RESUMEN
BACKGROUND: Outcomes and susceptibility to out-of-hospital cardiac arrest (OHCA) are known to differ by sex, yet little is known about changes in sex hormones after OHCA. We sought to determine the trajectory of sex hormones after OHCA and their association to survival and neurological outcome. METHODS: Plasma samples were collected from those that survived to hospital admission at four time points (1, 6, 24, and 48 hours) and estrone, estradiol, progesterone, and testosterone concentrations were quantified via liquid chromatography-mass spectrometry. Trends in hormones were plotted over time by sex and outcomes. The association between sex, hormone levels with survival and neurological outcome (cerebral performance category 1-2 indicating good outcome and 3-5 for poor outcome) were determined using generalized estimating equation models. RESULTS: Of the 94 OHCA patients, 50 were males and 44 females, with a mean age of 61.3 (+15.7) years. Despite older age and lower BCPR in females compared to males, females had higher proportion of good neurological outcome compared to males. Over the 48 hours, estrone increased, testosterone decreased, and estradiol and progesterone remained flat. Survivors had lower levels of estrone at all time points but only at early time points for estradiol, progesterone and testosterone. Lower estrone level predicted survival at discharge, even after adjusting for time, sex, age, and hormones independently (ß=-3.38, 95% CI= -5.71, -0.85). Females had better neurological scores compared to males after adjusting for estrone (ß=1.27, 95% CI= 0.01, 2.53) and estradiol (ß=2.92, 95% CI= 1.13, 4.70). CONCLUSIONS: Survivors and those with favorable neurological outcome had lower trend in estrone. The sex hormone estrone, present in both males and females, may be a predictor of survival. When adjusted for estrogens, female sex had better neurological recovery compared to males. The difference in neurological outcome by sex is not explained by estrogens. However, these finding open the door for exploration of other sex-specific pathways in resuscitation after OHCA.
RESUMEN
BACKGROUND: Outcomes from cardiopulmonary resuscitation (CPR) following sudden cardiac arrest are suboptimal. Postresuscitation targeted temperature management has been shown to have benefit in subjects with sudden cardiac arrest due to ventricular fibrillation, but there are few data for outcomes from sudden cardiac arrest due to pulseless electrical activity. In addition, intra-CPR cooling is more effective than postresuscitation cooling. Physical cooling is associated with increased protein kinase B activity. Therefore, our group developed a novel peptide, TAT-PHLPP9c, which regulates protein kinase B. We hypothesized that when given during CPR, TAT-PHLPP9c would improve survival and neurologic outcomes following pulseless electrical activity arrest. METHODS AND RESULTS: In 24 female pigs, pulseless electrical activity was induced by inflating balloon catheters in the right coronary and left anterior descending arteries for ≈7 minutes. Advanced life support was initiated. In 12 control animals, epinephrine was given after 1 and 3 minutes. In 12 peptide-treated animals, 7.5 mg/kg TAT-PHLPP9c was also administered at 1 and 3 minutes of CPR. The balloons were removed after 2 minutes of support. Animals were recovered and neurologically scored 24 hours after return of spontaneous circulation. Return of spontaneous circulation was more common in the peptide group, but this difference was not significant (8/12 control versus 12/12 peptide; P=0.093), while fully intact neurologic survival was significantly more common in the peptide group (0/12 control versus 11/12 peptide; P<0.00001). TAT-PHLPP9c significantly increased myocardial nicotinamide adenine dinucleotide levels. CONCLUSIONS: TAT-PHLPP9c resulted in improved survival with full neurologic function after sudden cardiac arrest in a swine model of pulseless electrical activity, and the peptide shows potential as an intra-CPR pharmacologic agent.
