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Opposite impact of inflammation and oxidative stress on vascular GGT activity
No full textPurpose. Gamma-glutamyltransferase (GGT) is a cell surface enzyme from the Meister cycle that catalyzes the first step in the catabolism of glutathione (GSH) and its derivative S-nitroso-glutathione (GSNO), thus allowing for both the recovery of precursor aminoacids and – with regard to GSNO – the release of nitric oxide (NO). Recent studies indicate that increased levels of GGT activity in the serum are correlated with increased cardiovascular morbidity/mortality, including atherosclerosis or hypertension, two contexts of vascular wall inflammation and oxidative stress [1]. However lesser is known on GGT activity inside the vascular wall. Our aim was to evaluate GGT activity in cells and tissue from the vascular wall in several models of inflammation and oxidative stress.
Methods. GGT activity (quantified using L-γ-glutamyl-p-nitroanilide) and intracellular GSH concentrations (measured by 2,3-naphtalene dicarboxaldeide) were measured in (i) smooth muscle cells isolated from rat aortic wall (A10 line) stimulated either with LPS (20 μg/ml, 24 h 37°C) or 2,2'-azobios(2-amidinopropane) dihydrochloride (AAPH; 50mM, 2 h 37°C) to mimic inflammation or oxidative stress respectively, and (ii) in aorta homogenates from male Spontaneously Hypertensive Rats (SHR) compared to their counterparts WKY normotensive Wistar Kyoto rats. In human atherosclerotic carotid plaques, GGT expression was assessed by SDS-PAGE analysis in homogenates, using macrophages infiltration as marker of inflammation.
Results. In human atherosclerotic plaques homogenates, the presence of a high-molecular weight GGT similar to that expressed by monocytes/macrophage was observed, with the highest GGT expression in presence of high vs. low macrophage infiltration (score 2: 5-10%, 3: >10% vs. 1: <0.5%). GGT activity increased in the cultured cell model of inflammation (Table 1), while it decreased in the cell and tissue models of oxidative stress in parallel with the decrease in GSH content.
Conclusion. We planned to further evaluate the possible modulation between activated macrophages and promotion of pro-atherogenic process in SMC. Future experiments may also help to fully elucidate the mechanisms of GGT release and/or activity and its role during inflammation and/or oxidative stress.
Keywords: Gamma-glutamyltransferase, inflammation, oxidative stress, atherosclerosis, smooth muscle cells
Bibliography:
1. Pompella A, Emdin M, Passino C, Paolicchi A. The significance of serum gamma-glutamyltransferase in cardiovascular diseases. Clin. Chem. Lab. Med., 2004, 42: 1085-91
Opposite impact of inflammation and oxidative stress on vascular GGT activity
No full textOpposite impact of inflammation and oxidative stress on vascular GGT activity
Endothelial gamma-glutamyltransferase contributes to the vasorelaxant effect of S-nitrosoglutathione in rat aorta
No full textS-nitrosoglutathione (GSNO) involved in storage and transport of nitric oxide (•NO), plays an important role in vascular homeostasis. Breakdown of GSNO can be catalyzed by γ−glutamyltransferase (GGT). We investigated whether vascular GGT influences the vasorelaxant effect of GSNO in isolated rat aorta.
Histochemical localization of GGT and its activity measurement were performed by using chromogenic substrates in sections and in aorta homogenates, respectively. The role of GGT in GSNO metabolism was evaluated by measuring GSNO consumption rate (absorbance decay at 334 nm), •NO release was visualized and quantified with the fluorescent probe 4,5-diaminofluorescein diacetate. The vasorelaxant effect of GSNO was assayed using isolated rat aortic rings (in the presence or absence of endothelium). In each experiment, the role of GGT was assessed using a γ-glutamyl acceptor, glycylglycine, and a non-competitive inhibitor of GGT, serine borate complex.
Specific GGT activity was histochemically localized in the endothelium. Consumption of GSNO and release of free •NO decreased and increased in presence of serine borate complex and glycylglycine, respectively. In endothelium-intact aorta, half maximal effective concentration (EC50) for GSNO (3.2 ± 0.5.10-7 M) was increased in the presence of serine borate complex serine borate complex (1.6 ± 0.2.10-6 M) and decreased with glycylglycine (4.7 ± 0.9.10-8 M). In endothelium-denuded aorta, EC50 for GSNO alone increased to 2.3 ± 0.3.10-6 M, with no change in the presence of serine borate complex.
These data demonstrate the important role of endothelial GGT activity in mediating the vasorelaxant effect of GSNO in rat aorta under physiological conditions. Because new therapeutics treatments based on GSNO are actually developed, this endothelium-dependent mechanism involved in the vascular effects of GSNO should be taken into account in a pharmacological perspective
Opposite impact of inflammation and oxidative stress on vascular GGT activity
No full textINTRODUCTION. Gamma-glutamyltransferase (GGT) is involved in the catabolism of glutathione (GSH) and S-nitroso-glutathione (GSNO), thus allowing the recovery of precursor aminoacids and the release of nitric oxide. Recent studies indicate that increased serum levels of GGT activity are correlated with increased cardiovascular morbidity/mortality, including atherosclerosis or hypertension, two contexts of vascular wall inflammation and oxidative stress [1]. However lesser is known on GGT activity inside the vascular wall. Our aim was to evaluate GGT activity in cells and tissue from the vascular wall in several models of inflammation and oxidative stress.
