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Importance: Rocuronium and succinylcholine are often used for rapid sequence intubation, although the comparative efficacy of these paralytic agents for achieving successful intubation in an emergency setting has not been evaluated in clinical trials. Succinylcholine use has been associated with several adverse events not reported with rocuronium. Objective: To assess the noninferiority of rocuronium vs succinylcholine for tracheal intubation in out-of-hospital emergency situations. Design, Setting and Participants: Multicenter, single-blind, noninferiority randomized clinical trial comparing rocuronium (1.2 mg/kg) with succinylcholine (1 mg/kg) for rapid sequence intubation in 1248 adult patients needing out-of-hospital tracheal intubation. Enrollment occurred from January 2014 to August 2016 in 17 French out-of-hospital emergency medical units. The date of final follow-up was August 31, 2016. Interventions: Patients were randomly assigned to undergo tracheal intubation facilitated by rocuronium (n = 624) or succinylcholine (n = 624). Main Outcomes and Measures: The primary outcome was the intubation success rate on first attempt. A noninferiority margin of 7% was chosen. A per-protocol analysis was prespecified as the primary analysis. Results: Among 1248 patients who were randomized (mean age, 56 years; 501 [40.1%] women), 1230 (98.6%) completed the trial and 1226 (98.2%) were included in the per-protocol analysis. The number of patients with successful first-attempt intubation was 455 of 610 (74.6%) in the rocuronium group vs 489 of 616 (79.4%) in the succinylcholine group, with a between-group difference of -4.8% (1-sided 97.5% CI, -9% to ∞), which did not meet criteria for noninferiority. The most common intubation-related adverse events were hypoxemia (55 of 610 patients [9.0%]) and hypotension (39 of 610 patients [6.4%]) in the rocuronium group and hypoxemia (61 of 616 [9.9%]) and hypotension (62 of 616 patients [10.1%]) in the succinylcholine group. Conclusions and Relevance: Among patients undergoing endotracheal intubation in an out-of-hospital emergency setting, rocuronium, compared with succinylcholine, failed to demonstrate noninferiority with regard to first-attempt intubation success rate. Trial Registration: ClinicalTrials.gov Identifier: NCT02000674.
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Servicios Médicos de Urgencia , Intubación e Inducción de Secuencia Rápida/métodos , Rocuronio/administración & dosificación , Succinilcolina/administración & dosificación , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Método Simple CiegoRESUMEN
Spontaneous spleen rupture with no recent report of trauma is an extremely rare and life-threatening cause of intraperitoneal hemorrhage.We present the first case of an atraumatic pathological splenic rupture following alteplase thrombolysis for ischemic stroke.
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Fibrinolíticos/efectos adversos , Rotura del Bazo/inducido químicamente , Accidente Cerebrovascular/tratamiento farmacológico , Terapia Trombolítica/efectos adversos , Activador de Tejido Plasminógeno/efectos adversos , Humanos , Masculino , Persona de Mediana Edad , Rotura Espontánea , TrombectomíaRESUMEN
Introduction: Acute vertigo is a frequent chief complaint in the emergency departments, and its efficient management requires thorough training. The HINTS protocol is a valid method to screen patients in the emergency room, but its application in routine is hindered by the lack of training. This study aimed to evaluate the training of emergency physicians for the HINTS method based on a mannequin-based virtual reality simulator (MBVRS). Methods: We conducted a monocenter, prospective, longitudinal, and randomized cohort study in an Emergency Department at a regional university hospital. We included 34 emergency physicians randomized into two equal groups matched by age and professional experience. The control group attended a theoretical lesson with video demonstrations and the test group received a simulation-based training in addition to the lecture. Results: We showed that the test group had a higher diagnosis performance for the HINTS method compared to the control group as evaluated by the simulator at 1 month (89% sensitivity versus 45, and 100% specificity versus 86% respectively, p < 001, Fisher's exact test). Evaluation at 6 months showed a similar advantage to the test group. Discussion: The MBVRS is a useful pedagogic tool for the HINTS protocol in the emergency department. The advantage of a unique training session can be measured up to 6 months after the lesson.