Asunto(s)
Reanimación Cardiopulmonar , Modelos Animales de Enfermedad , Paro Cardíaco , Animales , Reanimación Cardiopulmonar/métodos , Femenino , Paro Cardíaco/terapia , Paro Cardíaco/fisiopatología , Paro Cardíaco/tratamiento farmacológico , Porcinos , Péptidos/administración & dosificación , Péptidos/farmacología , Factores de TiempoRESUMEN
Metabolic suppression in the ischemic heart is characterized by reduced levels of NAD+ and ATP. Since NAD+ is required for most metabolic processes that generate ATP, we hypothesized that nicotinamide restores ischemic tissue NAD+ and improves cardiac function in cardiomyocytes and isolated hearts, and enhances survival in a mouse model of cardiac arrest. Mouse cardiomyocytes were exposed to 30 min simulated ischemia and 90 min reperfusion. NAD+ content dropped 40% by the end of ischemia compared to pre-ischemia. Treatment with 100 µM nicotinamide (NAM) at the start of reperfusion completely restored the cellular level of NAD+ at 15 min of reperfusion. This rescue of NAD+ depletion was associated with improved contractile recovery as early as 10 min post-reperfusion. In a mouse model of cardiac arrest, 100 mg/kg NAM administered IV immediately after cardiopulmonary resuscitation resulted in 100% survival at 4 h as compared to 50% in the saline group. In an isolated rat heart model, the effect of NAM on cardiac function was measured for 20 min following 18 min global ischemia. Rate pressure product was reduced by 26% in the control group following arrest. Cardiac contractile function was completely recovered with NAM treatment given at the start of reperfusion. NAM restored tissue NAD+ and enhanced production of lactate and ATP, while reducing glucose diversion to sorbitol in the heart. We conclude that NAM can rapidly restore cardiac NAD+ following ischemia and enhance glycolysis and contractile recovery, with improved survival in a mouse model of cardiac arrest.
Asunto(s)
Paro Cardíaco , NAD , Ratas , Animales , Ratones , Roedores , Paro Cardíaco/tratamiento farmacológico , Miocitos Cardíacos , Modelos Animales de Enfermedad , Ácido Láctico , Niacinamida/farmacología , Adenosina TrifosfatoRESUMEN
Out-of-hospital cardiac arrest is a leading cause of death in the US, with a mortality rate over 90%. Preclinical studies demonstrate that cooling during cardiopulmonary resuscitation (CPR) is highly beneficial, but can be challenging to implement clinically. No medications exist for improving long-term cardiac arrest survival. We have developed a 20-amino acid peptide, TAT-PHLPP9c, that mimics cooling protection by enhancing AKT activation via PH domain leucine-rich repeat phosphatase 1 (PHLPP1) inhibition. Complementary studies were conducted in mouse and swine. C57BL/6 mice were randomized into blinded saline control and peptide-treatment groups. Following a 12-minute asystolic arrest, TAT-PHLPP9c was administered intravenously during CPR and significantly improved the return of spontaneous circulation, mean arterial blood pressure and cerebral blood flow, cardiac and neurological function, and survival (4 hour and 5 day). It inhibited PHLPP-NHERF1 binding, enhanced AKT but not PKC phosphorylation, decreased pyruvate dehydrogenase phosphorylation and sorbitol production, and increased ATP generation in heart and brain. TAT-PHLPP9c treatment also reduced plasma taurine and glutamate concentrations after resuscitation. The protective benefit of TAT-PHLPP9c was validated in a swine cardiac arrest model of ventricular fibrillation. In conclusion, TAT-PHLPP9c may improve neurologically intact cardiac arrest survival without the need for physical cooling.
Asunto(s)
Reanimación Cardiopulmonar , Péptidos de Penetración Celular , Paro Cardíaco , Ratones , Animales , Porcinos , Reanimación Cardiopulmonar/efectos adversos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratones Endogámicos C57BL , Paro Cardíaco/terapia , Paro Cardíaco/etiología , Paro Cardíaco/metabolismo , Modelos Animales de EnfermedadRESUMEN
Therapeutic hypothermia (TH) provides cardioprotection from ischemia/reperfusion (I/R) injury. However, it remains unknown how TH regulates metabolic recovery. We tested the hypothesis that TH modulates PTEN, Akt, and ERK1/2, and improves metabolic recovery through mitigation of fatty acid oxidation and taurine release. Left ventricular function was monitored continuously in isolated rat hearts subjected to 20 min of global, no-flow ischemia. Moderate cooling (30°C) was applied at the start of ischemia and hearts were rewarmed after 10 min of reperfusion. The effect of TH on protein phosphorylation and expression at 0 and 30 min of reperfusion was investigated by western blot analysis. Post-ischemic cardiac metabolism was investigated by 13 C-NMR. TH enhanced recovery of cardiac function, reduced taurine release, and enhanced PTEN phosphorylation and expression. Phosphorylation of Akt and ERK1/2 was increased at the end of ischemia but decreased at the end of reperfusion. On NMR analysis, TH-treated hearts displayed decreased fatty acid oxidation. Direct cardioprotection by moderate intra-ischemic TH is associated with decreased fatty acid oxidation, reduced taurine release, enhanced PTEN phosphorylation and expression, and enhanced activation of both Akt and ERK1/2 prior to reperfusion.