MATERIAL&METHODS. GGT activity (quantified using L-γ-glutamyl-p-nitroanilide) and intracellular GSH concentrations (measured by 2,3-naphtalene dicarboxaldeide) were measured in (i) a rat smooth muscle cells line (SMC A-10) stimulated either with lipopolysaccharide (LPS; 20 μg/ml, 24 h 37°C) or 2,2'-azobios(2-amidinopropane) dihydrochloride (AAPH; 50 mM, 2 h 37°C) to mimic inflammation or oxidative stress respectively, and (ii) in aorta homogenates from male Spontaneously Hypertensive Rats (SHR) compared to normotensive Wistar Kyoto rats (WKY). In homogenates of human atherosclerotic carotid plaques, GGT expression was assessed by SDS-PAGE analysis, using macrophages infiltration as marker of inflammation.
RESULTS. In human atherosclerotic plaques homogenates, the presence of a high-molecular weight GGT similar to that expressed by macrophage was observed, with the highest GGT expression in presence of high vs. low macrophage infiltration (score 2: 5-10%, 3: >10% vs. 1: <0.5%). GGT activity increased in the cell model of inflammation (Table 1), while it decreased in the cell and tissue models of oxidative stress in parallel with the decrease in GSH content.
CONCLUSION. We planned to further evaluate the possible modulation between activated macrophages and promotion of pro-atherogenic process in SMC. Future experiments may also help to fully elucidate the mechanisms of GGT release and/or activity and its role during inflammation and/or oxidative stress
Oxidative stress enhances and modulates protein S-nitrosation in smooth muscle cells exposed to S-nitrosoglutathione
No full textBackground & Aims: Reduced availability or depletion of nitric oxide (NO) is often involved in pathogenesis and/or progression of cardiovascular diseases, such as atherosclerosis, thrombosis, pulmonary hypertension, ischemia and arrhythmia. Several NOrelated therapeutics have been developed to overcome the problem, such as e.g. organic nitrates, metal—NO complexes, nitrosamines etc., but all have relatively short half-lives and produce tolerance phenomena and oxidant stress. These drawbacks are absent in S-nitrosothiols (RSNOs), i.e. molecules in which NO is reversibly bound to SH groups, and in particular, S-nitrosoglutathione (GSNO) – the endogenous/physiological storage and transport form of NO – is under active investigation. However, the ability of GSNO to regulate NO bioavailability under oxidative stress is poorly studied. Using protein S-nitrosation (Pr-SNO), a post-translational protein modification, as a biomarker of the NO pool (1), the study aims to evaluated the capacity of GSNO to deliver NO to smooth muscle cells (SMCs) under oxidative stress. Furthermore, the implication of redox enzymes implicated in GSNO metabolism, such as gamma-glutamyl transferase (GGT) and protein disulfide isomerase (PDI), in Pr-SNO formation under oxidative stress will be assessed.
Experimental: Rat aorta embryonic SMCs (A-10 cell line) were exposed in vitro to a free radical generator, AAPH, as an oxidative
stress model. Oxidative stress impact on the expression/activity of redox enzymes implicated in GSNO metabolism, as well as NO release were evaluated. The intracellular thiol redox status was also monitored in relation with the extent and distribution of GSNO induced intracellular proteins S-nitrosation (mass spectrometry (LC-MALDI) analysis).
Results: Under oxidative stress, GSNO-metabolizing enzymes were differentially modulated: GGT (activity) was in fact decreased by 3.5-fold, while PDI (expression) was increased by 1.7- fold. Oxidative stress produced the increase of extracellular GSNO-catabolism into nitrite ions as well as an increase in seric proteins S-nitrosation. Moreover, oxidative stress caused both a decrease of SH groups in cellular proteins and an efflux of intracellular GSH to the extracellular space. However, only the first phenomenon was prevented by concomitant administration of GSNO. In agreement with the increased NO release, GSNO-dependent protein S-nitrosation was approx. 2-fold increased under oxidative stress. Experiments with GGT inhibitor serine-borate complex (SBC) and PDI inhibitor bacitracin confirmed that even oxidative stress modified their activity/localization, both enzymes participated in GSNO catabolism
and subsequent Pr-SNO. LC-MALDI analysis revealed that the number of proteins S-nitrosated by GSNO was increased under oxidative stress (51 proteins, vs. 32 in basal condition). Compared to basal condition, oxidative stress promoted the S-nitrosation of three additional classes of proteins, implicated in cell adhesion, transfer/carrier functions and cellular transport, and in particular favored the S-nitrosation of a higher number of cytoskeletal proteins (17 vs. 8 in basal condition).
Conclusions: Data obtained so far confirmed that oxidative stress such as those occurring in inflammation can modify the activity and/or expression of two critical enzymes in GSNO metabolism, GGT and PDI. Overall, oxidative stress induced higher levels of GSNO-dependent NO release and RSNOs formation. Protein S-nitrosation effected by GSNO under oxidative stress was more extensive, due to the involvement of additional proteins of SMCs cytoskeleton and contractile machinery. Further studies will likely elucidate the pathophysiological significance of these observation
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
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