Développement d'un modèle in vitro d'exposition chronique aux forces de cisaillement pariétal sur primocultures de cellules endothéliales coronaires de rat pour l'évaluation de la dysfonction endothéliale in vitro

Le rôle de l'endothélium vasculaire dans les pathologies cardiovasculaires a gagné un intérêt croissant au cours des dernières années de recherche. Autrefois considéré comme un tissu inerte, celui-ci s'est peu à peu doté de propriétés d'interaction et de régulation qui sont à la base même de l'homéostasie du système cardiovasculaire. Les capacités d'interaction de l'endothélium avec son environnement direct ne facilitent pas son étude in vitro lorsque les cellules endothéliales sont cultivées hors de tout contexte physiologique. Le développement d'un modèle in vitro permettant de recréer et de moduler de façon indépendante les paramètres physico-chimiques du micro-environnement endothélial (forces de cisaillement pariétal, pression hydrostatique, pression partielle en oxygène) s'avère nécessaire afin d'évaluer finement les relations qui les lient. Le système des rollers permet d'évaluer de façon chronique et de façon indépendante des paramètres physiques (forces de cisaillement pariétal), chimiques (pression partielle en oxygène) et physiologiques (substances vasoactives). Les études récentes ayant mis l'accent sur le rôle primordial de la balance oxydo-réductrice de la cellule endothéliale dans la survenue ou non de phénomènes pathologiques, nous nous sommes intéressés aux réponses cellulaires pour différentes conditions micro-environnementales. Nos résultats montrent que l'augmentation des forces de cisaillement pariétal induisent un stress oxydant transitoire ainsi qu'une induction pérenne des systèmes de défense anti-oxydant alors qu'une diminution chronique semble provoquer un déséquilibre de la balance oxydo-réductrice cellulaire. L'hypoxie ne provoque pas la génération d'espèces radicalaires mais module le stress oxydant induit par le flux et augmente la biodisponibilité du monoxyde d'azote produit. L'étude de l'angiotensine II a montré un effet anti-oxydant prédominant de la voie des récepteurs AT2 sur les cellules endothéliales coronaires alors que les effets pro-inflammatoires et pro-oxydant du TNF alpha sont plus marquées sur les cellules ayant subi une diminution chronique de flux. Ainsi, il apparaît que le micro-environnement physico-chimique des cellules endothéliales module fortement leur physiologie et leurs réponses aux stress exogènes. Le rôle de l'hypoxie au niveau tissulaire ainsi que le type cellulaire impliqué dans l'oxyception restent à déterminer au moyen de systèmes de co-culture cardiomyocytes/cellules endothéliales. Par ailleurs, l'action prédominante de la voie AT2 au niveau de l'endothélium coronaire ouvre une voie de recherche supplémentaire pour le développement de stratégies thérapeutiques. En conclusion, le modèle développé permet d'évaluer finement et de façon chronique les interrelations qui existent entre l'endothélium et son environnement tant physico-chimique qu'humoral. Le concept de culture dynamique des cellules endothéliales apparaît alors primordial pour le développement d'études offrant une alternative à l'expérimentation animale.Vascular endothelium role in cardiovascular diseases has gained more and more interest for a few years. Being considered just as a barrier, research gave it many properties which became the foundations of cardiovascular homeostasis. As endothelium may interact with its micro-environment, in vitro studies are not easy since endothelial cells are grown under non-physiological conditions. Development of an in vitro model which could simulate and modulate independently physical, chemical and physiological parameters of endothelium micro-environment appears to be necessary for the in vitro studies relevance. The "Roller" system allows study in a chronic and independent fashion those parameters. As recent studies pointed out the crucial role of oxidative status of endothelial cells into the outcome of physio-pathological events, we have been interested in endothelial oxidative status cellular responses when exposed to various micro-environmental conditions. Our results show that shear stress increase induces transient oxidative stress as well as long-term anti-oxidant enzymatic systems; chronic decrease in shear stress seems to induce an imbalance in endothelial cell oxidative state. Hypoxia did not induce reactive oxygen species production but modulates flow-induced oxidative stress and enhances NO bioavailability. Angiotensin II showed a dominant anti-oxidant effect mediated by AT2 receptors and pro-inflammatory and pro-oxidant TNF alpha effects were stronger when cells had undergone chronic flow decrease. Then, it appears that micro-environment strongly affects endothelial cell in culture and their response to exogenous stresses. Hypoxia role and mechanism at the tissue level remains to be assessed with co-culture systems (cardiomyocytes/endothelial cells). Furthermore, AT2 receptor presence at the coronary level could lead to novel research in terms of therapeutical strategies. To conclude, this model allows fine and chronic evaluation of endothelium interaction with its environment. Dynamic culture concept appears to be uncircumventable for development of studies which could offer an alternative to animal experimentation.ROUEN-BU Sciences (764512102) / SudocSudocFranceF