Asunto(s)
Hipotermia , Daño por Reperfusión Miocárdica , Animales , Ratas , Ácidos Grasos , Isquemia , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fosfohidrolasa PTEN/metabolismo , Sistema de Señalización de MAP QuinasasRESUMEN
Extracorporeal cardiopulmonary resuscitation (eCPR) is emerging as an effective, lifesaving resuscitation strategy for select patients with prolonged or refractory cardiac arrest. Currently, a paucity of evidence-based recommendations is available to guide clinical management of eCPR patients. Despite promising results from initial clinical trials, neurological injury remains a significant cause of morbidity and mortality. Neuropathology associated with utilization of an extracorporeal circuit may interact significantly with the consequences of a prolonged low-flow state that typically precedes eCPR. In this narrative review, we explore current gaps in knowledge about cerebral perfusion over the course of cardiac arrest and resuscitation with a focus on patients treated with eCPR. We found no studies which investigated regional cerebral blood flow or cerebral autoregulation in human cohorts specific to eCPR. Studies which assessed cerebral perfusion in clinical eCPR were small and limited to near-infrared spectroscopy. Furthermore, no studies prospectively or retrospectively evaluated the relationship between epinephrine and neurological outcomes in eCPR patients. In summary, the field currently lacks a comprehensive understanding of how regional cerebral perfusion and cerebral autoregulation are temporally modified by factors such as pre-eCPR low-flow duration, vasopressors, and circuit flow rate. Elucidating these critical relationships may inform future strategies aimed at improving neurological outcomes in patients treated with lifesaving eCPR.
Asunto(s)
Reanimación Cardiopulmonar , Oxigenación por Membrana Extracorpórea , Paro Cardíaco , Paro Cardíaco Extrahospitalario , Humanos , Estudios Retrospectivos , Oxigenación por Membrana Extracorpórea/métodos , Reanimación Cardiopulmonar/métodos , Paro Cardíaco/terapia , Perfusión , Paro Cardíaco Extrahospitalario/terapiaRESUMEN
Preconditioning has a powerful protective potential against myocardial ischemia-reperfusion injury (I/R). Our prior work demonstrated that baicalein, a flavonoid derived from the root of Scatellaria baicalensis Georgi (also known as Huangqin), confers this preconditioning protection. This study further explored the mechanisms of baicalein preconditioning (BC-PC) in mouse cardiomyocytes. Cells were treated with baicalein (10 µM) for a brief period of time (10 min) prior to simulated ischemia 90 min/reperfusion for 180 min. Baicalein triggered an induction of a small amount of mitochondrial reactive oxygen species (ROS) prior to the initiation of ischemia, assessed by 6-carboxy-2', 7'-dichlorodihydrofluorescein diacetate (6-carboxy-H2DCFDA). It also significantly increased cell viability measured by propidium iodide (PI) and lactate dehydrogenase and preserved mitochondrial membrane potential assessed by TMRM fluorescence intensity. Myxothiazol, a mitochondrial electron transport chain complex III inhibitor, partially blocked ROS generation induced by BC-PC and reduced cell viability. BC-PC increased phosphorylation of Akt (Thr308 and Ser473) and eNOS Ser1177, and nitric oxide (NO) production measured using 4,5-diaminofluorescein diacetate (DAF-2 DA, 1 µM). Akt inhibitor API-2 abolished Akt phosphorylation and reduced DAF-2 production and cell viability. In addition, BC-PC decreased phosphorylation of pyruvate dehydrogenase (PDH) reflecting upregulated PDH activity, and increased ATP production at 30 min during reperfusion. Taken together, baicalein preconditioning-induced cardioprotection involves pro-oxidant generation, activates survival signaling Akt/eNOS/NO, and improves metabolic recovery after I/R injury. Our work provides new perspectives on the effect of baicalein on cardiac preconditioning against I/R injury.
