1,720,976 research outputs found
Intracellular shunting of O2− contributes to charge compensation and preservation of neutrophil respiratory burst in the absence of voltage-gated proton channel activity
No full textProton efflux via voltage-gated proton channels (Hv1) is considered to mediate the charge compensation necessary to preserve NADPH oxidase activity during the respiratory burst. Using the Hv1 inhibitor Zn(2+), we found that the PMA-induced respiratory burst of human neutrophils is inhibited when assessed as extracellular production of O2(-) and H2O2, in accordance with literature studies, but, surprisingly, unaffected when measured as oxygen consumption or total (extracellular plus intracellular) H2O2 production. Furthermore, we show that inhibiting Hv1 with Zn(2+) results in an increased production of intracellular ROS. Similar results, i.e. decreased extracellular and increased intracellular ROS production, were obtained using a human granulocyte-like cell line with severely impaired Hv1 expression. Acidic extracellular pH, which dampens proton efflux, also augmented intracellular production of H2O2. Zinc caused an increase in the rate but not in the extent of depolarization and cytosolic acidification indicating that mechanisms other than proton efflux take part in charge compensation. Our results suggest a hitherto unpredicted mechanism of charge compensation whereby, in the absence of proton efflux, part of O2(-) generated within gp91(phox) in the plasma membrane is shunted intracellularly down electrochemical gradient to dampen excessive depolarization. This would preserve NADPH oxidase activity under conditions such as the inflammatory exudate in which the acidic pH hinders charge compensation by proton efflux
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
LA BACTERICIDIE DES NEUTROPHILES : MECANISMES CELLULAIRES ET MOLECULAIRES ET LEURS DYSFONCTIONS
No full textMicrobicidal activity of neutrophils requires the NADPH oxidase dependent reactive oxygen species (ROS) production and myeloperoxidase (MPO)-H2O2-halide system-catalyzed halogenations (oxidative mechanisms) and the release of granular proteins into the phagosomal lumen (non oxidative mechanisms). The long-established direct role of ROS and MPO has been recently questioned. In this work, using our improved method to assess microbicidal activity, we have re-examined the role of the diverse mechanisms proposed to be involved in the antimicrobial activity of neutrophils. We show that the NADPH oxidase activity is indispensable for the killing of certain microorganisms (e.g. Staphylococcus aureus, Candida albicans) but not of others (e.g. Escherichia coli) which are efficiently killed even in the absence of a respiratory burst. Our data also indicate that NADPH oxidase-dependent killing of S. aureus and C. albicans is largely dependent on MPO. Alkalinization of phagosomal pH and K+ fluxes do not appear to significantly contribute to the killing of these microorganisms. K+ channels activity may account for the partial recovery of killing observed in MPO-deficient neutrophils at long times of incubation.We have also investigated the neutrophils’ microbicidal mechanisms in a cellular model, the human myelomonoblastic cellular line inducible in granulocytes, PLB-985. The ROS production in PLB-985 cells stimulated by opsonized microorganisms is very weak compared to human neutrophils, but sustains a NADPH oxidase dependent killing activity of S. aureus and C. albicans similar to PMN after short times of incubation (<10 min). However, the killing activity of the PLB cells is abolished after long times of incubation. NADPH oxidase independent killing of E. coli is also defective. The ROS overproduction of the mutant DloopNox4-Nox2 (in these cells, the D-loop of Nox2 was replaced by the D-loop of its homolog Nox4) after soluble and particulate agonists dependent activation does not increase their microbicidal power. The weak killing activity of the PLB-985 cellular line can be explained by the faint amounts of cytochrome b558 and by the more or less lack of granular proteins content (MPO, elastase, cathepsine G, lactoferrine, lysozyme, MMP8 and MMP9) involved in the microbicidal event. In conclusion, in PLB-985 DMF-differentiated cells, the oxygen dependent and independent mechanisms are both defective. A lack of NADPH oxidase complex results in a rare inherited disorder, the chronic granulomatose disease (CGD). We characterized an atypical and extremely rare case of CGD: the X91- variant: the new identified point mutation in the CYBB gene promoter (insertion of a T at position -54T/-56T) appeared to be related to the loss of association of ets transcription factors within this region, prevent the normally functional expression of the gene and finally, correlate with the low NADPH oxidase activity. The residual respiratory burst supported no killing of S. aureus and C. albicans, in vitro, and is not sufficient to protect the patient