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Nitric oxide (NO) deficiency is often associated with several acute and chronic diseases. NO donors and especially S -nitrosothiols such as S -nitrosoglutathione (GSNO) have been identified as promising therapeutic agents. Although their permeability through the intestinal barrier have recently be proved, suitable drug delivery systems have to be designed for their oral administration. This is especially challenging due to the physico-chemical features of these drugs: high hydrophilicity and high lability. In this paper, three types of particles were prepared with an Eudragit® polymer: nanoparticles and microparticles obtained with a water-in-oil-in-water emulsion/evaporation process versus microparticles obtained with a solid-in-oil-in-water emulsion/evaporation process. They had a similar encapsulation efficiency (around 30%), and could be freeze-dried then be stored at least one month without modification of their critical attributes (size and GSNO content). However, microparticles had a slightly slower in vitro release of GSNO than nanoparticles, and were able to boost by a factor of two the drug intestinal permeability (Caco-2 model). Altogether, this study brings new data about GSNO intestinal permeability and three ready-to-use formulations suitable for further preclinical studies with oral administration. Image 1 • GSNO is a promising drug, but challenging to formulate (hydrophilicity, lability). • Nano and microparticles suitable for GSNO oral delivery were produced and compared. • Their critical attributes were maintained after freeze-drying and stable upon storage. • They improved the drug permeability through a Caco-2 model of intestinal barrier. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A series of aromatic and aliphatic mono-amidoximes and bis-amidoximes were synthesized. • Amidoximes were tested for their NO release ability on rat liver microsomes and human vascular cells. • Amidoximes are cytocompatible with human vascular smooth muscle cells. • Mono-amidoximes 2a and 2b exhibit high NO release. • Bis-amidoxime 2d releases the same amount of NO than 2a and 2b at a twice lower concentration. Abstract Four compounds bearing amidoxime functions were synthetized: (1) 2a,b bearing an aromatic amidoxime function, (2) 2c bearing an aliphatic amidoxime function, and (3) 2d bearing aromatic and aliphatic amidoximes functions. The ability of these compounds to release NO was evaluated in vitro using the oxidative metabolism of cytochrome P450 from rat liver microsomes. Results obtained demonstrate that all amidoximes were able to release NO with a highest amount of NO produced by the 2a aromatic amidoxime. Moreover, all amidoximes exhibit cytocompatibility with human aorta smooth muscle cells. Using intracellular S -nitrosothiol formation as a marker of NO bioavailability, compounds 2a–c were demonstrated to deliver a higher amount of NO in the intracellular environment than the reference. Considering that the concentration of the bis-amidoxime 2d was two times lower that than of 2a and 2b , we can assume that 2d is the most potent molecule among the tested compounds for NO release. [ABSTRACT FROM AUTHOR]

Decades of chemical, biochemical and pathophysiological research have established the relevance of post-translational protein modifications induced by processes related to oxidative stress, with critical reflections on cellular signal transduction pathways. A great deal of the so-called 'redox regulation' of cell function is in fact mediated through reactions promoted by reactive oxygen and nitrogen species on more or less specific aminoacid residues in proteins, at various levels within the cell machinery. Modifications involving cysteine residues have received most attention, due to the critical roles they play in determining the structure/function correlates in proteins. The peculiar reactivity of these residues results in two major classes of modifications, with incorporation of NO moieties (S-nitrosation, leading to formation of protein S-nitrosothiols) or binding of low molecular weight thiols (S-thionylation, i.e. in particular S-glutathionylation, S-cysteinylglycinylation and S-cysteinylation). A wide array of proteins have been thus analyzed in detail as far as their susceptibility to either modification or both, and the resulting functional changes have been described in a number of experimental settings. The present review aims to provide an update of available knowledge in the field, with a special focus on the respective (sometimes competing and antagonistic) roles played by protein S-nitrosations and S-thionylations in biochemical and cellular processes specifically pertaining to pathogenesis of cardiovascular diseases. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) produced by endothelial nitric oxide synthase (eNOS) activates signaling pathways responsible for smooth muscle cell relaxation, leading to vasodilation and thus plays an important role in controlling vascular homeostasis, thrombosis and inflammation. Recent studies indicate that S- nitrosothiols produced in vivo as well as synthetic ones might be important reservoirs of NO. Based on a broad range of NO functions within the living organisms, this review highlights the impact of S -nitrosothiols on cardiovascular cell cycle. The cell membrane transport and the decomposition patterns responsible of S -nitrosothiols actions are presented. The effects of NO delivery through S -nitrosothiols have a significant potential in cardiovascular diseases with various underlying causes. The challenges related to their application in the pharmacotherapy of patients with various cardiovascular diseases are also discussed. [ABSTRACT FROM AUTHOR]

