11 research outputs found

    Glucose and Pharmacological Modulators of ATP-Sensitive K+ Channels Control [Ca2+]c by Different Mechanisms in Isolated Mouse α-Cells

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    OBJECTIVE—We studied how glucose and ATP-sensitive K+ (KATP) channel modulators affect α-cell [Ca2+]c

    Clinical Pathway Evaluation for Left and Sigmoid Colectomy in Abdominal Surgery

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    At the end of 2008, a new left colon clinical pathway was implemented in our hospital and set up by a multidisciplinary team, monitored by a clinical pathway coordinator. Our aim was to evaluate the quality of left and sigmoid colectomy management, to simplify the clinical pathway and to assess its impact on the patient, the medical and nursing staffs. A sample of 290 patients with benign or malignant disease requiring a laparoscopic of laparotomy left colon resection (mainly sigmoid) was included in this clinical pathway during the years 2009–2017. Our analysis focused particularly on the compliance with the protocol, the pain felt, the suture leak rate, the hospital stay, the re-hospitalization rate and redo surgery within 30 days. Our work leads to the conclusion that the introduction of a clinical pathway, when it is well prepared and brings together all the implicated persons with the same goal, is feasible with convincing results. These are directly beneficial to the patient and to the quality of its management

    Effets du donneur de peroxynitrite, la 3-morpholinosydnonimine, sur les cellules endothéliales. RÎle du facteur de transcription Nrf2 et de la voie UPR.