Comparative mutagenicity and genotoxicity of particles and aerosols emitted by the combustion of standard vs. rapeseed methyl ester supplemented bio-diesel fuels: Impact of after treatment devices: Oxidation catalyst and particulate filter

International audienceDiesel exhausts are partly responsible for the deleterious effects on human health associated with urban pollution, including cardiovascular diseases, asthma, COPD, and possibly lung cancer. Particulate fraction has been incriminated and thus largely investigated for its genotoxic properties, based on exposure conditions that are, however, not relevant for human risk assessment. In this paper, original and more realistic protocols were used to investigate the hazards induced by exhausts emitted by the combustion of standard (DF0) vs. bio-diesel fuels (DF7 and DF30) and to assess the impact of exhaust treatment devices (DOC and DPF). Mutagenicity and genotoxicity were evaluated for (1) resuspended particles (“off line” exposure that takes into account the bioavailability of adsorbed chemicals) and for (2) the whole aerosols (particles + gas phase components) under continuous flow exposure (“on line” exposure). Native particles displayed mutagenic properties associated with nitroaromatic profiles (YG1041), whereas PAHs did not seem to be involved. After DOC treatment, the mutagenicity of particles was fully abolished. In contrast, the level of particle deposition was low under continuous flow exposure, and the observed mutagenicity in TA98 and TA102 was thus attributable to the gas phase. A bactericidal effect was also observed in TA102 after DOC treatment, and a weak but significant mutagenicity persisted after DPF treatment for bio-diesel fuels. No formation of bulky DNA-adducts was observed on A549 cells exposed to diesel exhaust, even in very drastic conditions (organic extracts corresponding to 500 μg equivalent particule/mL, 48 h exposure). Taken together, these data indicate that the exhausts issued from the bio-diesel fuels supplemented with rapseed methyl ester (RME), and generated by current diesel engines equipped with after treatment devices are less mutagenic than older ones. The residual mutagenicity is linked to the gas phase and could be due to pro-oxydants, mainly for RME-supplemented fuels

Nitrogen Dioxide Inhalation Exposures Induce Cardiac Mitochondrial Reactive Oxygen Species Production, Impair Mitochondrial Function and Promote Coronary Endothelial Dysfunction

International audienceTraffic air pollution is a major health problem and is recognized as an important risk factor for cardiovascular (CV) diseases. In a previous experimental study, we showed that diesel exhaust (DE) exposures induced cardiac mitochondrial and CV dysfunctions associated with the gaseous phase. Here, we hypothesized that NO(2)exposures to levels close to those found in DE induce a mitochondrial reactive oxygen species (ROS) production, which contribute to an endothelial dysfunction, an early indicator for numerous CV diseases. For this, we studied the effects of NO(2)on ROS production and its impacts on the mitochondrial, coronary endothelial and cardiac functions, after acute (one single exposure) and repeated (three h/day, five days/week for three weeks) exposures in Wistar rats. Acute NO(2)exposure induced an early but reversible mitochondrial ROS production. This event was isolated since neither mitochondrial function nor endothelial function were impaired, whereas cardiac function assessment showed a reversible left ventricular dysfunction. Conversely, after three weeks of exposure this alteration was accompanied by a cardiac mitochondrial dysfunction highlighted by an alteration of adenosine triphosphate (ATP) synthesis and oxidative phosphorylation and an increase in mitochondrial ROS production. Moreover, repeated NO(2)exposures promoted endothelial dysfunction of the coronary arteries, as shown by reduced acetylcholine-induced vasodilatation, which was due, at least partially, to a superoxide-dependent decrease of nitric oxide (NO) bioavailability. This study shows that NO(2)exposures impair cardiac mitochondrial function, which, in conjunction with coronary endothelial dysfunction, contributes to cardiac dysfunction. Together, these results clearly identify NO(2)as a probable risk factor in ischemic heart diseases

Oxidative stress induction and genotoxicity of diesel exhausts on A549 lung cells continuously exposed at the air-liquid interface. Impact of rapeseed methyl ester supplementation and of after treatment devices: oxidation catalyst and particulate filter.