S -nitrosoglutathione (GSNO) is a nitric oxide (NO) donor with therapeutic potential for cardiovascular disease treatment. Chronic oral treatment with GSNO is limited by high drug sensitivity to the environment and limited oral bioavailability, requiring the development of delivery systems able to sustain NO release. The present work describes new platforms based on polymer nanocomposite particles for the delivery of GSNO. Five types of optimized nanocomposite particles have been developed (three based on chitosan, two based on alginate sodium). Those nanocomposite particles encapsulate GSNO with high efficiency from 64% to 70% and an average size of 13 to 61 μm compatible with oral delivery. Sustained release of GSNO in vitro was achieved. Indeed, chitosan nanocomposites discharged their payload within 24 h; whereas alginate nanocomposites released GSNO more slowly (10% of GSNO was still remaining in the dosage form after 24 h). Their cytocompatibility toward intestinal Caco-2 cells (MTT assay) was acceptable (IC 50 : 6.07 ± 0.07–9.46 ± 0.08 mg/mL), demonstrating their suitability as oral delivery systems for GSNO. These delivery systems presented efficient GSNO loading and sustained release as well as cytocompatibility, showing their promise as a means of improving the oral bioavailability of GSNO and as a potential new treatment. [ABSTRACT FROM AUTHOR]

Hemoglobin-based oxygen carriers (HBOCs) may generate oxidative stress, vasoconstriction and inflammation. To reduce these undesirable vasoactive properties, we increased hemoglobin (Hb) molecular size by genetic engineering with octameric Hb, recombinant (r) HbβG83C. We investigate the potential side effects of rHbβG83C on endothelial cells. The rHbβG83C has no impact on cell viability, and induces a huge repression of endothelial nitric oxide synthase gene transcription, a marker of vasomotion. No induction of Intermolecular-Adhesion Molecule 1 and E-selectin (inflammatory markers) transcription was seen. In the presence of rHbβG83C, the transcription of heme oxygenase-1 (oxidative stress marker) is weakly increased compared to the two other HBOCs (references) or Voluven (control). This genetically engineered octameric Hb, based on a human Hb βG83C mutant, leads to little impact at the level of endothelial cell inflammatory response and thus appears as an interesting molecule for HBOC development. [ABSTRACT FROM AUTHOR]

Physiological S -nitrosothiols (RSNO), such as S -nitrosoglutathione (GSNO), can be used as nitric oxide (NO) donor for the treatment of vascular diseases. However, despite a half-life measured in hours, the stability of RSNO, limited by enzymatic and non-enzymatic degradations, is too low for clinical application. So, to provide a long-lasting effect and to deliver appropriate NO concentrations to target tissues, RSNO have to be protected. RSNO encapsulation is an interesting response to overcome degradation and provide protection. However, RSNO such as GSNO raise difficulties for encapsulation due to its hydrophilic nature and the instability of the S-NO bound during the formulation process. To our knowledge, the present study is the first description of the direct encapsulation of GSNO within polymeric nanoparticles (NP). The GSNO-loaded NP (GSNO-NP) formulated by a double emulsion process, presented a mean diameter of 289 ± 7 nm. They were positively charged (+40 mV) due to the methacrylic acid and ethylacrylate polymer (Eudragit® RL) used and encapsulated GSNO with a satisfactory efficiency (i.e. 54% or 40 mM GSNO loaded in the NP). In phosphate buffer (37 °C; pH 7.4), GSNO-NP released 100% of encapsulated GSNO within 3 h and remained stable still 6 h. However, in contact with smooth muscle cells, maximum protein nitrosation (a marker of NO bioavailability) was delayed from 1 h for free GSNO to 18 h for GSNO-NP. Therefore, protection and sustained release of NO were achieved by the association of a NO donor with a drug delivery system (such as polymeric NP), providing opportunities for vascular diseases treatment. [ABSTRACT FROM AUTHOR]