against severe and recurrent infections.L’élimination des microorganismes par les neutrophiles (PMN) est mediée par l’activité des formes réactives de l’oxygène (FRO), produites suite à l’activation de la NADPH oxydase, dont l’efficacité est sensiblement augmentée par la myélopéroxydase, MPO, (mécanismes oxydatifs) et par l’action des protéines bactéricides contenues dans les granules cytoplasmiques (mécanismes non oxydatifs). L’implication directe des FRO et de la MPO dans la bactéricidie des PMN a été récemment remise en question. Nous nous sommes proposés de réexaminer les mécanismes impliqués dans la bactéricidie des neutrophiles sur la base des nouvelles hypothèses récemment proposées en appliquant une méthode de mesure du « killing » des microorganismes que nous avons récemment mise au point et qui permet une estimation plus correcte de la bactéricidie. Nous avons démontré que l’activité NADPH oxydase est indispensable pour le « killing » de certains microorganismes (S. aureus et C. albicans) mais pas pour d’autres (E. coli) qui sont efficacement éliminés même en l’absence du burst respiratoire. Les flux d’ions K+ et l’alcalinisation du pH intraphagosomale, induits par l’activation du complexe oxydase, ne sont pas nécessaires au killing de S. aureus et C. albicans, microorganismes dont la bactéricidie NADPH oxydase dépendante est mediée presque exclusivement par la MPO. Les courants des ions K+, à travers des canaux BKCa-like, semblent être responsables du killing résiduel des PMN MPO déficients, après une longue incubation. Nous avons aussi étudié les mécanismes impliqués dans la bactéricidie des PMN en utilisant comme modèle cellulaire la lignée PLB-985 différentiable en « pseudo-neutrophiles ». Nous avons prouvé que la réponse de ces cellules aux stimuli particulaires est sensiblement plus faible par rapport aux PMN. Ce burst respiratoire support e une bactéricidie comparable à celles des PMN pour des temps courts d’incubation (<10 min) vis-à-vis des microorganismes sensibles aux mécanismes de « killing » oxygène-dépendants (S. aureus et C. albicans); toutefois, en prolongeant les temps d’incubation, le pouvoir bactéricides ce ces cellules est abolit. Le « killing » NADPH oxydase indépendant d’E. coli est aussi partiellement déficitaire. La superproduction de FRO mise en évidence dans les cellules mutantes PLB-985 DloopNox4-Nox2 (où la deuxième boucle intracellulaire de Nox2 a été remplacée par celle de l’oxydase homologue Nox4) en réponse aux stimuli solubles et particulaires n’est pas accompagnée par une augmentation du « killing ». Le faible pouvoir bactéricide de cette lignée cellulaire est du au contenu réduit en cytochrome b558 (un dixième par rapport aux PMN) et aux défauts plus ou moins importants des protéines granulaires (MPO, elastase, cathepsine G, lactoferrine, lysozyme, MMP8 et MMP9) impliquées dans le processus de bactéricidie: dans les cellules PLB-985 différentiées au DMF, les mécanismes bactéricides oxygènes dépendants et indépendants sont donc tous deux compromis.Le dysfonctionnement du complexe NADPH oxydase est à la base d’une maladie génétique rare, la granulomatose septique chronique (CGD). Nous nous sommes intéressés à la caractérisation d’un cas atypique de CGD, la forme X91-. Nous avons démontré que la nouvelle mutation identifiée (insertion d’un T en position -54T/-56T) compromet l’association des facteurs de transcription ets avec la région promotrice, empêche, par conséquence, l’expression normale du gène dans les neutrophiles et détermine une activité oxydase réduite. Ce burst respiratoire résiduel (6% du control) n’est pas suffisante pour soutenir, in vitro, une activité de « killing » vis-à-vis de S. aureus et C. albicans ni pour protéger le patient des infections sévères et récidivantes
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
LA BACTERICIDIE DES NEUTROPHILES : MECANISMES CELLULAIRES ET MOLECULAIRES ET LEURS DYSFONCTIONS
Full text linkMicrobicidal activity of neutrophils requires the NADPH oxidase dependent reactive oxygen species (ROS) production and myeloperoxidase (MPO)-H2O2-halide system-catalyzed halogenations (oxidative mechanisms) and the release of granular proteins into the phagosomal lumen (non oxidative mechanisms). The long-established direct role of ROS and MPO has been recently questioned. In this work, using our improved method to assess microbicidal activity, we have re-examined the role of the diverse mechanisms proposed to be involved in the antimicrobial activity of neutrophils. We show that the NADPH oxidase activity is indispensable for the killing of certain microorganisms (e.g. Staphylococcus aureus, Candida albicans) but not of others (e.g. Escherichia coli) which are efficiently killed even in the absence of a respiratory burst. Our data also indicate that NADPH oxidase-dependent killing of S. aureus and C. albicans is largely dependent on MPO. Alkalinization of phagosomal pH and K+ fluxes do not appear to significantly contribute