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Oxidative stress induction and genotoxicity of diesel exhausts on A549 lung cells continuously exposed at the air-liquid interface. Impact of rapeseed methyl ester supplementation and of after treatment devices: oxidation catalyst and particulate filter.

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An integrated functional and transcriptomic analysis reveals that repeated exposure to diesel exhaust induces sustained mitochondrial and cardiac dysfunctions

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Abstract Number ‐ 240: Association of intravenous thrombolysis and pre‐interventional reperfusion: a post‐hoc analysis of the SWIFT DIRECT trial

Introduction One potential benefit of intravenous thrombolysis (IVT) before mechanical thrombectomy (MT) is pre‐interventional reperfusion. Currently, there is a paucity of data regarding the occurrence of pre‐interventional reperfusion in patients randomized to IVT or no‐IVT before MT. Methods SWIFT DIRECT was a randomized controlled trial including acute ischemic stroke IVT‐eligible patients being directly admitted to a comprehensive stroke center, with allocation to either MT alone or IVT + MT. Primary endpoint of this analysis was the occurrence of pre‐interventional reperfusion defined as pre‐interventional expanded Thrombolysis in Cerebral Infarction score ≥ 2a. The effect of IVT and potential treatment effect heterogeneity were analyzed using logistic regression analyses. Results Out of the 396 patients analyzed, pre‐interventional reperfusion occurred in 20 (10.0%) of patients randomized to IVT+MT, and 7 (3.6%) of patients randomized to MT alone. Receiving IVT favored the occurrence of pre‐interventional reperfusion (aOR 2.91 [95% CI 1.23 – 6.87]). There was no IVT treatment effect heterogeneity on the occurrence of pre‐interventional reperfusion with different strata of Randomization‐to‐Groin‐Puncture (p for interaction = 0.33), although the effect tended to be stronger in patients with Randomization‐to‐Groin‐Puncture >28 minutes (aOR 4.65 [95% CI 1.16 – 18.68]). There were no significant difference in rates of functional outcomes between patients with and without pre‐interventional reperfusion. Conclusions Even for patients with proximal large vessel occlusions and direct access to MT, IVT leads towards an absolute increase of 6.9% (95% CI 1.7‐12.2%) in the rates of pre‐interventional reperfusion. The effect of IVT tended to be more pronounced when Randomization‐to‐Groin‐Puncture intervals were longer, but this heterogeneity did not reach statistical significance

Comparative study of diesel and biodiesel exhausts repeated exposure on lung genotoxicity in rats

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Intravenous Fibrinolysis for Central Retinal Artery Occlusion: A Cohort Study and Updated Patient-Level Meta-Analysis

BACKGROUND AND PURPOSE: Central retinal artery occlusion results in sudden, painless, usually permanent loss of vision in the affected eye. There is no proven, effective treatment to salvage visual acuity and a clear, unmet need for an effective therapy. In this work, we evaluated the efficacy of intravenous tissue-type plasminogen activator (IV alteplase) in a prospective cohort study and an updated systematic review and meta-analysis. METHODS: We enrolled consecutive patients with acute central retinal artery occlusion within 48 hours of symptoms onset and with a visual acuity of \u3c20/200 from January 2009 until May 2019. The primary outcomes were safety and functional visual acuity recovery. We compared rates of visual recovery between those treated with alteplase within 4.5 hours of symptom onset to those who did not receive alteplase (including an analysis restricted to untreated patients presenting within the window for treatment). We incorporated these results into an updated systematic review and patient-level meta-analysis. RESULTS: We enrolled 112 patients, of whom 25 (22.3% of the cohort) were treated with IV alteplase. One patient had an asymptomatic intracerebral hemorrhage after IV alteplase treatment. Forty-four percent of alteplase-treated patients had recovery of visual acuity when treated within 4.5 hours versus 13.1% of those not treated with alteplase (P=0.003) and 11.6% of those presenting within 4 hours who did not receive alteplase (P=0.03). Our updated patient-level meta-analysis of 238 patients included 67 patients treated with alteplase within 4.5 hours since time last known well with a recovery rate of 37.3%. This favorably compares with a 17.7% recovery rate in those without treatment. In linear regression, earlier treatment correlated with a higher rate of visual recovery (P=0.01). CONCLUSIONS: This study showed that the administration of intravenous alteplase within 4.5 hours of symptom onset is associated with a higher likelihood of a favorable visual outcome for acute central retinal artery occlusion. Our results strongly support proceeding to a randomized, placebo-controlled clinical trial