Ferroptosis is a type of cell death driven by iron overload and lipid peroxidation. It is considered a key mechanism in the development of various diseases such as atherosclerosis, Alzheimer, diabetes, cancer, and renal failure. The redox status of cells, such as the balance between intracellular oxidants (lipid peroxides, reactive oxygen species, free iron ions) and antioxidants (glutathione, glutathione Peroxidase 4), plays a major role in ferroptosis regulation and constitutes its principal biomarkers. Therefore, the induction and inhibition of ferroptosis are promising strategies for disease treatments such as cancer or neurodegenerative and cardiovascular diseases, respectively. Many drugs have been developed to exert ferroptosis-inducing and/or inhibiting reactions, such as erastin and iron-chelating compounds, respectively. In addition, many natural bioactive compounds have significantly contributed to regulating ferroptosis and ferroptosis-induced oxidative stress. Natural bioactive compounds are largely abundant in food and plants and have been for a long time, inspiring the development of various low-toxic therapeutic drugs. Currently, functional bioactive peptides are widely reported for their antioxidant properties and application in human disease treatment. The scientific evidence from biochemical and in vitro tests of these peptides strongly supports the existence of a relationship between their antioxidant properties (such as iron chelation) and ferroptosis regulation. In this review, we answer questions concerning ferroptosis milestones, its importance in physiopathology mechanisms, and its downstream regulatory mechanisms. We also address ferroptosis regulatory natural compounds as well as provide promising thoughts about bioactive peptides. [ABSTRACT FROM AUTHOR]

Aiming to prepare oily core pH-sensitive nanocapsules (NCs) for anticancer drugs delivery, the use of a dextran-based transurf (DexN 3- τ CTA γ) as both stabilizer and macromolecular chain transfer agent in methyl methacrylate/2-(diethylamino)ethyl methacrylate (MMA/DEAEMA) miniemulsion copolymerization was investigated. NCs of about 195 nm with an oily-core of Miglyol 810 (M810) and a dextran coverage covalently linked to the poly(MMA- co -DEAEMA) intern shell have been obtained. Compared to the non-sensitive PMMA-based NCs (prepared in a similar way), these novel objects were shown to swell in acidic media and to trigger Coumarin 1 release in physiological relevant pH range. As a starting point of NCs biological effects, cytotoxicity and NCs-proteins interactions studies were performed with both PMMA and poly(MMA- co -DEAEMA)-based NCs. Finally, free azide functions from dextran-based coverage were successfully exploited to attach fluorescent model dyes to NCs surface. The overall results suggest that this novel NCs platform could be potentially used as drug nanocarriers for intravenous injection. [ABSTRACT FROM AUTHOR]

Rationale: The potency of S‐nitrosoglutathione (GSNO) as a nitric oxide (NO) donor to treat cardiovascular diseases (CVDs) has been highlighted in numerous studies. In order to study its bioavailability after oral administration, which represents the most convenient route for the chronic treatment of CVDs, it is essential to develop an analytical method permitting (i) the simultaneous measurement of GSNO metabolites, i.e. nitrite, S‐nitrosothiols (RSNOs) and nitrate and (ii) to distinguish them from other sources (endogenous synthesis and diet). Methods: Exogenous GSNO was labeled with 15N, and the GS15NO metabolites after conversion into the nitrite ion were derivatized with 2,3‐diaminonaphthalene. The resulting 2,3‐naphthotriazole was quantified by liquid chromatography/tandem ion trap mass spectrometry (LC/ITMS/MS) in multiple reaction monitoring mode after Higher‐energy Collision‐induced Dissociation (HCD). Finally, the validated method was applied to an in vitro model of the intestinal barrier (monolayer of Caco‐2 cells) to study GS15NO intestinal permeability. Results: A LC/ITMS/MS method based on an original transition (m/z 171 to 156) for sodium 15N‐nitrite, GS15NO and sodium 15N‐nitrate measurements was validated, with recoveries of 100.8 ± 3.8, 98.0 ± 2.7 and 104.1 ± 3.3%, respectively. Intra‐ and inter‐day variabilities were below 13.4 and 12.6%, and the limit of quantification reached 5 nM (signal over blank = 4). The permeability of labeled GS15NO (10–100 μM) was evaluated by calculating its apparent permeability coefficient (Papp). Conclusions: A quantitative LC/ITMS/MS method using HCD was developed for the first time to selectively monitor GS15NO metabolites. The assay allowed evaluation of GS15NO intestinal permeability and situated this drug candidate within the middle permeability class according to FDA guidelines. In addition, the present method has opened the perspective of a more fundamental work aiming at studying the fragmentation mechanism leading to the ion at m/z 156 in HCD tandem mass spectrometry in the presence of acetonitrile. [ABSTRACT FROM AUTHOR]