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Cuidados Críticos , Vías Clínicas , Accidente Cerebrovascular/terapia , Cuidados Críticos/métodos , Cuidados Críticos/organización & administración , Vías Clínicas/organización & administración , Urgencias Médicas , Fibrinolíticos/uso terapéutico , Hospitalización , Humanos , Tiempo de Tratamiento/organización & administraciónRESUMEN
BACKGROUND: In this study, we primarily sought to assess the ability of flow cytometry to predict early clinical deterioration and overall survival in patients with sepsis admitted in the ED and ICU. METHODS: Patients admitted for community-acquired acute sepsis from 11 hospital centers were eligible. Early (day 7) and late (day 28) deaths were notified. Levels of CD64pos granulocytes, CD16pos monocytes, CD16dim immature granulocytes (IGs), and T and B lymphocytes were assessed by flow cytometry using an identical, cross-validated, robust, and simple consensus standardized protocol in each center. RESULTS: Among 1,062 patients screened, 781 patients with confirmed sepsis were studied (age, 67 ± 48 years; Simplified Acute Physiology Score II, 36 ± 17; Sequential Organ Failure Assessment, 5 ± 4). Patients were divided into three groups (sepsis, severe sepsis, and septic shock) on day 0 and on day 2. On day 0, patients with sepsis exhibited increased levels of CD64pos granulocytes, CD16pos monocytes, and IGs with T-cell lymphopenia. Clinical severity was associated with higher percentages of IGs and deeper T-cell lymphopenia. IG percentages tended to be higher in patients whose clinical status worsened on day 2 (35.1 ± 35.6 vs 43.5 ± 35.2, P = .07). Increased IG percentages were also related to occurrence of new organ failures on day 2. Increased IG percentages, especially when associated with T-cell lymphopenia, were independently associated with early (P < .01) and late (P < .01) death. CONCLUSIONS: Increased circulating IGs at the acute phase of sepsis are linked to clinical worsening, especially when associated with T-cell lymphopenia. Early flow cytometry could help clinicians to target patients at high risk of clinical deterioration. TRIAL REGISTRY: ClinicalTrials.gov; No.: NCT01995448; URL: www.clinicaltrials.gov.
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Deterioro Clínico , Citometría de Flujo/normas , Sepsis/sangre , Anciano , Femenino , Humanos , Masculino , Puntuaciones en la Disfunción de Órganos , Valor Predictivo de las Pruebas , Pronóstico , Estudios Prospectivos , Análisis de SupervivenciaRESUMEN
OBJECTIVE: Although diagnostic guidelines are similar, there is a huge difference in pulmonary embolism (PE) prevalence between the United States of America (US) and countries outside the USA (OUS) in the emergency care setting. In this study, we prospectively analyze patients' characteristics and differences in clinical care that may influence PE prevalence in different countries. METHODS: An international multicenter prospective diagnostic study was conducted in a standard-of-care setting. Consecutive outpatients presenting to the emergency unit and suspected for PE were managed using the Wells score, STA-Liatest® D-Dimers and imaging. RESULTS: The prevalence of PE in the study was 7.9% in low and moderate risk patients. Among the 1060 patients with low or moderate pre-test probability (PTP), PE prevalence was four times higher in OUS (10.7%) than in the US (2.5%) (P < 0.0001). The mean number of imaging procedures performed for one new PE diagnosis was 3.3 in OUS vs 17 in the US (P < 0.001). Stopping investigation in the case of negative D-dimers (DD combined) with low/moderate PTP was more frequent in OUS (92.7%) than in the US (75.7%) (P < 0.01). Moreover, the use of imaging was much higher in the US (44.4% vs 19.2% in OUS) in the case of moderate PTP combined with negative DD. CONCLUSION: Differences between US and OUS PE prevalence in emergency setting might be explained by differences in patients' characteristics and mostly in care patterns. US physicians performed computed tomographic pulmonary angiography more often than in Europe in cases of low/moderate PTP combined with negative DD. TRIAL REGISTRATION: ClinicalTrials.gov NCT01221805.