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    In physiological conditions, endothelial cells produce superoxide anions (O2‱-) and nitric oxide (NO). O2‱- are precursors, among others, of hydrogen peroxide, involved in signaling processes leading to the regulation of gene expression. At higher concentrations, hydrogen peroxide promotes the formation of the cytotoxic hydroxyl radicals. NO, in turn, exerts well-recognized beneficial effects on the cardiovascular system by inducing in particular the relaxation of smooth muscle cells. But when the NO concentration becomes too high, it competes with superoxide dismutases and reacts with O2‱- to form peroxynitrite. The latter is a highly reactive molecule, attacking fatty acids, nucleic acids and proteins (with the formation of, for instance, 3-nitrotyrosine) and involved in endothelial dysfunction, a phenomenon at the basis of cardiovascular diseases. Reactive oxygen or nitrogen species can therefore, depending on the conditions, act as second messengers at low concentrations or as toxic molecules inducing oxidative stress at higher concentrations. Under normal conditions, the production of peroxynitrite is low and the damage that it could cause is limited by various defense systems, such as the Nrf2 (nuclear transcription factor erythroid 2p45 - related factor) and the UPR pathways (Unfolded Protein Response). Nrf2 is a transcription factor that responds to oxidative stress and regulates the expression of genes encoding detoxifying or antioxidant proteins such as heme oxygenase-1 (HO-1) or NAD(P)H quinone oxidoreductase 1 (NQO1). The UPR is an intracellular signaling pathway activated by the accumulation of misfolded proteins in the endoplasmic reticulum leading to the activation of various proteins such as the PERK kinase and the ATF6 transcription factor. The UPR aims to increase the synthesis of chaperonnes including BiP and GRP94, and thus increase the cellular capacity to fold or eliminate misfolded proteins. However, if the stress is too intense or too long, the UPR can induce cell death by apoptosis in particular via the activation of the transcription factor CHOP (CAAT/ enhancer binding protein (C/EBP) homologous protein). The objective of this thesis was to better understand the effects of peroxynitrite on endothelial cells, by discriminating, on one hand, its role as second messenger triggering defense responses to stress and, on the other hand, its toxicity leading to apoptosis. To achieve this goal, we chose an experimental system for peroxynitrite formation, SIN-1 (3-morpholinosydnonimine). We also tried to develop "more physiological" systems of peroxynitrite generation in vitro. Among the systems described to generate peroxynitrite, we chose homocysteine, hypoxia/reoxygenation, oxidized LDL and angiotensin-II. In this study, we developped an experimental model of endothelial cell exposure to peroxynitrite by incubating human endothelial cells in culture (the EAhy926 cell line and the primary culture, HUVEC) in the presence of SIN-1. Firstly, we tested the cytotoxicity of SIN-1 and we characterized the peroxynitrite formation induced by SIN-1 in these cellular models by following the oxidation of the hydroxyphenyl fluorescein probe and the formation of 3-nitrotyrosine by Western blot with anti-3-NT antibodies. Secondly, we demonstrated the activation of the Nrf2 and UPR pathways by SIN-1. We also showed that SIN-1 was able to exert a protective effect in serum-starved EAhy926 cells. By investigating some of the molecular mechanisms involved in the cytoprotective effect of SIN-1, we found that, via Nrf2 and HO-1 proteins, SIN-1 induced a decrease in DNA fragmentation and increased LC3-II formation in serum-starved endothelial cells. We also evaluated the effects of more physiological conditions (homocysteine, hypoxia/reoxygenation, oxidized LDL and angiotensin-II) on the peroxynitrite formation. It seems that the latter is generated in the presence of oxidized LDL and angiotensin-II combined with a NO donor. However, more investigations are necessary to further optimize some of these "more physiological" conditions on the peroxynitrite formation and to confirm the cytoprotective effect of peroxynitrite observed in serum-starved endothelial cells exposed to SIN-1. Altogether, our results indicate that in a narrow range of concentration, peroxynitrite formed after SIN-1 stimulation, activates the Nrf2 and UPR pathways, leading to a cytoprotective effect in serum-starved endothelial cells. This protective effect is achieved by stimulating the endothelial cells either after or before a period of serum starvation. The latter is in agreement with so called conditioning experiments in vivo. This work provides some new insights in the balance between survival signals or cell death triggered by peroxynitrite and highlights under which conditions this balance leads to endothelial cell death, in the context of endothelial dysfunction, involved in the early stages of atherosclerosis. It also suggests that a drastic anti-oxidant therapy targetting cardiovascular diseases could have drawbacks, as it abolishes the positive effects of reactive oxygen and nitrogen species.En conditions physiologiques, les cellules endothĂ©liales produisent des anions superoxyde (O2‱-) et du monoxyde d’azote (NO). Les O2‱- sont des prĂ©curseurs, entre autres, du peroxyde d’hydrogĂšne, impliquĂ© dans des processus de signalisation aboutissant Ă  des rĂ©gulations d’expression gĂ©nique. A plus forte concentration, le peroxyde d’hydrogĂšne favorise la formation de radicaux hydroxyl, cytotoxiques. Le NO, quant Ă  lui, a une action bĂ©nĂ©fique reconnue sur le systĂšme cardiovasculaire en