[Display omitted] • Higher loading capacity of collagen-based hydrogels for unfractionated heparin. • Decrease storage modulus along heparin content in collagen-based hydrogel. • Heparin release from collagen-based hydrogel is biologically active. • Long lasting heparinization, at least 7 days, of collagen-based hydrogels. As the main protein forming the vascular extracellular matrix, collagen has a weak antigenicity, making it an attractive candidate for coatings of vascular grafts. In order to bring antithrombotic properties to collagen for obtaining suitable blood compatibility of surfaces and further bioactive molecule carrying capacity, heparinization appears as a method of choice. Thus, in this article, pH-driven self-assembly was used to form collagen-based hydrogels with physical incorporation of heparins, especially low molecular weight heparin or unfractionated heparin at 1 IU/mL and 6 IU/mL. These heparinized hydrogels were evaluated for their physicochemical properties including gelation kinetic, spreading, viscoelasticity, microstructure and heparin quantification, and their biocompatibility such as cytocompatibility and their capacity to release bioactive heparins with antithrombotic properties. The loading capacity of collagen-based hydrogels was higher for unfractionated heparin (60 to 80 %) than for low molecular weight heparin (20 %). Interestingly, the highest concentration (6 IU/mL) of heparin used to form collagen-based hydrogels resulted in the formation of a softer hydrogel owning a better spreadability compared to 1 IU/mL and non heparinized collagen-based hydrogel. The 3D structure observation showed a layered formation with visible pores or spaces between the layers in all types of collagen-based hydrogels. These layers are interconnected by fibrous structures, suggesting a networked architecture. Moreover, the amount of heparin released from collagen-based hydrogel prepared with 6 IU /mL was higher than from those heparinized with 1 IU/mL attested by a delay in blood coagulation (activated partial thromboplastin time and thrombin time) and the abolishment of thrombin generation. Those hydrogels were also biocompatible, with low albumin adsorption and no impact on cell viability. Finally, heparin is retained in these hydrogels for at least seven days after cell seeding, providing the possibility of long-term antithrombotic properties. Thus, we succeeded in develop/obtain coatings with antithrombogenic properties, biocompatible and able to retain heparins at least seven days, making them good candidates with a great potential for the development and application of efficient blood-contacting materials. [ABSTRACT FROM AUTHOR]