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Angiografía/tendencias , Servicio de Urgencia en Hospital/estadística & datos numéricos , Productos de Degradación de Fibrina-Fibrinógeno/metabolismo , Embolia Pulmonar/diagnóstico , Embolia Pulmonar/epidemiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/metabolismo , Canadá/epidemiología , Europa (Continente)/epidemiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pacientes Ambulatorios , Prevalencia , Estudios Prospectivos , Embolia Pulmonar/metabolismo , Tomografía Computarizada por Rayos X/métodos , Estados Unidos/epidemiología , Adulto JovenRESUMEN
Alterations of the microtubule network, which is involved in many vital processes, occur in several pathological conditions, such as cardiac ischemia. However, the connection between the microtubule assembly state and the factors affecting myocardial reperfusion injury, especially oxidative stress, is unknown. We aimed thus to study the effects of different tubulin ligands on the changes in the microtubule network and in several markers of cell injury and oxidative activity in cardiac muscle cells submitted to a reversible substrate-free, hypoxia-reoxygenation model of ischemia-reperfusion. The microtubule network was visualized by immunocytochemistry. Cell injury was evaluated via lactate dehydrogenase release and the mitochondrial function by the MTT test. Superoxide production was detected using dihydroethidium. The activity of NADPH oxidase and mRNA subunit expression were investigated. The microtubule disassembly induced by simulated ischemia was reversed by placing cardiomyocytes under normoxic conditions. This post-"ischemic" restoration of microtubule assembly was modulated by microtubule stabilizers (taxol: paclitaxel) and by microtubule disrupting drugs (nocodazole, colchicine). In addition, nocodazole decreased superoxide anion production as well as NADPH oxidase activity and mRNA expression of the NADPH oxidase subunit p22phox. These results demonstrated that the "ischemia"-induced microtubule network alteration is reversible and suggest a possible relationship between "reperfusion"-induced reassembly of microtubules and free radical generation in post-"ischemic" cardiomyocytes.
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Microtúbulos/metabolismo , Isquemia Miocárdica/metabolismo , Miocitos Cardíacos/metabolismo , NADPH Oxidasas/metabolismo , Animales , Células Cultivadas , Regulación de la Expresión Génica/efectos de los fármacos , L-Lactato Deshidrogenasa/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Mitocondrias Cardíacas/metabolismo , Reperfusión Miocárdica , Miocitos Cardíacos/patología , NADPH Oxidasa 2 , NADPH Oxidasas/genética , Nocodazol/farmacología , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Superóxidos/metabolismo , Tubulina (Proteína)/metabolismoRESUMEN
We investigated the influence of PUFA in phospholipids (PL) on the functional characteristics of cultured cardiomyocytes (CM) in basal conditions and during free radical (FR) stress provoked either by the xanthine/xanthine oxidase (X/XO) system or by a (9Z, 11E, 13 (S), 15Z)-13-hydroperoxyoctadecatrienoic acid (13-HpOTrE). CM were grown in media containing either n - 3 (eicosapentaenoic acid, EPA, and docosahexaenoic acid, DHA) or n - 6 (arachidonic acid, AA). These two groups of CM displayed different PUFA n - 6/n - 3 ratio in PL. However, their basal electromechanical characteristics were similar. The X/XO system drastically altered CM functions, without difference between the two groups of CM. 13-HpOTrE caused a moderate and reversible depression in action potential parameters, which was dependent upon the PL PUFA, since the n - 3-enriched CM exhibited an earlier functional depression but faster recovery. Thus, the peroxidative damage of CM depended on a cross relationship between FR species and the PL PUFA composition.
