262,268 research outputs found
S-nitrosoglutathione potentiates protein S-nitrosation under oxidative stress, a potential improvement of NO storage into smooth muscle cells
No full textIntroduction To counteract NO deficiency occurring with oxidative stress (OS) in cardiovascular diseases, administration of S-nitrosothiols (RSNO) like S-nitrosoglutathione (GSNO), the main storage form of NO in tissues [1], represents an alternative to other NO-donors, with no tolerance nor OS induction. However, their ability to regulate NO bioavailability under OS is unknown. As S-nitrosation of proteins, the formation of high molecular weight RSNOs, is also considered as a form of NO storage in tissues [2], we evaluated whether an administration of GSNO will regulate protein S-nitrosation in an OS model of rat smooth muscle cells. Material and methods A rat smooth muscle cell line (SMC A-10) was stimulated by 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH; 50 mM, 2 h, 37°C) to mimic OS. Intracellular thiol status as content of reduced glutathione (GSH) (2,3-naphthalene dicarboxaldehyde assay) and reduced thiol containing proteins (Ellman’s method) were monitored. The activity of γ-glutamyl transpeptidase (GGT), specifically implied in GSNO catabolism, was measured using L-γ-glutamyl-p-nitroanilide as chromogenic substrate. Then, the thiol status modifications and intracellular peptides/proteins S-nitrosation (2,3-diaminonaphthalene/Hg2+assay) were monitored in stressed SMC incubated for 1 h with 50 μM GSNO. S-nitrosated proteins were purified (biotin switch technique) and identified by mass spectrometry. Results Under OS, the intracellular content of reduced thiols was greatly decreased for GSH (59±4 to 29 ± 5 nmol/mg proteins, n = 3) compared to proteins (148 ± 6 to 125 ± 4 nmol/mg proteins, n = 3), with no impact of GSNO. However, GSNO increased the global content of intracellular S-nitrosated peptides/proteins upon OS (0.53 ± 0.04 to 1.07±0.09 nmol/mg proteins, n = 3). Although the GGT activity decreased (1.35 ± 0.20 to 0.39±0.14 nmol/min/mg proteins) under OS, it was still implied at 38±5% (using serine borate complex, a GGT specific inhibitor) into the intracellular peptides/proteins S-nitrosation. The final mass spectrometry identification revealed that 71 proteins were S-nitrosated under control condition and this rose to 93 under OS. Discussion/conclusion The increase in intracellular S-nitrosated proteins in smooth muscle cells submitted to OS and treated with GSNO can be the starting point for GSNO to restore the NO pool. How and when this NO pool can be released has to be further evaluated. References 1. Maron BA et al. Antioxid Redox Signal (2013) 18, 270-287. 2. Rayner BS et al. J Biol Chem (2005) 280, 9985-9993
S-nitrosoglutathione potentiates protein S-nitrosation under oxidative stress, a potential improvement of NO storage into smooth muscle cells
No full textCardiovascular diseases are associated with oxidative stress and reduced nitric oxide (NO) bioavailability. The ability of NO donors like S-nitrosoglutathione (GSNO) to regulate NO bioavailability under oxidative stress is poorly studied. Here, we monitored protein S-nitrosation (Pr-SNO), a post-translational protein modification in smooth muscle cells exposed to GSNO under oxidative stress. Intracellular thiol redox status in relation with the extent and distribution of GSNO-induced intracellular Pr-SNO (LC-MALDI MS) were assessed. The role of the gammaglutamyl transferase (GGT), a redox enzyme metabolizing GSNO, in Pr-SNO formation was also studied. GSNO prevented the oxidation of proteins SH groups. Concomitantly, a 2-fold increase of GSNO-dependent Pr-SNO formation still depending on GGT activity
was observed. Mass spectrometry identified 51 proteins S-nitrosated by GSNO under oxidative stress (vs 32 in basal
condition), including a higher number of cytoskeletal proteins (17 vs 8 in basal condition) related to cell morphogenesis and movement. Furthermore, additional proteins belong to cell adhesion and protein trafficking were S-nitrosated under oxidative stress. Oxidative stress modifies the extent and distribution of GSNO induced Pr-SNO formation, a NO storage form in tissue. Further studies will likely elucidate the pathophysiological significance of these observations
Neocytherideis labyrinthoidea Guernet & Huyghe & Lartaud & Merle & Emmanuel & Gély & Michel & Pilet 2012, n. sp.