Background and Purpose: Angiotensin II (AngII) and NO regulate the cerebral circulation. AngII AT1 receptors exert ligand‐dependent and ligand‐independent (myogenic tone [MT]) vasoconstriction of cerebral vessels. NO induces post‐translational modifications of proteins such as S‐nitrosation (redox modification of cysteine residues). In cultured cells, S‐nitrosation decreases AngII's affinity for the AT1 receptor. The present work evaluated the functional consequences of S‐nitrosation on both AngII‐dependent and AngII‐independent cerebrovascular responses. Experimental Approach S‐Nitrosation was induced in rat isolated middle cerebral arteries by pretreatment with the NO donors, S‐nitrosoglutathione (GSNO) or sodium nitroprusside (SNP). Agonist‐dependent activation of AT1 receptors was evaluated by obtaining concentration–response curves to AngII. Ligand‐independent activation of AT1 receptors was evaluated by calculating MT (active vs. passive diameter) at pressures ranging from 20 to 200 mmHg in the presence or not of a selective AT1 receptor inverse agonist. Key Results: GSNO or SNP completely abolished the AngII‐dependent AT1 receptor‐mediated vasoconstriction of cerebral arteries. GSNO had no impact on responses to other vasoconstrictors sharing (phenylephrine, U46619) or not (5‐HT) the same signalling pathway. MT was reduced by GSNO, and the addition of losartan did not further decrease MT, suggesting that GSNO blocks AT1 receptor‐dependent MT. Ascorbate (which reduces S‐nitrosated compounds) restored the response to AngII but not the soluble GC inhibitor ODQ, suggesting that these effects are mediated by S‐nitrosation rather than by S‐nitrosylation. Conclusions and Implications: In rat middle cerebral arteries, GSNO pretreatment specifically affects the AT1 receptor and reduces both AngII‐dependent and AngII‐independent activation, most likely through AT1 receptor S‐nitrosation. [ABSTRACT FROM AUTHOR]

A healthy ageing process is important when it is considered that one-third of the population of Europe is already over 50 years old, although there are regional variations. This proportion is likely to increase in the future, and maintenance of vitality at an older age is not only an important measure of the quality of life but also key to participation and productivity. So, the binomial "nutrition and ageing" has different aspects and poses considerable challenges, providing a fertile ground for research and networks. The NutRedOx network will focus on the impact of redox-active compounds in food on healthy ageing, chemoprevention, and redox control in the context of major age-related diseases. The main aim of the NutRedOx network is to gather experts from Europe, and neighbouring countries, and from different disciplines that are involved in the study of biological redox active food components and are relevant to the ageing organism, its health, function, and vulnerability to disease. Together, these experts will form a major and sustainable EU-wide cluster in form of the NutRedOx Centre of Excellence able to address the topic from different perspectives, with the long-term aim to provide a scientific basis for improved nutritional and lifestyle habits, to train the next generation of multidisciplinary researchers in this field, to raise awareness of such habits among the wider population, and also to engage with industry to develop age-adequate foods and medicines. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract S -Nitrosothiols, a class of NO donors, demonstrate potential benefits for cardiovascular diseases. Drugs for such chronic diseases require long term administration preferentially through the oral route. However, the absorption of S -nitrosothiols by the intestine, which is the first limiting barrier for their vascular bioavailability, is rarely evaluated. Using an in vitro model of intestinal barrier, based on human cells, the present work aimed at elucidating the mechanisms of intestinal transport (passive or active, paracellular or transcellular pathway) and at predicting the absorption site of three S -nitrosothiols: S -nitrosoglutathione (GSNO), S -nitroso- N -acetyl- l -cysteine (NACNO) and S -nitroso- N -acetyl- d -penicillamine (SNAP). These S -nitrosothiols include different skeletons carrying the nitroso group, which confer different physico-chemical characteristics and biological activities (antioxidant and anti-inflammatory). According to the values of apparent permeability coefficient, the three S -nitrosothiols belong to the medium class of permeability. The evaluation of the bidirectional apparent permeability demonstrated a passive diffusion of the three S -nitrosothiols. GSNO and NACNO preferentially cross the intestinal barrier though the transcellular pathway, while SNAP followed both the trans- and paracellular pathways. Finally, the permeability of NACNO was favoured at pH 6.4, which is close to the pH of the jejunal part of the intestine. Through this study, we determined the absorption mechanisms of S -nitrosothiols and postulated that they can be administrated through the oral route. [ABSTRACT FROM AUTHOR]