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Ácidos Grasos Insaturados/metabolismo , Miocitos Cardíacos/metabolismo , Estrés Oxidativo , Fosfolípidos/metabolismo , Animales , Células Cultivadas , Radicales Libres/metabolismo , Ácidos Linoleicos/metabolismo , Peróxidos Lipídicos/metabolismo , Miocitos Cardíacos/citología , Ratas , Ratas Wistar , Xantina/metabolismo , Xantina Oxidasa/metabolismoRESUMEN
Cyclosporin A is an immunosuppressor that prolongs graft survival but its use is limited by cardiotoxicity. The effects of cyclosporin A on several functional and biological characteristics were thus evaluated in rat cardiomyocytes in normal conditions and in a substrate-free, hypoxia-reoxygenation model of ischemia-reperfusion. Cyclosporin A (100 and 1000 ng/ml) did not induce cardiocytotoxicity in basal conditions. Simulated ischemia gradually decreased and then blocked the spontaneous electromechanical activity. Cyclosporin A at 100 and 1000 ng/ml permitted the maintenance of electromechanical functions that were abolished in control cells. Cyclosporin A also improved the post-"ischemic" functional recovery. Cyclosporin A reduced the "ischemia"-induced lactate dehydrogenase and troponine I releases and the successive rises in heat shock protein mRNA observed after "ischemia" and reoxygenation. Moreover, cyclosporin A improved the resumption of the mitochondrial function. To conclude, cyclosporin A displayed a direct, pleiotropic protection of isolated cardiomyocytes against physiological, metabolic, structural and stress signaling changes induced by ischemia-reperfusion mimicked in vitro.
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Ciclosporina/farmacología , Miocitos Cardíacos/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Animales , Hipoxia de la Célula , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Relación Dosis-Respuesta a Droga , Proteínas HSP70 de Choque Térmico/genética , Inmunosupresores/farmacología , L-Lactato Deshidrogenasa/metabolismo , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/fisiología , Oxígeno/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo , Transcripción Genética/efectos de los fármacos , Troponina I/metabolismoRESUMEN
The management of stroke is now recognized as a real medical emergency as well as myocardial infarct, because we have now an efficacious treatment in cerebral infarct, intravenous fibrinolysis that decreases the risk of death and motor and cognitive handicap. The second characteristic is its very important frequency, and its risk that increases in young people. This medical emergency enforces the care systems because it needs a speedy network for the patient, his family and the care professionals, useful for intravenous fibrinolysis before 3 hours after 80 years and before 4 hours and a half before 80 years. It is necessary to start treatment as soon as possible because it is most effective when given early. The consequences to avoid the lost of chance, need several actions: inform the public about the interest of FAST score to identify the first signs (facial palsy, palsy of arm, aphasia and time of stroke onset); call 15; translate the patient towards an appropriate medical center; use tele-stroke when the hospital has no neurologist; and manage the patient in a stroke unit, to introduce in a second time secondary prevention thanks to therapeutical education. Therefore, stroke care is a real multiprofessional emergency around the neurologist.
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Accidente Cerebrovascular/terapia , Adulto , Anciano , Anciano de 80 o más Años , Urgencias Médicas , Servicios Médicos de Urgencia/organización & administración , Historia del Siglo XXI , Hospitalización , Humanos , Persona de Mediana Edad , Accidente Cerebrovascular/epidemiología , Terapia Trombolítica/normas , Terapia Trombolítica/tendenciasRESUMEN
BACKGROUND: We investigated the influence of deep hypothermia (4 degrees C) during ischemia-reperfusion in the isolated rat heart model. METHODS: Isolated, perfused rat hearts underwent either 30 minutes of normothermic ischemia (control group) or 30 minutes of hypothermic ischemia (hypothermia-treated group), followed by 30 minutes of reperfusion in both groups. We recorded functional parameters and used electron spin resonance (ESR) spectroscopy to detect ascorbyl radicals, as markers of free-radical production, in samples of coronary effluents. RESULTS: Functional parameters were stable in the 2 groups during pre-ischemic and ischemic periods. During reperfusion, coronary flow, left diastolic ventricular pressure, left ventricular developed pressure, and heart rate more rapidly recovered to values close to those obtained during the pre-ischemic period in the hypothermia-treated group than in the control group. Moreover, the post-ischemic contracture observed in the control group did not appear in the hypothermia-treated group. Finally, ESR analysis showed that the post-ischemic release of ascorbyl radicals decreased in the hypothermia-treated group. CONCLUSIONS: These results demonstrate that the protective effect of hypothermia against functional injury caused by ischemia-reperfusion may decrease the free-radical burst at reperfusion.
