflammatory cytokine tumor necrosis factor-alpha were diminished by pioglitazone treatment. Therefore, pioglitazone protection seems to be linked to an inhibition of glial cell proinflammatory activities following anti-MOG induced demyelination. We show that PPAR-gamma agonists act not only on T cells but also on antibody-mediated demyelination. This may represent a significant benefit in treating multiple sclerosis patients.

TRAIL receptors 1 (DR4) and 2 (DR5) signal FADD-dependent apoptosis and activate NF-kappaB.

TRAIL induces apoptosis through two closely related receptors, TRAIL-R1 (DR4) and TRAIL-R2 (DR5). Here we show that TRAIL-R1 can associate with TRAIL-R2, suggesting that TRAIL may signal through heteroreceptor signaling complexes. Both TRAIL receptors bind the adaptor molecules FADD and TRADD, and both death signals are interrupted by a dominant negative form of FADD and by the FLICE-inhibitory protein FLIP. The recruitment of TRADD may explain the potent activation of NF-kappaB observed by TRAIL receptors. Thus, TRAIL receptors can signal both death and gene transcription, functions reminiscent of those of TNFR1 and TRAMP, two other members of the death receptor family.

Factors determining the spontaneous activation of splenic dendritic cells in culture.

Dendritic cells (DCs) serve as a link between the innate and adaptive immune systems. The activation state of DCs is crucial in this role. However, when DCs are isolated from lymphoid tissues, purified and placed in culture they undergo 'spontaneous' activation. The basis of this was explored, using up-regulation of DC surface MHC II, CD40, CD80 and CD86 as indicators of DC activation. No evidence was found for DC damage during isolation or for microbial products causing the activation. The culture activation of spleen DCs differed from that of Langerhans cells when released from E-cadherin-mediated adhesions, since E-cadherin was not detected and activation still occurred with β-catenin null DCs. Much of the activation could be attributed to DC-DC interactions. Although increases in surface MHC II levels occurred under all culture conditions tested, the increase in expression of CD40, CD80 and CD86 was much less under culture conditions where such interactions were minimised. DC-to-DC contact under the artificial conditions of high DC concentration in culture induced the production of soluble factors and these, in turn, induced the up-regulation of co-stimulatory molecules on the DC surface.

Green vs. Lempert functions: a minimal example

The Lempert function for a set of poles in a domain of Cn at a point z is obtained by taking a certain infimum over all analytic disks going through the poles and the point z, and majorizes the corresponding multi-pole pluricomplex Green function. Coman proved that both coincide in the case of sets of two poles in the unit ball. We give an example of a set of three poles in the unit ball where this equality fails.

On the system of the functions x (s) / (s-r)k

"Vegeu el resum a l'inici del document del fitxer adjunt."

The spleen is an important site of T cell activation during human hepatosplenic schistosomiasis

We have undertaken a comparative immunephenotypic study of spleen cells from hepatosplenic patients (HS) and uninfected individuals (NOR) using flow cytometry. Our data did not show any significant differences in the mean percentage of T-cells and B-cells between the two groups. Analysis of activated T-cells demonstrated that HS present an increased percentage of CD3+HLA-DR+ splenocytes in comparison to NOR. Analysis of T-cell subsets demonstrated a significant increase on the percentage of both activated CD4+ T-splenocytes and CD8+ cells in HS. We did not find any difference in the mean percentage of CD28+ T-cells. Analysis of the B-cell compartment did not show any difference on the percentage of B1-splenocytes. However, the spleen seems to be an important reservoir/source for B1 lymphocytes during hepatosplenic disease, since after splenectomy we found a decreased the percentage of circulating B1-lymphocytes. We observed an increase on the percentage of CD2+CD3- lymphocytes in the spleen of HS suggesting that the loss of CD3 by activated T-cells or the expansion of NK-cells might play a role in the development/maintenance of splenomegaly.

Activation of the heat shock response in plants by chlorophenols: transgenic Physcomitrella patens as a sensitive biosensor for organic pollutants.

