Argon Attenuates Multiorgan Failure in Relation with HMGB1 Inhibition.

Argon inhalation attenuates multiorgan failure (MOF) after experimental ischemic injury. We hypothesized that this protection could involve decreased High Mobility Group Box 1 (HMGB1) systemic release. We investigated this issue in an animal model of MOF induced by aortic cross-clamping. Anesthetized rabbits were submitted to supra-coeliac aortic cross-clamping for 30 min, followed by 300 min of reperfusion. They were randomly divided into three groups (n = 7/group). The Control group inhaled nitrogen (70%) and oxygen (30%). The Argon group was exposed to a mixture of argon (70%) and oxygen (30%). The last group inhaled nitrogen/oxygen (70/30%) with an administration of the HMGB1 inhibitor glycyrrhizin (4 mg/kg i.v.) 5 min before aortic unclamping. At the end of follow-up, cardiac output was significantly higher in Argon and Glycyrrhizin vs. Control (60 ± 4 and 49 ± 4 vs. 33 ± 8 mL/kg/min, respectively). Metabolic acidosis was attenuated in Argon and Glycyrrhizin vs. Control, along with reduced amount of norepinephrine to reverse arterial hypotension. This was associated with reduced interleukin-6 and HMGB1 plasma concentration in Argon and Glycyrrhizin vs. Control. End-organ damages were also attenuated in the liver and kidney in Argon and Glycyrrhizin vs. Control, respectively. Argon inhalation reduced HMGB1 blood level after experimental aortic cross-clamping and provided similar benefits to direct HMGB1 inhibition.

Effectiveness of pure argon for renal transplant preservation in a preclinical pig model of heterotopic autotransplantation.

BACKGROUND: In kidney transplantation, the conditions of organ preservation following removal influence function recovery. Current static preservation procedures are generally based on immersion in a cold-storage solution used under atmospheric air (approximately 78 kPa N2, 21 kPa O2, 1 kPa Ar). Research on static cold-preservation solutions has stalled, and modifying the gas composition of the storage medium for improving preservation was considered. Organoprotective strategies successfully used noble gases and we addressed here the effects of argon and xenon on graft preservation in an established preclinical pig model of autotransplantation. METHODS: The preservation solution Celsior saturated with pure argon (Argon-Celsior) or xenon (Xenon-Celsior) at atmospheric pressure was tested versus Celsior saturated with atmospheric air (Air-Celsior). The left kidney was removed, and Air-Celsior (n = 8 pigs), Argon-Celsior (n = 8) or Xenon-Celsior (n = 6) was used at 4 °C to flush and store the transplant for 30 h, a duration that induced ischemic injury in our model when Air-Celsior was used. Heterotopic autotransplantation and contralateral nephrectomy were performed. Animals were followed for 21 days. RESULTS: The use of Argon-Celsior vs. Air-Celsior: (1) improved function recovery as monitored via creatinine clearance, the fraction of excreted sodium and tubulopathy duration; (2) enabled diuresis recovery 2-3 days earlier; (3) improved survival (7/8 vs. 3/8 pigs survived at postoperative day-21); (4) decreased tubular necrosis, interstitial fibrosis, apoptosis and inflammation, and preserved tissue structures as observed after the natural death/euthanasia; (5) stimulated plasma antioxidant defences during the days following transplantation as shown by monitoring the "reduced ascorbic acid/thiobarbituric acid reactive substances" ratio and Hsp27 expression; (6) limited the inflammatory response as shown by expression of TNF-alpha, IL1-beta and IL6 as observed after the natural death/euthanasia. Conversely, Xenon-Celsior was detrimental, no animal surviving by day-8 in a context where functional recovery, renal tissue properties and the antioxidant and inflammation responses were significantly altered. Thus, the positive effects of argon were not attributable to the noble gases as a group. CONCLUSIONS: The saturation of Celsior with argon improved early functional recovery, graft quality and survival. Manipulating the gas composition of a preservation medium constitutes therefore a promising approach to improve preservation.

Argon pharmacokinetics: a solubility measurement technique.