[Display omitted] Chronic diseases such as cystic fibrosis, inflammatory bowel diseases, rheumatoid arthritis, and cardiovascular illness have been linked to a decrease in selenium levels and an increase in oxidative stress. Selenium is an essential trace element that exhibits antioxidant properties, with selenocysteine enzymes like glutathione peroxidase being particularly effective at reducing peroxides. In this study, a series of synthetic organoselenium compounds were synthesized and evaluated for their potential antioxidant activities. The new selenohydantoin molecules were inspired by selenoneine and synthesized using straightforward methods. Their antioxidant potential was evaluated and proven using classical radical scavenging and metal-reducing methods. The selenohydantoin derivatives exhibited glutathione peroxidase-like activity, reducing hydroperoxides. Theoretical calculations using Density Functional Theory (DFT) revealed the selenone isomer to be the only one occurring in solution, with selenolate as a possible tautomeric form in the presence of a basic species. Cytocompatibility assays indicated that the selenohydantoin derivatives were non-toxic to primary human aortic smooth muscle cells, paving the way for further biological evaluations of their antioxidant activity. The results suggest that selenohydantoin derivatives with trifluoro-methyl (-CF 3) and chlorine (-Cl) substituents have significant activities and could be potential candidates for further biological trials. These compounds may contribute to the development of effective therapies for chronic diseases such cardiovascular diseases. [ABSTRACT FROM AUTHOR]

Solution structures of new pyrazine-based pseudotripeptide with amidoxime function and simplified pseudodipeptide analogs were determined by a combination of IR and NMR spectroscopic studies and molecular dynamic simulations using explicit chloroform as a solvent. It was found that proline-phenylalanine dipeptide residue and amidoxime moiety in o-position are essential for intramolecular hydrogen bonding including a seven-membered γ-turn formation. In addition, a cis/trans equilibrium study was present for prolyl amides in polar solvents (DO and DMSO). A phenylalanine substituent was found to exhibit profound effect on thermodynamic parameters in prolyl peptides. The presence of intramolecular hydrogen bonds dramatically increases the amount of trans isomer in non-hydrogen-bonding CHCl and significantly favor cis isomer in hydrogen-bonding solvents such as DMSO and DO. All molecules are not cytotoxic therefore they can be further studied in relation to potent biological activities. [ABSTRACT FROM AUTHOR]

[Display omitted] • Soy and Pea hydrolysates prepared by different enzymatic hydrolysis conditions. • Sequential enzymatic hydrolysis is the most efficient regarding degree of hydrolysis. • Similar molecular weights and amino acid compositions for ultrafiltrated hydrolysates. • Metal-chelating activity determined in hydrolysates by spectrophotometry and SPR. • SPR revealed better sensitivity to detect metal-chelation than spectrophotometry. Soy and pea proteins are two rich sources of essential amino acids. The hydrolysis of these proteins reveals functional and bioactive properties of the produced small peptide mixtures. In our study, we employed the hydrolysis of soy and pea protein isolates with the endopeptidases Alcalase® and Protamex®, used alone or followed by the exopeptidase Flavourzyme®. The sequential enzyme treatments were the most efficient regarding the degree of hydrolysis. Then, soy and pea protein hydrolysates (SPHs and PPHs, respectively) were ultrafiltrated in order to select peptides of molecular weight ≤ 1 kDa. Whatever the protein source or the hydrolysis treatment, the hydrolysates showed similar molecular weight distributions and amino acid compositions. In addition, all the ultrafiltrated hydrolysates possess metal-chelating activities, as determined by UV-spectrophotometry and Surface Plasmon Resonance (SPR). However, the SPR data revealed better chelating affinities in SPHs and PPHs when produced by sequential enzymatic treatment. [ABSTRACT FROM AUTHOR]

Organogel nanoparticles of triglyceride oils were produced using an aminoacid-based low-molecular weight gelator and a polysaccharide-based stabilizer synthesized by hydrophobic modification of dextran (a non ionic bacterial polysaccharide). The hot liquid organic phase containing the organogelator was emulsified at 70 °C into an aqueous solution of amphiphilic polysaccharide by application of ultrasounds. The resulting sub micrometric oil-in-water emulsion was let cooling down to room temperature leading to an aqueous suspension of gelled oil nanoparticles. The encapsulation of alkyl esters of gallic acid with various alkyl chain lengths (2, 3 and 8 carbon atoms) was carried-out by dissolving these molecules into the hot liquid oil. Octyl gallate (OG) appeared as the most suitable molecule with about 70% of the loaded drug encapsulated into the final particles. The influences of various formulation and process parameters on the final particle size distribution and the yield of encapsulation were examined. The kinetics of release of OG from gelled oil particles was monitored and it was shown that the transport at interface was limiting the rate of release of OG both for particles of gelled oil and for oil droplets without gelator. The cytotoxicity of gelator was studied with smooth muscle cells from mouse. These tests showed that it was possible to prepare formulations encapsulating enough OG and keeping the content of gelator below its limit of cytotoxicity. [ABSTRACT FROM AUTHOR]