The ability to detect early molecular responses to various chemicals is central to the understanding of biological impact of pollutants in a context of varying environmental cues. To monitor stress responses in a model plant, we used transgenic moss Physcomitrella patens expressing the beta-glucuronidase reporter (GUS) under the control of the stress-inducible promoter hsp17.3B. Following exposure to pollutants from the dye and paper industry, GUS activity was measured by monitoring a fluorescent product. Chlorophenols, heavy metals and sulphonated anthraquinones were found to specifically activate the hsp17.3B promoter (within hours) in correlation with long-term toxicity effects (within days). At mildly elevated physiological temperatures, the chemical activation of this promoter was strongly amplified, which considerably increased the sensitivity of the bioassay. Together with the activation of hsp17.3B promoter, chlorophenols induced endogenous chaperones that transiently protected a recombinant thermolabile luciferase (LUC) from severe heat denaturation. This sensitive bioassay provides an early warning molecular sensor to industrial pollutants under varying environments, in anticipation to long-term toxic effects in plants. Because of the strong cross-talk between abiotic and chemical stresses that we find, this P. patens line is more likely to serve as a direct toxicity bioassay for pollutants combined with environmental cues, than as an indicator of absolute toxicity thresholds for various pollutants. It is also a powerful tool to study the role of heat shock proteins (HSPs) in plants exposed to combined chemical and environmental stresses.

Orthogonal higher order structure and confirmatory factor analysis of the French Wechsler Adult Intelligence Scale (WAIS-III).

According to the most widely accepted Cattell-Horn-Carroll (CHC) model of intelligence measurement, each subtest score of the Wechsler Intelligence Scale for Adults (3rd ed.; WAIS-III) should reflect both 1st- and 2nd-order factors (i.e., 4 or 5 broad abilities and 1 general factor). To disentangle the contribution of each factor, we applied a Schmid-Leiman orthogonalization transformation (SLT) to the standardization data published in the French technical manual for the WAIS-III. Results showed that the general factor accounted for 63% of the common variance and that the specific contributions of the 1st-order factors were weak (4.7%-15.9%). We also addressed this issue by using confirmatory factor analysis. Results indicated that the bifactor model (with 1st-order group and general factors) better fit the data than did the traditional higher order structure. Models based on the CHC framework were also tested. Results indicated that a higher order CHC model showed a better fit than did the classical 4-factor model; however, the WAIS bifactor structure was the most adequate. We recommend that users do not discount the Full Scale IQ when interpreting the index scores of the WAIS-III because the general factor accounts for the bulk of the common variance in the French WAIS-III. The 4 index scores cannot be considered to reflect only broad ability because they include a strong contribution of the general factor.

Effects of a dual inhibitor of angiotensin converting enzyme and neutral endopeptidase, MDL 100,240, on endocrine and renal functions in healthy volunteers.

OBJECTIVE: To investigate the endocrine and renal effects of the dual inhibitor of angiotensin converting enzyme and neutral endopeptidase, MDL 100,240. DESIGN: A randomized, placebo-controlled, crossover study was performed in 12 healthy volunteers. METHODS: MDL 100,240 was administered intravenously over 20 min at single doses of 6.25 and 25 mg in subjects with a sodium intake of 280 (n = 6) or 80 (n = 6) mmol/day. Measurements were taken of supine and standing blood pressure, plasma angiotensin converting enzyme activity, angiotensin II, atrial natriuretic peptide, urinary atrial natriuretic peptide and cyclic GMP excretion, effective renal plasma flow and the glomerular filtration rate as p-aminohippurate and inulin clearances, electrolytes and segmental tubular function by endogenous lithium clearance. RESULTS: Supine systolic blood pressure was consistently decreased by MDL 100,240, particularly after the high dose and during the low-salt intake. Diastolic blood pressure and heart rate did not change. Plasma angiotensin converting enzyme activity decreased rapidly and dose-dependently. In both the high- and the low-salt treatment groups, plasma angiotensin II levels fell and renin activity rose accordingly, while plasma atrial natriuretic peptide levels remained unchanged. In contrast, urinary atrial natriuretic peptide excretion increased dose-dependently under both diets, as did urinary cyclic GMP excretion. Effective renal plasma flow and the glomerular filtration rate did not change. The urinary flow rate increased markedly during the first 2 h following administration of either dose of MDL 100,240 (P < 0.001) and, similarly, sodium excretion tended to increase from 0 to 4 h after the dose (P = 0.07). Potassium excretion remained stable. Proximal and distal fractional sodium reabsorption were not significantly altered by the treatment. Uric acid excretion was increased. The safety and clinical tolerance of MDL 100,240 were good. CONCLUSIONS: The increased fall in blood pressure in normal volunteers together with the preservation of renal hemodynamics and the increased urinary volume, atrial natriuretic peptide and cyclic GMP excretion distinguish MDL 100,240 as a double-enzyme inhibitor from inhibitors of the angiotensin converting enzyme alone. The differences appear to be due, at least in part, to increased renal exposure to atrial natriuretic peptide following neutral endopeptidase blockade.