The noble gas argon has demonstrated biological activity that may prove useful as a medical intervention. Pharmacokinetics, the disposition of the drug molecule in the body through time, is fundamental necessary knowledge to drug discovery, development and even post-marketing. The fundamental measurement in pharmacokinetic studies is blood concentration of the molecule (and its metabolites) of interest. While a physiologically based model of argon pharmacokinetics has appeared in the literature, no experimental data have been published. Thus, argon pharmaceutical development requires measurement of argon solubility in blood. This paper reports on the development of a technique based on mass spectrometry for measuring argon solubility in liquids, including blood, to be further employed in pharmacokinetics testing of argon. Based on a prototype, results are reported from sensitivity experiments using ambient air, water and rabbit blood. The key takeaway is that the system was sensitive to argon during all of the testing. We believe the technique and prototype of the quadrupole mass spectrometer gas analyzer will be capable of inferring argon pharmacokinetics through the analysis of blood samples.

Computational fluid dynamics applied to the ventilation of small-animal laboratory cages.

Several studies based on in vivo or in vitro models have found promising results for the noble gas argon in neuroprotection against ischaemic pathologies. The development of argon as a medicinal product includes the requirement for toxicity testing through non-clinical studies. The long exposure period of animals (rats) during several days results in technical and logistic challenges related to the gas administration. In particular, a minimum of 10 air changes per hour (ACH) to maintain animal welfare results in extremely large volumes of experimental gas required if the gas is not recirculated. The difficulty with handling the many cylinders prompted the development of such a recirculation-based design. To distribute the recirculating gas to individually ventilated cages and monitor them properly was deemed more difficult than constructing a single large enclosure that will hold several open cages. To address these concerns, a computational fluid dynamics (CFD) analysis of the preliminary design was performed. A purpose-made exposure chamber was designed based on the CFD simulations. Comparisons of the simulation results to measurements of gas concentration at two cage positions while filling show that the CFD results compare well to these limited experiments. Thus, we believe that the CFD results are representative of the gas distribution throughout the enclosure. The CFD shows that the design provides better gas distribution (i.e. a higher effective air change rate) than predicted by 10 ACH.

Numerical analysis of mechanical ventilation using high concentration medical gas mixtures in newborns.

When administered in relatively high concentrations the mechanical properties of inhaled gas can become significantly different from air. This fact has implications in mechanical ventilation where adequate respiration and injury to the lungs or respiratory muscles can worsen morbidity and mortality. Here we use an engineering pressure loss model to analyze the administration of medical gas mixtures in newborns. The model is used to determine the pressure distribution along the gas flow path. Numerical experiments comparing medical gas mixtures with helium, nitrous oxide, argon, xenon, and medical air as a control, with and without an endotracheal tube obstruction were performed. The engineering pressure loss model was incorporated into a model of mechanical ventilation during pressure control mode, a ventilator mode that is often used for neonates. Results are presented in the form of Rohrer equations relating pressure loss to flow rate for each gas mixture with and without obstruction. These equations were incorporated into a model for mechanical ventilation resulting in pressure, flow rate, and volume curves for the inhalation-exhalation cycle. In terms of accuracy, published values of airway resistance range from 50 to 150 cmH2O/L per second for a normal 3 kg infant. With air, the current results are 55 to 80 cmH2O/L per second for 0.3 to 5 L/min. It is shown that density through inertial pressure losses has a greater influence on airway resistance than viscosity in spite of relatively low flow rates and small airway dimensions of newborns. The results indicate that the high-density xenon mixture can be problematic during mechanical ventilation. On the other hand, low density heliox (a mixture of helium and oxygen) provides a wider margin of safety for mechanical ventilation than the other gas mixtures. The argon or nitrous oxide mixtures considered are only slightly different from air in terms of mechanical ventilation performance.

A porcine ex vivo lung perfusion model with maximal argon exposure to attenuate ischemia-reperfusion injury.