induisant notamment la relaxation des cellules musculaires lisses. Mais lorsque la concentration en NO est trop importante, il entre en compĂ©tition avec les superoxyde dismutases et rĂ©agit avec les O2‱- pour former le peroxynitrite. Ce dernier est une molĂ©cule hautement rĂ©actionnelle, rĂ©agissant avec les acides gras, les acides nuclĂ©iques et les protĂ©ines (avec par exemple, la formation de 3-nitrotyrosines) et impliquĂ©e dans le dysfonctionnement endothĂ©lial, phĂ©nomĂšne Ă  la base des maladies cardiovasculaires. Les espĂšces rĂ©actives de l’oxygĂšne ou de l’azote peuvent donc, selon les conditions, jouer un rĂŽle de messagers secondaires Ă  faible concentration ou de molĂ©cules toxiques induisant un stress oxydatif Ă  plus forte concentration. En conditions normales, la production de peroxynitrite est faible et les dommages qu’il pourrait induire sont limitĂ©s par divers systĂšmes de dĂ©fense. Parmi ces moyens de dĂ©fense, il y a la voie du facteur de transcription Nrf2 (« nuclear transcription factor erythroid 2p45 – related factor ») et la voie UPR (« Unfolded Protein Response »). Nrf2 est un facteur de transcription qui rĂ©pond aux stress oxydatifs et rĂ©gule l’expression des gĂšnes codant pour des protĂ©ines anti-oxydantes ou dĂ©toxifiantes telles que l’hĂšme oxygĂ©nase-1 (HO-1) ou la NAD(P)H quinone oxydorĂ©ductase 1 (NQO1). La voie UPR est une voie de signalisation intracellulaire cytoprotectrice activĂ©e par l’accumulation de protĂ©ines mal repliĂ©es dans le rĂ©ticulum endoplasmique aboutissant Ă  l’activation de divers acteurs molĂ©culaires dont la kinase PERK et le facteur de transcription ATF6. La voie UPR a pour but d’augmenter la synthĂšse des chaperonnes dont BiP et Grp94, et ainsi d’augmenter la capacitĂ© de bien replier et d’éliminer les protĂ©ines mal repliĂ©es. Cependant, si le stress est trop intense ou de trop longue durĂ©e, la voie UPR peut induire la mort cellulaire par apoptose notamment via le facteur de transcription CHOP (« CAAT/Enhancer binding protein (C/EBP) homologous protein »). L’objectif de ce travail de thĂšse Ă©tait de mieux comprendre les effets du peroxynitrite dans les cellules endothĂ©liales, en discriminant d’une part sa fonction de messager secondaire dĂ©clenchant des rĂ©ponses de dĂ©fense au stress, et d’autre part, sa toxicitĂ© pouvant mener Ă  l’apoptose. Pour ce faire, nous avons choisi un systĂšme expĂ©rimental de formation de peroxynitrite, le SIN-1 (3-morpholinosydnonimine). Nous avons aussi essayĂ© de mettre au point diffĂ©rents systĂšmes dits « plus physiologiques » de gĂ©nĂ©ration de peroxynitrite in vitro. Parmi les systĂšmes dĂ©crits pour gĂ©nĂ©rer du peroxynitrite, nous avons choisi l’homocystĂ©ine, l’hypoxie/rĂ©oxygĂ©nation, les LDL oxydĂ©es et l’angiotensine-II. Au cours de ce travail, nous avons dĂ©veloppĂ© un modĂšle expĂ©rimental d’exposition des cellules endothĂ©liales au peroxynitrite en incubant des cellules endothĂ©liales humaines en culture (lignĂ©e EAhy926 et cellules en primo-culture HUVEC) en prĂ©sence de SIN-1. Dans un premier temps, nous avons testĂ© la cytotoxicitĂ© du SIN-1 et nous avons caractĂ©risĂ© la formation de peroxynitrite Ă  partir de SIN-1 dans ces modĂšles cellulaires en suivant l’oxydation de la sonde hydroxyphĂ©nyl fluorescĂ©ine et la formation de 3-nitrotyrosines par une analyse en Western blot avec des anticorps dirigĂ©s contre les 3-nitrotyrosines. Dans un second temps, nous avons mis en Ă©vidence l’activation de la voie Nrf2 et de la voie UPR par le SIN-1 et nous avons dĂ©montrĂ© que le SIN-1 exerce un effet protecteur dans les cellules EAhy926 soumises Ă  une privation de sĂ©rum. En dĂ©cortiquant les mĂ©canismes molĂ©culaires impliquĂ©s dans l’effet cytoprotecteur du SIN-1, nous avons constatĂ© que, via les protĂ©ines Nrf2 et HO-1, le SIN-1 induit une diminution de la fragmentation de l’ADN et une augmentation de la formation de LC3-II dans les cellules endothĂ©liales soumises Ă  une privation de sĂ©rum. Au cours de ce travail, nous avons Ă©galement Ă©valuĂ© l’effet de conditions particuliĂšres (homocystĂ©ine, hypoxie/rĂ©oxygĂ©nation, LDL oxydĂ©es et angiotensine-II) sur la formation de peroxynitrite. Il semblerait que ce dernier soit gĂ©nĂ©rĂ© en prĂ©sence de LDL oxydĂ©es et d’angiotensine-II combinĂ©e Ă  un donneur de NO. Toutefois, de plus amples investigations sont encore nĂ©cessaires afin de pouvoir optimaliser ces conditions dites « plus physiologiques » sur la formation de peroxynitrite et confirmer l’effet cytoprotecteur du peroxynitrite observĂ© dans des cellules endothĂ©liales soumises Ă  une privation de sĂ©rum et exposĂ©es au SIN-1. Les rĂ©sultats obtenus au cours de ce travail montrent donc que, dans une gamme de concentration Ă©troite, le peroxynitrite formĂ© suite Ă  la stimulation des cellules endothĂ©liales avec le SIN-1, permet de les protĂ©ger contre la toxicitĂ© induite par une privation de sĂ©rum, que la stimulation avec le SIN-1 soit rĂ©alisĂ©e aprĂšs ou avant la privation de sĂ©rum, cette derniĂšre condition se rapprochant des expĂ©riences dites de conditionnement in vivo. Ce travail nous a donc permis de mieux comprendre l’équilibre entre les signaux de survie ou de mort cellulaires dĂ©clenchĂ©s par le peroxynitrite et de prĂ©ciser dans quelles conditions cet Ă©quilibre bascule vers la mort des cellules endothĂ©liales, dans le contexte du dysfonctionnement endothĂ©lial, impliquĂ© dans les Ă©tapes prĂ©coces de l'athĂ©rosclĂ©rose. Il suggĂšre Ă©galement qu’une thĂ©rapie anti-oxydante trop drastique des maladies cardiovasculaires risque de supprimer les actions positives des espĂšces rĂ©actives de l’oxygĂšne et des espĂšces rĂ©actives de l’azote prĂ©sents en faible concentration.(DOCSC03) -- FUNDP, 201