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Circulación Coronaria/fisiología , Ácido Deshidroascórbico/análogos & derivados , Ácido Deshidroascórbico/metabolismo , Hipotermia Inducida , Isquemia Miocárdica/metabolismo , Isquemia Miocárdica/terapia , Daño por Reperfusión/prevención & control , Animales , Modelos Animales de Enfermedad , Espectroscopía de Resonancia por Spin del Electrón , Ratas , Ratas Wistar , Recuperación de la Función/fisiología , Presión Ventricular/fisiologíaRESUMEN
This work aimed to detect functional adenosine receptors in isolated rat cardiomyocytes and to study the influence of stimulation of these receptors in an in vitro model of ischemia. Cultures of cardiomyocytes were prepared from newborn rat ventricles. The contractions were photometrically monitored. In this preparation, adenosine induced a positive chronotropic response. This effect was reproduced by CGS 21680 (2-(4-[2-carboxyethyl]-phen-ethyl-amino) adenosine-5'N-ethylunosamide), a specific adenosine A(2) receptor agonist, and antagonized by DMPX (3,7-dimethyl-1-propargylxanthine), an adenosine A(2) receptor antagonist. However, R-PIA (R-N(6)-(2-phenylisopropyl)-adenosine; a specific adenosine A(1) receptor agonist) induced a negative chronotropic effect that was abolished by its corresponding adenosine A(1) antagonist DPCPX (1,3-dipropyl-8-cyclo-pentyl-adenosine). Substrate-free hypoxia, as simulation of ischemia, induced a progressive decrease and then arrest of spontaneous cell contractions. The spontaneous rhythmic contractile activity was restored during reoxygenation following simulated ischemia. Adenosine A(1) receptor stimulation with R-PIA induced a decrease of hypoxia-induced damage. This effect was antagonized by DPCPX, an adenosine A(1) receptor antagonist. Conversely, the cells treated with CGS 21680 did not display complete recovery after reoxygenation. In addition, this effect was abolished by DMPX, since the cells recovered normal function after reoxygenation. To conclude, it appeared that cardiomyocytes possess both functional adenosine A(1) and A(2) receptors and that only the activation of adenosine A(1) receptor had a cytoprotective effect against simulated ischemia-induced cardiac cell injury.
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Adenosina/farmacología , Citoprotección/efectos de los fármacos , Isquemia Miocárdica , Miocitos Cardíacos/efectos de los fármacos , Adenosina/uso terapéutico , Animales , Hipoxia de la Célula/efectos de los fármacos , Hipoxia de la Célula/fisiología , Células Cultivadas , Citoprotección/fisiología , Isquemia Miocárdica/tratamiento farmacológico , Isquemia Miocárdica/fisiopatología , Miocitos Cardíacos/fisiología , Agonistas del Receptor Purinérgico P1 , Antagonistas de Receptores Purinérgicos P1 , Ratas , Ratas Wistar , Receptores Purinérgicos P1/fisiologíaRESUMEN
Mycophenolate mofetil (MMF) is a new immunosuppressive drug used to reduce acute rejection after heart transplantation. As with other immunosuppressive drugs, MMF therapy is associated with several adverse effects. However, the direct effects of MMF on myocardial tissue has not been yet evaluated. The aim of the work was thus to evaluate the effects of MMF on isolated cardiomyocytes (CM) in normal conditions and in an in vitro model of simulated ischemia (SI; substrate-free hypoxia) and reperfusion (R; reoxygenation). Myocyte-enriched cultures were prepared from newborn rat heart ventricles. The transmembrane potentials were recorded using conventional microelectrodes and the cell contractions were monitored with a photoelectric device. In basal conditions, MMF (10(-6) and 10(-5) M) exerted no significant effects on the survival and on the electrical and contractile activities of CM in culture, even during long-term exposure (up to 48 h). SI per se led to a gradual decrease and then an abortion of the spontaneous automaticity and electromechanical activity of CM. Pretreating CM with either 10(-6) or 10(-5) M MMF was able to reduce the SI-induced cell dysfunctions. The presence of MMF at these concentrations did not hamper the post-SI functional recovery of CM during reoxygenation. At 10(-5) M, MMF applied during reoxygenation only permitted a better recovery of CM. However, the mitochondrial function after reoxygenation, as assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl-tetrazolium bromide (MTT) test, was not significantly influenced by the addition of MMF before as well as after ischemia. Conversely, MMF was able to reduce in this model the postischemic rise in xanthine and hypoxanthine. These data from CM-enriched model show that MMF: (i) had no cytotoxic effect, (ii) displayed a cytoprotective effect during SI, and (iii) exerted its beneficial effect at least partly through the decrease in the xanthine oxidase-dependent free radical production.