to the killing of these microorganisms. K+ channels activity may account for the partial recovery of killing observed in MPO-deficient neutrophils at long times of incubation.We have also investigated the neutrophils’ microbicidal mechanisms in a cellular model, the human myelomonoblastic cellular line inducible in granulocytes, PLB-985. The ROS production in PLB-985 cells stimulated by opsonized microorganisms is very weak compared to human neutrophils, but sustains a NADPH oxidase dependent killing activity of S. aureus and C. albicans similar to PMN after short times of incubation (<10 min). However, the killing activity of the PLB cells is abolished after long times of incubation. NADPH oxidase independent killing of E. coli is also defective. The ROS overproduction of the mutant DloopNox4-Nox2 (in these cells, the D-loop of Nox2 was replaced by the D-loop of its homolog Nox4) after soluble and particulate agonists dependent activation does not increase their microbicidal power. The weak killing activity of the PLB-985 cellular line can be explained by the faint amounts of cytochrome b558 and by the more or less lack of granular proteins content (MPO, elastase, cathepsine G, lactoferrine, lysozyme, MMP8 and MMP9) involved in the microbicidal event. In conclusion, in PLB-985 DMF-differentiated cells, the oxygen dependent and independent mechanisms are both defective. A lack of NADPH oxidase complex results in a rare inherited disorder, the chronic granulomatose disease (CGD). We characterized an atypical and extremely rare case of CGD: the X91- variant: the new identified point mutation in the CYBB gene promoter (insertion of a T at position -54T/-56T) appeared to be related to the loss of association of ets transcription factors within this region, prevent the normally functional expression of the gene and finally, correlate with the low NADPH oxidase activity. The residual respiratory burst supported no killing of S. aureus and C. albicans, in vitro, and is not sufficient to protect the patient against severe and recurrent infections.L’élimination des microorganismes par les neutrophiles (PMN) est mediée par l’activité des formes réactives de l’oxygène (FRO), produites suite à l’activation de la NADPH oxydase, dont l’efficacité est sensiblement augmentée par la myélopéroxydase, MPO, (mécanismes oxydatifs) et par l’action des protéines bactéricides contenues dans les granules cytoplasmiques (mécanismes non oxydatifs). L’implication directe des FRO et de la MPO dans la bactéricidie des PMN a été récemment remise en question. Nous nous sommes proposés de réexaminer les mécanismes impliqués dans la bactéricidie des neutrophiles sur la base des nouvelles hypothèses récemment proposées en appliquant une méthode de mesure du « killing » des microorganismes que nous avons récemment mise au point et qui permet une estimation plus correcte de la bactéricidie. Nous avons démontré que l’activité NADPH oxydase est indispensable pour le « killing » de certains microorganismes (S. aureus et C. albicans) mais pas pour d’autres (E. coli) qui sont efficacement éliminés même en l’absence du burst respiratoire. Les flux d’ions K+ et l’alcalinisation du pH intraphagosomale, induits par l’activation du complexe oxydase, ne sont pas nécessaires au killing de S. aureus et C. albicans, microorganismes dont la bactéricidie NADPH oxydase dépendante est mediée presque exclusivement par la MPO. Les courants des ions K+, à travers des canaux BKCa-like, semblent être responsables du killing résiduel des PMN MPO déficients, après une longue incubation. Nous avons aussi étudié les mécanismes impliqués dans la bactéricidie des PMN en utilisant comme modèle cellulaire la lignée PLB-985 différentiable en « pseudo-neutrophiles ». Nous avons prouvé que la réponse de ces cellules aux stimuli particulaires est sensiblement plus faible par rapport aux PMN. Ce burst respiratoire support e une bactéricidie comparable à celles des PMN pour des temps courts d’incubation (<10 min) vis-à-vis des microorganismes sensibles aux mécanismes de « killing » oxygène-dépendants (S. aureus et C. albicans); toutefois, en prolongeant les temps d’incubation, le pouvoir bactéricides ce ces cellules est abolit. Le « killing » NADPH oxydase indépendant d’E. coli est aussi partiellement déficitaire. La superproduction de FRO mise en évidence dans les cellules mutantes PLB-985 DloopNox4-Nox2 (où la deuxième boucle intracellulaire de Nox2 a été remplacée par celle de l’oxydase homologue Nox4) en réponse aux stimuli solubles et particulaires n’est pas accompagnée par une augmentation du « killing ». Le faible pouvoir bactéricide de cette lignée cellulaire est du au contenu réduit en cytochrome b558 (un dixième par rapport aux PMN) et aux défauts plus ou moins importants des protéines granulaires (MPO, elastase, cathepsine G, lactoferrine, lysozyme, MMP8 et MMP9) impliquées dans le processus de bactéricidie: dans les cellules PLB-985 différentiées au DMF, les mécanismes