    Clinical pathway evaluation for left colectomy in abdominal surgery

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    peer reviewedFin 2008, l’itinĂ©raire clinique (IC) «colectomie gauche» a Ă©tĂ© mis en place au sein de la Clinique Saint-Joseph (CHC) de LiĂšge. Une sĂ©rie de 213 patients prĂ©sentant une pathologie bĂ©nigne ou maligne nĂ©cessitant une rĂ©section du cĂŽlon gauche par laparoscopie a Ă©tĂ© incluse dans cet IC entre 2009 et 2015. Nous nous sommes intĂ©ressĂ©s Ă  l’observance du protocole de l’IC ainsi qu’aux taux de complications et de rĂ©-hospitalisations dans les 30 jours post-opĂ©ratoires. Nous avons constatĂ©, aprĂšs comparaison avec un groupe tĂ©moin historique, que l’adhĂ©sion au protocole IC a Ă©tĂ© d’emblĂ©e excellente (> 80 %) tout au long de la durĂ©e de l’étude. Il n’y a pas eu de modification du taux de rĂ©-hospitalisations et le taux de lĂąchage de suture a Ă©tĂ© rĂ©duit. Bien que la diminution de la durĂ©e de sĂ©jour n’était pas l’objectif premier lors de la mise en place de cet IC, elle s’est significativement rĂ©duite passant, en moyenne, de 8 Ă  4 jours. En conclusion, l’introduction d’un IC, pour autant qu’il soit bien prĂ©parĂ© et rassemble dans le mĂȘme objectif l’ensemble des acteurs de soins, est directement bĂ©nĂ©fique pour le patient et la qualitĂ© de sa prise en charge

    Rectal cancer treatment in a teaching hospital

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    Background: Rectal adenocarcinomas surgery morbidity and mortality might be impaired by neoadjuvant therapy. We performed this retropsective study to be compared with the PROCARE study running afterwards. Methods: We performed a retrospective study of 95 patients operated on for rectal denocarcinoma in a single institution during the period 2007-2009. We used logistic regression to estimate the relationship between possible predictive parameters of AL. Results: The laparoscopic approach is favored in 63.1% of the cases with a conversion rate of 11.6%, mainly in man (6 out of 7). For low rectal cancer though, laparotomy was the first choice (92.3%). From a carcinological point of view, laparoscopy allowed a complete tumor resection according to the PME (n=27) and TME (n=26) standards. Multivariate analysis revealed that women, lower BMI, lower rectum tumor, laparoscopic surgery, neoadjuvant treatment and anal suture were associated with higher risk of AL. The mean hospital stay was 15.4 days (3 – 46 days). In-hospital mortality was 3.1%. Adjuvant chemotherapy was completed in 42.1% of the patients. Despite these treatments, we registered a recurrence rate of 26.6%. Of these, 72% were distally localized and 12% exclusively locally. Among the patients operated on by laparoscopy, there was one local recurrence and one local with distant metastases (3.7%). The one- and three-years survival rates were 91.5% and 80.4% respectively. Conclusions: Our study showed a higher rate of AL than expected (18%). In our series recorded in PROCARE-Home, our leak rate has dropped to 10%. It may be indicating a positive effect of PROCARE

    Copper and myeloperoxidase-modified LDLs activate Nrf2 through different pathways of ROS production in macrophages.

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    Low-density lipoprotein (LDL) oxidation is a key step in atherogenesis, promoting the formation of lipid-laden macrophages. Here, we compared the effects of copper-oxidized LDLs (OxLDLs) and of the more physiologically relevant myeloperoxidase-oxidized LDLs (MoxLDLs) in murine RAW264.7 macrophages and in human peripheral blood monocyte-derived macrophages. Both oxidized LDLs, contrary to native LDLs, induced foam cell formation and an intracellular accumulation of reactive oxygen species (ROS). This oxidative stress was responsible for the activation of the NF-E2-related factor 2 (Nrf2) transcription factor, and the subsequent Nrf2-dependent overexpression of the antioxidant genes, Gclm and HO-1, as evidenced by the invalidation of Nrf2 by RNAi. MoxLDLs always induced a stronger response than OxLDLs. These differences could be partly explained by specific ROS-producing mechanisms differing between OxLDLs and MoxLDLs. Whereas both types of oxidized LDLs caused ROS production partly by NADPH oxidase, only MoxLDLs-induced ROS production was dependent on cytosolic PLA2. This study highlights that OxLDLs and MoxLDLs induce an oxidative stress, through distinct ROS-producing mechanisms, which is responsible for the differential activation of the Nrf2 pathway. These data clearly suggest that results obtained until now with copper oxidized-LDLs should be carefully reevaluated, taking into consideration physiologically more relevant oxidized LDLs.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

    [Ca2+]c in single mouse α-cells is controlled by modulators of K+ATP channels but not by glucose

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    BACKGROUND AND AIMS: Stimulus-secretion coupling in pancreatic α-cells is poorly understood. Changes in the free cytosolic Ca2+ concentration ([Ca2+]c) are implicated in the control of glucagon release, but the mechanisms by which secretagogues modify α-cell [Ca2+]c remain highly controversial. These were studied here in single α-cells isolated from a mouse model expressing a fluorescent protein (FP) under the control of the glucagon promoter. [...
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