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Inmunosupresores/efectos adversos , Ácido Micofenólico/análogos & derivados , Ácido Micofenólico/efectos adversos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Adenosina/metabolismo , Animales , Animales Recién Nacidos , Hipoxia de la Célula/efectos de los fármacos , Células Cultivadas , Hipoxantina/metabolismo , Potenciales de la Membrana , Isquemia Miocárdica/metabolismo , Reperfusión Miocárdica , Miocitos Cardíacos/fisiología , Ratas , Ratas Wistar , Xantina/metabolismoRESUMEN
Before transplantation, the heart graft is preserved by the use of cold storage in order to limit ischemia-reperfusion stress. However, sustained exposure to low temperature may induce myocardial ultrastructural damage, particularly microtubules (MT) disruption. Previous data suggested that tubulin-binding agents are able to attenuate cold-induced cytoskeleton alterations. Thus, the aim of the present work was to study the influence of docetaxel (DX, a tubulin-binding taxane) on the effects of deep hypothermia (4 degrees C) and of simulated cold ischemia-reperfusion on the MT network and oxidative stress of cardiomyocyte (CM) in monolayer cultures prepared from newborn rat ventricles. The MT network was explored by immunocytochemistry and Western-blotting, the cell stress by tetrazolium dye assay (MTT) and lactate dehydrogenase (LDH) release, and the superoxide production by the dihydroethidium probe (DHE). The MT assembly remained stable after 4 and 8 h of hypothermia. Tubulin acetylation was promoted in CM subjected to 4-h hypothermia. Low temperature reduced the mitochondrial function and increased the basal LDH release. The cold ischemia during 4 and 8 h preserved MT network. Docetaxel promoted MT polymerization and tubulin acetylation in basal and in cold conditions. This drug decreased the release of LDH induced by cold ischemia. Moreover, hypothermia (4 h) significantly raised the anion superoxide production. Docetaxel decreased this oxidative stress in the control CM and in CM submitted to 4 h of hypothermia. These data demonstrated that stabilizing MT with DX exerted a protective effect on CM subjected to hypothermia and to cold ischemia-reperfusion. Tubulin-ligands should be thus considered to improve the tolerance of the heart graft toward stressing conservative conditions.
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Isquemia Fría/efectos adversos , Citoprotección , Microtúbulos/fisiología , Miocitos Cardíacos/patología , Daño por Reperfusión/patología , Acetilación , Acetiltransferasas/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Citoprotección/efectos de los fármacos , Dimerización , Docetaxel , Radicales Libres/metabolismo , Hipotermia/patología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Unión Proteica , Ratas , Ratas Wistar , Daño por Reperfusión/metabolismo , Taxoides/farmacología , Tubulina (Proteína)/metabolismoRESUMEN
Defining the substrate that influences the most favourably the myocardial post-ischemic recovery is subject of debates, due to dissociation between functional and biochemical benefits. Hence, we studied the effects of either glucose or different fatty acids on the functional and metabolic recovery of post-ischemic cardiomyocytes in a substrate-free hypoxia model of simulated ischemia-reperfusion. Rat cardiomyocytes were submitted to a 2.5 h simulated ischemia followed by a 2 h reoxygenation without substrate (control), or with either glucose, octanoic acid, oleic acid, or elaidic acid. During simulated ischemia, electromechanical function gradually disappeared while the cellular viability and mitochondrial function declined. During control simulated reperfusion, cardiomyocytes recovered near normal function but a significant reduction in the action potential amplitude and rate persisted. The addition of glucose or oleic acid during simulated reperfusion promoted a faster, better and sustain functional recovery. Amongst the fatty acids, the functional recovery was slower with elaidic and octanoic acids as compared with oleic acid. The mitochondrial function was better improved during simulated reperfusion with glucose than with the tested fatty acids, among which elaidic acid was the less unfavourable. Paradoxically, the addition of whichever substrate during simulated reperfusion tended to worsen the cellular viability. Thus, cardiomyocytes recovery strongly relies on the characteristics of the substrate supplied at the onset of simulated reperfusion: glucidic or lipidic nature, chain-length, insaturation degree. Moreover, these data suggest that defining the appropriateness of a given substrate for the post-ischemic cardiomyocyte recovery is closely related to the functional and the biological endpoints in consideration.