bactéricides oxygènes dépendants et indépendants sont donc tous deux compromis.Le dysfonctionnement du complexe NADPH oxydase est à la base d’une maladie génétique rare, la granulomatose septique chronique (CGD). Nous nous sommes intéressés à la caractérisation d’un cas atypique de CGD, la forme X91-. Nous avons démontré que la nouvelle mutation identifiée (insertion d’un T en position -54T/-56T) compromet l’association des facteurs de transcription ets avec la région promotrice, empêche, par conséquence, l’expression normale du gène dans les neutrophiles et détermine une activité oxydase réduite. Ce burst respiratoire résiduel (6% du control) n’est pas suffisante pour soutenir, in vitro, une activité de « killing » vis-à-vis de S. aureus et C. albicans ni pour protéger le patient des infections sévères et récidivantes
La bactéricidie des neutrophiles: mécanismes cellulaires et moléculaires et leurs dysfonctions
No full text2008/2009In questa tesi ci siamo proposti di riesaminare i meccanismi implicati nell’attività microbicida dei neutrofili, alla luce delle nuove ipotesi proposte recentemente, impiegando una metodica corretta di misura del killing microbico da noi recentemente messa a punto. Nel complesso, i risultati da noi ottenuti provano che l’attività NADPH ossidasica è indispensabile per il killing di certi microrganismi (quali S. aureus e C. albicans) ma non per altri (quali E. coli) che sono uccisi efficacemente anche in assenza di burst respiratorio. I flussi di ioni potassio e l’alcalinizzazione del pH intrafagosomale, indotti dall’attivazione del complesso ossidasi, non sono necessari al killing di S. aureus e C. albicans, il cui killing NADPH ossidasi-dipendente è mediato quasi esclsivamente dalla mieloperossidasi. Le correnti di ioni K+ sembrano responsabili dell’attività microbicida residua dei neutrofili MPO-deficienti, a lunghi tempi di incubazione.
I meccanismi implicati nell’attività microbicida dei neutrofili sono stati studiati anche ricorrendo ad un modello sperimentale ampiamente utilizzato in letteratura per le ricerche sul funzionamento del complesso NADPH ossidasi: la linea cellulare PLB-985 differenziabile in neutrophils-like. In questo contesto, i risultati da noi ottenuti suggeriscono che la produzione di ROS da parte delle linea cellulare PLB-985 in risposta a stimoli particolati (microrganismi opsonizzati) è sensibilmente più debole rispetto ai PMN. Tale burst respiratorio è responsabile di un’attività microbicida, nei confronti di microrganismi sensibili ai meccanismi di killing ossigeno-dipendenti (S. aureus e C. albicans), paragonabile a quella dei PMN a tempi brevi di incubazione; prolungando i tempi di fagocitosi, il potere microbicida di queste cellule risulta invece abolito. Nei confronti di E. coli, battere sensibile ai processi di killing ossigeno-indipendenti, l’attività microbicida delle cellule PLB-985 risulta parzialmente difettosa se paragonata ai neutrofili La super-produzione di anioni superossido, evidenziata nelle cellule PLB-985 DloopNox4-Nox2 (cellule in cui la seconda ansa intracellulare di No2 é sostituita da quella dell’ossidasi omologa Nox4) in risposta a stimoli solubili e particolati, non è accompagnata da un’aumentata capacità microbicida di queste cellule nei confronti di S. aureus, C. albicans ed E. coli. Nelle cellule PLB-985 i processi microbicidi non ossidativi risultano compromessi, a causa della carenza parziale (MPO, elastasi, catepsina G, β-glucuronidasi, CD11b) o totale (lattoferrina, lisozima, MMP-8, MMP-9) degli enzimi e delle proteine granulari coinvolti nell’attività antimicrobica e a causa della loro difettosa esocitosi.. In particolare, i nostri risultati suggeriscono un difetto a livello della biogenesi dei granuli specifici e terziari.
Il ridotto funzionamento del complesso enzimatico NADPH ossidasi è alla base di una rara malattia genetica, la malattia granulomatosa cronica. Parte di questo lavoro di tesi è stata dedicata alla caratterizzazione di un caso atipico di CGD, la forma X91-. I risultati ottenuti ci hanno permesso di dimostrare che la nuova mutazione da noi descritta nel promotore del gene CYBB (inserzione di una T nella posizione da -54T a -57T) é responsabile della ridotta associazione dei fattori di trascrizione ets con la regione promotrice, impedisce una normale espressione del gene nei neutrofili dei pazienti ed é correlata alla ridotta attività ossidasica misurata in tutta la popolazione di granulociti; l’espressione e l’attività funzionale della NADPH ossidasi appaiono invece normali negli eosinofili, suggerendo che l’espressione di gp91phox in queste cellule é regolata da fattori di trascrizione differenti da quelli operanti nei neutrofili. Infine, l’attività ossidasica residua (5-7% del normale) non é di per sé sufficiente a garantire un’attività di killing nei confronti di S. aureus e C. albicans e a proteggere il paziente contro le infezioni .XXI Cicl
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