Argon (Ar) is a noble gas with known organoprotective effects in rodents and in vitro models. In a previous study we failed to find a postconditioning effect of Ar during ex vivo lung perfusion (EVLP) on warm-ischemic injury in a porcine model. In this study, we further investigated a prolonged exposure to Ar to decrease cold ischemia-reperfusion injury after lung transplantation in a porcine model with EVLP assessment. Domestic pigs (n = 6/group) were pre-conditioned for 6 hours with 21% O2 and 79% N2 (CONTR) or 79% Ar (ARG). Subsequently, lungs were cold flushed and stored inflated on ice for 18 hours inflated with the same gas mixtures. Next, lungs were perfused for 4 hours on EVLP (acellular) while ventilated with 12% O2 and 88% N2 (CONTR group) or 88% Ar (ARG group). The perfusate was saturated with the same gas mixture but with the addition of CO2 to an end-tidal CO2 of 35-45 mmHg. The saturated perfusate was drained and lungs were perfused with whole blood for an additional 2 hours on EVLP. Evaluation at the end of EVLP did not show significant effects on physiologic parameters by prolonged exposure to Ar. Also wet-to-dry weight ratio did not improve in the ARG group. Although in other organ systems protective effects of Ar have been shown, we did not detect beneficial effects of a high concentration of Ar on cold pulmonary ischemia-reperfusion injury in a porcine lung model after prolonged exposure to Ar in this porcine model with EVLP assessment.

Red wine polyphenol compounds favor neovascularisation through estrogen receptor α-independent mechanism in mice.

Red wine polyphenol compounds (RWPC) exert paradoxical effects depending on the dose on post-ischemic neovascularisation. Low dose RWPC (0.2 mg/kg/day) is pro-angiogenic, whereas high dose (20 mg/kg/day) is anti-angiogenic. We recently reported that the endothelial effect of RWPC is mediated through the activation of a redox-sensitive pathway, mitochondrial biogenesis and the activation of α isoform of the estrogen receptor (ERα). Here, we investigated the implication of ERα on angiogenic properties of RWPC. Using ovariectomized mice lacking ERα treated with high dose of RWPC after hindlimb ischemia, we examined blood flow reperfusion, vascular density, nitric oxide (NO) production, expression and activation of proteins involved in angiogenic process and muscle energy sensing network. As expected, high dose of RWPC treatment reduced both blood flow and vascular density in muscles of mice expressing ERα. These effects were associated with reduced NO production resulting from diminished activity of eNOS. In the absence of RWPC, ERα deficient mice showed a reduced neo-vascularisation associated with a decreased NO production. Surprisingly in mice lacking ERα, high dose of RWPC increased blood flow and capillary density in conjunction with increased NO pathway and production as well as VEGF expression. Of particular interest is the activation of Sirt-1, AMPKα and PGC-1α/ß axis in ischemic hindlimb from both strains. Altogether, the results highlight a pro-angiogenic property of RWPC via an ERα-independent mechanism that is associated with an up-regulation of energy sensing network. This study brings a corner stone of a novel pathway for RWPC to correct cardiovascular diseases associated with failed neovascularisation.

Hepatic protein tyrosine phosphatase 1B (PTP1B) deficiency protects against obesity-induced endothelial dysfunction.

Growing evidence suggests that hepatic-insulin resistance is sufficient to promote progression to cardiovascular disease. We have shown previously that liver-specific protein-tyrosine-phosphatase 1B (PTP1B) deficiency improves hepatic-insulin sensitivity and whole-body glucose homeostasis. The aim of this study was to investigate the impact of liver-specific PTP1B-deficiency (L-PTP1B-/-) on cardiac and peripheral vascular function, with special emphasis on endothelial function in the context of high-fat diet (HFD)-induced obesity. L-PTP1B-/- mice exhibited an improved glucose and lipid homeostasis and increased insulin sensitivity, without changes in body weight. HFD-feeding increased systolic blood pressure (BP) in both L-PTP1B-/- and control littermates; however, this was significantly lower in L-PTP1B-/- mice. HFD-feeding increased diastolic BP in control mice only, whilst the L-PTP1B-/- mice were completely protected. The analysis of the function of the left ventricle (LV) revealed that HFD-feeding decreased LV fractional shortening in control animals, which was not observed in L-PTP1B-/- mice. Importantly, HFD feeding significantly impaired endothelium-dependent vasorelaxation in response to acetylcholine in aortas from control mice, whilst L-PTP1B-/- mice were fully protected. This was associated with alterations in eNOS phosphorylation. Selective inhibition of COX-2, using NS-398, decreased the contractile response in response to serotonin (5-HT) only in vessels from control mice. HFD-fed control mice released enhanced levels of prostaglandin E, a vasoconstrictor metabolite; whilst both chow- and HFD-fed L-PTP1B-/- mice released higher levels of prostacylin, a vasorelaxant metabolite. Our data indicate that hepatic-PTP1B inhibition protects against HFD-induced endothelial dysfunction, underscoring the potential of peripheral PTP1B inhibitors in reduction of obesity-associated cardiovascular risk in addition to its anti-diabetic effects.