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Caprilatos/farmacología , Glucosa/farmacología , Isquemia Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Miocitos Cardíacos/metabolismo , Ácido Oléico/farmacología , Animales , Animales Recién Nacidos , Antihipertensivos/farmacología , Biomarcadores/metabolismo , Hipoxia de la Célula , Isquemia Miocárdica/tratamiento farmacológico , Isquemia Miocárdica/patología , Reperfusión Miocárdica , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/citología , Ácidos Oléicos , Oxígeno/metabolismo , Ratas , Ratas Wistar , Especificidad por Sustrato , Edulcorantes/farmacologíaRESUMEN
The main factors of myocardial ischemia are hypoxia, substrate deprivation, acidosis, and high extracellular potassium concentration ([K+]e), but the influence of each of these factors has not yet been evaluated in a cardiomyocyte (CM) culture system. Electromechanical responses to the individual and combined components of ischemia were studied in CM cultured from newborn rat ventricles. Action potentials (APs) were recorded using glass microelectrodes and contractions were monitored photometrically. Glucose-free hypoxia initially reduced AP duration, amplitude, and rate and altered excitation-contraction coupling, but AP upstroke velocity (Vmax) remained unaffected. Early afterdepolarizations appeared, leading to bursts of high-rate triggered impulses before the complete arrest of electromechanical activity after 120 min. Acidosis reduced Vmax whereas AP amplitude and rate were moderately decreased. Combining acidosis and substrate-free hypoxia also decreased Vmax but attenuated the effects of substrate-free hypoxia on APs and delayed the cessation of the electrical activity (180 min). Raising [K+]e reduced the maximal diastolic potential and Vmax. Total ischemia (substrate deletion, hypoxia, acidosis, and high [K+]e) decreased AP amplitude and Vmax without changing AP duration. Moreover, delayed afterdepolarizations appeared, initiating triggered activity. Ultimately, 120 min of total ischemia blocked APs and contractions. To conclude, glucose-free hypoxia caused severe functional defects, acidosis delayed the changes induced by substrate-free hypoxia, and total ischemia induced specific dysfunctions differing from those caused by the former conditions. Heart-cell cultures thus represent a valuable tool to scrutinize the individual and combined components of ischemia on CMs.
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Potenciales de Acción/fisiología , Isquemia Miocárdica , Miocitos Cardíacos/fisiología , Animales , Hipoxia de la Célula/fisiología , Células Cultivadas , Isquemia Miocárdica/fisiopatología , Miocitos Cardíacos/citología , Ratas , Ratas WistarRESUMEN
Cytoskeleton damage, particularly microtubule (MT) alterations, may play an important role in the pathogenesis of ischemia-induced myocardial injury. However, this disorganization has been scarcely confirmed in the cellular context. We evaluated MT network disassembly in myoblast cell line H9c2 and in neonatal rat cardiomyocytes in an in vitro substrate-free hypoxia model of simulated ischemia (SI). After different duration of SI from 30 up to 180 min, the cells were fixed and the microtubule network was revealed by immunocytochemistry. The microtubule alterations were quantified using a house-developed image analysis program. Additionally, the tubulin fraction were extracted and quantified by Western blotting. The cell respiration, the release of cellular LDH and the cell viability were evaluated at the same periods. An early MT disassembly was observed after 60 min of SI. The decrease in MT fluorescence intensity at 60 and 90 min was correlated with a microtubule disassembly. Conversely, SI-induced significant LDH release (35%) and decrease in cell viability (34%) occurred after 120 min only. These results suggest that the simulated ischemia-induced changes in MT network should not be considered as an ultrastructural hallmark of the cell injury and could rather be an early ultrastructural correlate of the cellular reaction to the metabolic challenge.