No full textNeocytherideis labyrinthoidea n. sp. (Fig. 5 G-I) ÉTYMOLOGIE. — Ornementation constituée de côtes flexueuses plus ou moins anastomosées. MATÉRIEL TYPE. — Holotype: une valve (CGT 1242) déposée dans la collection du laboratoire de micropaléontologie de l’Université Pierre et Marie Curie Paris à Paris. Paratype: une valve (CGT 1242bis) déposée dans la même collection. DIMENSIONS (en mm). — Holotype, valve droite, L = 0,685 ± 0,01; h = 0,25 ±0,01; Paratype, valve gauche, L = 0,69 ± 0,01, h = 0,25 ± 0,01. LOCALITÉ TYPE. — Grignon (Bassin de Paris, France). NIVEAU TYPE. — Lutétien moyen (faluns à miliolidés et à Orbitolites complanatus, éch. GRS 7). RÉPARTITION STRATIGRAPHIQUE. — Lutétien moyen à Orbitolites complanatus. DESCRIPTION Valves allongées et de petite taille, bords dorsaux et ventraux subrectilignes; extrémité antérieure oblique, arrondie dans sa partie inférieure; extrémité postérieure étirée vers le haut, concave dorsalement et convexe ventralement; zone marginale modérement large, vestibule profond antérieurement; charnière lophodonte. Surface des valves ponctuée et parcourue d’une quinzaine de côtes flexueuses plus ou moins anastomosées. REMARQUE L’appartenance générique de ces valves est sans ambiguité tandis que leur ornementation est très différente de celle des autres espèces connues du genre Neocytherideis.Published as part of Guernet, Claude, Huyghe, Damien, Lartaud, Franck, Merle, Didier, Emmanuel, Laurent, Gély, Jean-Pierre, Michel, Florent & Pilet, Ophélie, 2012, Les Ostracodes de la falunière de Grignon (Lutétien du Bassin de Paris): implications stratigraphiques, pp. 909-959 in Geodiversitas 34 (4) on pages 932-933, DOI: 10.5252/g2012n4a12, http://zenodo.org/record/459749
Going Beyond Counting First Authors in Author Co-citation Analysis
No full textThe 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
Accurate measurement of reduced glutathione in gamma-glutamyltransferase-rich brain microvessel fractions
No full textS-nitrosoglutathione potentiates protein S-nitrosation under oxidative stress, a potential improvement of NO storage into smooth muscle cells
No full textCardiovascular diseases are associated with oxidative stress and reduced nitric oxide (NO) bioavailability. The ability of NO donors like S-nitrosoglutathione (GSNO) to regulate NO bioavailability under oxidative stress is poorly studied. Here, we monitored protein S-nitrosation (Pr-SNO), a post-translational protein modification in smooth muscle cells exposed to GSNO under oxidative stress. Intracellular thiol redox status in relation with the extent and distribution of GSNO-induced intracellular Pr-SNO (LC-MALDI MS) were assessed. The role of the gammaglutamyl transferase (GGT), a redox enzyme metabolizing GSNO, in Pr-SNO formation was also studied. GSNO prevented the oxidation
of proteins SH groups. Concomitantly, a 2-fold increase of GSNO-dependent Pr-SNO formation still depending on GGT activity was observed. Mass spectrometry identified 51 proteins S-nitrosated by GSNO under oxidative stress (vs 32 in basal condition), including a higher number of cytoskeletal proteins (17 vs 8 in basal condition) related to cell morphogenesis and movement. Furthermore, additional proteins belong to cell adhesion and protein trafficking were S-nitrosated under oxidative stress. Oxidative stress modifies the extent and distribution of GSNO induced Pr-SNO formation, a NO storage form in tissue. Further studies will likely elucidate the pathophysiological significance of these observations
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
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 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
Protecting Animals 36: Author Witi Ihimaera
No full textIn this very special episode of Knowing Animals I am joined by beloved New Zealand author Witi Ihimaera. Witi has written many books featuring nonhuman animals. He offers us a non-colonial lens through which to think about the human/nonhuman relationship
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