S-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 measurement of its activity 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). The role of GGT was assessed by stimulating enzyme activity with cosubstrate glycylglycine, as well as using two independent inhibitors, competitive serine borate complex and non-competitive acivicin. 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 vasorelaxation experiments with endothelium-intact aorta, the half maximal effective concentration of GSNO (EC50 = 3.2±0.5.10-7 M) increased in the presence of the two distinct GGT inhibitors, serine borate complex (1.6±0.2.10-6 M) and acivicin (8.3±0.6.10-7 M), while it decreased with glycylglycine (4.7±0.9.10-8 M). In endothelium-denuded aorta, EC50 for GSNO alone increased to 2.360.3.1026 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 therapeutic treatments based on GSNO are presently under development, this endothelium-dependent mechanism involved in the vascular effects of GSNO should be taken into account in a pharmacological perspective. [ABSTRACT FROM AUTHOR]

Endothelial dysfunction predisposing to cardiovascular diseases is defined as an imbalance in the production of vasodilating factors, such as nitric oxide (NO), and vasoconstrictive factors. To insure its physiological role, NO, a radical with very short half-life, requires to be stored and transported to its action site. S -nitrosothiols (RSNOs) like S -nitrosoglutathione (GSNO) represent the main form of NO storage within the vasculature. The NO store formed by RSNOs is still bioavailable to trigger vasorelaxation. In this way, RSNOs are an emerging class of NO donors with a potential to restore NO bioavailability within cardiovascular disorders. The aim of this study was to compare S -nitrosothiols ability, formed of peptide (glutathione) like the physiologic GSNO or derived from amino acids (cysteine, valine) like the synthetics S -nitroso- N -acetylcysteine (NACNO) and S -nitroso -N -acetylpenicillamine (SNAP), respectively, to produce a vascular store of NO either in endothelium-intact or endothelium-removed aortae in order to evaluate whether RSNOs can be used as therapeutics to compensate endothelial dysfunction. Sodium nitroprusside (SNP), a marketed drug already in clinics, was used as a non-RSNO NO-donor. Endothelium-intact or endothelium-removed aortae, isolated from normotensive Wistar rats, were exposed to RSNOs or SNP. Then, NO-derived (NOx) species, representing the NO store inside the vascular wall, were quantified using the diaminonaphthalene probe coupled to mercuric ions. The bioavailability of the NO store and its ability to induce vasodilation was tested using N -acetylcysteine, then its ability to counteract vasoconstriction was challenged using phenylephrine (PHE). All the studied RSNOs were able to generate a NO store materialized by a three to five times increase in NOx species inside aortae. NACNO was the most potent RSNO to produce a vascular NO store bioavailable for vasorelaxation and the most efficient to induce vascular hyporeactivity to PHE in endothelium-removed aortae. GSNO and SNAP were equivalent and more efficient than SNP. In endothelium-intact aortae, the NO store was also formed whereas it seemed less available for vasorelaxation and did not influence PHE-induced vasoconstriction. In conclusion, RSNOs – NACNO in a better extent – are able to restore NO bioavailability as a functional NO store within the vessel wall, especially when the endothelium is removed. This was associated with a hyporeactivity to the vasoconstrictive agent phenylephrine. Treatment with RSNOs could present a benefit to restore NO-dependent functions in pathological states associated with injured endothelium. [ABSTRACT FROM AUTHOR]