Propionyl-L-carnitine corrects metabolic and cardiovascular alterations in diet-induced obese mice and improves liver respiratory chain activity.

AIMS: Obesity is a primary contributor to acquired insulin resistance leading to the development of type 2 diabetes and cardiovascular alterations. The carnitine derivate, propionyl-L-carnitine (PLC), plays a key role in energy control. Our aim was to evaluate metabolic and cardiovascular effects of PLC in diet-induced obese mice. METHODS: C57BL/6 mice were fed a high-fat diet for 9 weeks and then divided into two groups, receiving either free- (vehicle-HF) or PLC-supplemented water (200 mg/kg/day) during 4 additional weeks. Standard diet-fed animals were used as lean controls (vehicle-ST). Body weight and food intake were monitored. Glucose and insulin tolerance tests were assessed, as well as the HOMA(IR), the serum lipid profile, the hepatic and muscular mitochondrial activity and the tissue nitric oxide (NO) liberation. Systolic blood pressure, cardiac and endothelial functions were also evaluated. RESULTS: Vehicle-HF displayed a greater increase of body weight compared to vehicle-ST that was completely reversed by PLC treatment without affecting food intake. PLC improved the insulin-resistant state and reversed the increased total cholesterol but not the increase in free fatty acid, triglyceride and HDL/LDL ratio induced by high-fat diet. Vehicle-HF exhibited a reduced cardiac output/body weight ratio, endothelial dysfunction and tissue decrease of NO production, all of them being improved by PLC treatment. Finally, the decrease of hepatic mitochondrial activity by high-fat diet was reversed by PLC. CONCLUSIONS: Oral administration of PLC improves the insulin-resistant state developed by obese animals and decreases the cardiovascular risk associated to this metabolic alteration probably via correction of mitochondrial function.

Estrogen receptor alpha as a key target of red wine polyphenols action on the endothelium.

BACKGROUND: A greater reduction in cardiovascular risk and vascular protection associated with diet rich in polyphenols are generally accepted; however, the molecular targets for polyphenols effects remain unknown. Meanwhile evidences in the literature have enlightened, not only structural similarities between estrogens and polyphenols known as phytoestrogens, but also in their vascular effects. We hypothesized that alpha isoform of estrogen receptor (ERalpha) could be involved in the transduction of the vascular benefits of polyphenols. METHODOLOGY/PRINCIPAL FINDINGS: Here, we used ERalpha deficient mice to show that endothelium-dependent vasorelaxation induced either by red wine polyphenol extract, Provinols, or delphinidin, an anthocyanin that possesses similar pharmacological profile, is mediated by ERalpha. Indeed, Provinols, delphinidin and ERalpha agonists, 17-beta-estradiol and PPT, are able to induce endothelial vasodilatation in aorta from ERalpha Wild-Type but not from Knock-Out mice, by activation of nitric oxide (NO) pathway in endothelial cells. Besides, silencing the effects of ERalpha completely prevented the effects of Provinols and delphinidin to activate NO pathway (Src, ERK 1/2, eNOS, caveolin-1) leading to NO production. Furthermore, direct interaction between delphinidin and ERalpha activator site is demonstrated using both binding assay and docking. Most interestingly, the ability of short term oral administration of Provinols to decrease response to serotonin and to enhance sensitivity of the endothelium-dependent relaxation to acetylcholine, associated with concomitant increased NO production and decreased superoxide anions, was completely blunted in ERalpha deficient mice. CONCLUSIONS/SIGNIFICANCE: This study provides evidence that red wine polyphenols, especially delphinidin, exert their endothelial benefits via ERalpha activation. It is a major breakthrough bringing new insights of the potential therapeutic of polyphenols against cardiovascular pathologies.