• Multi-reactive Dextran-based transurfs (RAFT agent and surfactant) are designed. • Dithiobenzoate groups of transurf confine RAFT polymerization at o/w interface. • RAFT miniemulsion polymerization of MMA from transurf is controlled. • Dextran-covered nanocapsules with PMMA shell and oily core are produced. • Nanocapsules seem to be cytocompatible with vascular smooth muscle cells. A multi-reactive polysaccharide-based transurf (acting both as macro-Chain Transfer Agent and stabilizer) was used to confine RAFT polymerization of methyl methacrylate (MMA) at the oil/water (o/w) miniemulsion interface. Dithiobenzoate groups and hydrophobic aliphatic side chains were introduced onto dextran, conferring it both transfer agent properties and ability to stabilize direct miniemulsion of MMA in the presence of a biocompatible oil, used as co-stabilizer. Because of their amphiphilic character, transurfs were initially adsorbed at the (o/w) interface and their reactive sites mediated RAFT polymerization via the R-group approach. PMMA-grafted dextran glycopolymers were consequently produced at the o/w interface, thus leading to dextran coverage/PMMA shell/oily core nanocapsules (NCs) as evidenced by Cryo-TEM analyses. The influence of dextran-based transurf chemistry and oil amount on MMA RAFT polymerization control was investigated. Positive preliminary results on NCs cytotoxicity suggest the potential of these objects for biomedical applications. [ABSTRACT FROM AUTHOR]

Background and Purpose: Angiotensin II (AngII) and NO regulate the cerebral circulation. AngII AT1 receptors exert ligand-dependent and ligand-independent (myogenic tone [MT]) vasoconstriction of cerebral vessels. NO induces post-translational modifications of proteins such as S-nitrosation (redox modification of cysteine residues). In cultured cells, S-nitrosation decreases AngII's affinity for the AT1 receptor. The present work evaluated the functional consequences of S-nitrosation on both AngII-dependent and AngII-independent cerebrovascular responses.Experimental Approach: S-Nitrosation was induced in rat isolated middle cerebral arteries by pretreatment with the NO donors, S-nitrosoglutathione (GSNO) or sodium nitroprusside (SNP). Agonist-dependent activation of AT1 receptors was evaluated by obtaining concentration-response curves to AngII. Ligand-independent activation of AT1 receptors was evaluated by calculating MT (active vs. passive diameter) at pressures ranging from 20 to 200 mmHg in the presence or not of a selective AT1 receptor inverse agonist.Key Results: GSNO or SNP completely abolished the AngII-dependent AT1 receptor-mediated vasoconstriction of cerebral arteries. GSNO had no impact on responses to other vasoconstrictors sharing (phenylephrine, U46619) or not (5-HT) the same signalling pathway. MT was reduced by GSNO, and the addition of losartan did not further decrease MT, suggesting that GSNO blocks AT1 receptor-dependent MT. Ascorbate (which reduces S-nitrosated compounds) restored the response to AngII but not the soluble GC inhibitor ODQ, suggesting that these effects are mediated by S-nitrosation rather than by S-nitrosylation.Conclusions and Implications: In rat middle cerebral arteries, GSNO pretreatment specifically affects the AT1 receptor and reduces both AngII-dependent and AngII-independent activation, most likely through AT1 receptor S-nitrosation. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is a gaseous messenger which plays an important role in the vascular system homeostasis. Its origin includes both endogenous (catabolism of arginine catalyzed by endothelial NO synthase) and exogenous (intake of drinking water and foods) sources. The endogenous NO production decreases within ageing and cardiovascular diseases (CVDs) related to endothelial dysfunction, thus exogenous source of NO has to supply. Drugs actually used as NO donors have major drawbacks such as oxidative stress induction and tolerance phenomenon. Numerous studies highlight the potency of low molecular weight S-nitrosothiols as NO donors because they do not exhibit the previously cited side effects. However, very few reports have been focused on their bioavailability after oral administration which represents the most convenient route for the chronic treatment of CVDs. We presently study the intestine permeability of S-nitrosoglutathione (GSNO) as a NO donor by using an ex vivo model: monolayer of differentiated Caco-2 cells. NO species (nitrite, nitrate and S-nitrosothiols) apparent permeabilities were measured with the help of a fluorogenic probe (i.e. 2,3-diaminonaphthalene) and compared with reference drugs commonly used to evaluate intestine permeability. Some perspectives to improve the oral bioavailability of GSNO are indicated. [ABSTRACT FROM AUTHOR]

Cardiovascular 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 gamma-glutamyl 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. [ABSTRACT FROM AUTHOR]