Asunto(s)
Flavanonas , Proteínas Proto-Oncogénicas c-akt , Animales , Flavanonas/farmacología , Ratones , Óxido Nítrico/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Piruvatos , Especies Reactivas de Oxígeno/metabolismoRESUMEN
We have recently shown that pharmacologic inhibition of PTEN significantly increases cardiac arrest survival in a mouse model, however, this protection required pretreatment 30 min before the arrest. To improve the onset of PTEN inhibition during cardiac arrest treatment, we have designed a TAT fused cell-permeable peptide (TAT-PTEN9c) based on the C-terminal PDZ binding motif of PTEN for rapid tissue delivery and protection. Western blot analysis demonstrated that TAT-PTEN9c peptide significantly enhanced Akt activation in mouse cardiomyocytes in a concentration- and time-dependent manner. Mice were subjected to 8 min asystolic arrest followed by CPR, and 30 mice with successful CPR were then randomly assigned to receive either saline or TAT-PTEN9c treatment. Survival was significantly increased in TAT-PTEN9c-treated mice compared with that of saline control at 4 h after CPR. The treated mice had increased Akt phosphorylation at 30 min resuscitation with significantly decreased sorbitol content in heart or brain tissues and reduced release of taurine and glutamate in blood, suggesting improved glucose metabolism. In an isolated rat heart Langendorff model, direct effects of TAT-PTEN9c on cardiac function were measured for 20 min following 20 min global ischemia. Rate pressure product was reduced by >20% for both TAT vehicle and nontreatment groups following arrest. Cardiac contractile function was completely recovered with TAT-PTEN9c treatment given at the start of reperfusion. We conclude that TAT-PTEN9c enhances Akt activation and decreases glucose shunting to the polyol pathway in critical organs, thereby preventing osmotic injury and early cardiovascular collapse and death.NEW & NOTEWORTHY We have designed a cell-permeable peptide, TAT-PTEN9c, to improve cardiac arrest survival. It blocked endogenous PTEN binding to its adaptor and enhanced Akt signaling in mouse cardiomyocytes. It improved mouse survival after cardiac arrest, which is related to improved glucose metabolism and reduced glucose shunting to sorbitol in critical organs.
Asunto(s)
Cardiotónicos/uso terapéutico , Paro Cardíaco/tratamiento farmacológico , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Fosfohidrolasa PTEN/antagonistas & inhibidores , Animales , Encéfalo/metabolismo , Cardiotónicos/farmacología , Modelos Animales de Enfermedad , Ácido Glutámico/sangre , Paro Cardíaco/metabolismo , Ratones , Daño por Reperfusión Miocárdica/metabolismo , Miocardio/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Taurina/sangreRESUMEN
Therapeutic hypothermia initiated during cardiopulmonary resuscitation (CPR) in pre-clinical studies appears to be highly protective against sudden cardiac arrest injury. Given the challenges to implementing CPR cooling clinically, insights into its critical mechanisms of protection could guide development of new CPR drugs that mimic hypothermia effects without the need for physical cooling. Here, we used Akt1-deficient mice that lose CPR hypothermia protection to identify hypothermia targets. Adult female C57BL/6 mice (Akt1+/+ and Akt1+/-) underwent 8 min of KCl-induced asystolic arrest and were randomized to receive hypothermia (30 ± 0.5°C) or normothermia. Hypothermia was initiated during CPR and extended for 1 h after resuscitation. Neurologically scored survival was measured at 72 h. Other outcomes included mean arterial pressure and target measures in heart and brain related to contractile function, glucose utilization and inflammation. Compared to northothermia, hypothermia improved both 2h mean arterial pressure and 72h neurologically intact survival in Akt1+/+ mice but not in Akt1+/- mice. In Akt1+/+ mice, hypothermia increased Akt and GSK3ß phosphorylation, pyruvate dehydrogenase activation, and NAD+ and ATP production while decreasing IκBα degradation and NF-κB activity in both heart and brain at 30 min after CPR. It also increased phospholamban phosphorylation in heart tissue. Further, hypothermia reduced metabolic and inflammatory blood markers lactate and Pre-B cell Colony Enhancing Factor. Despite hypothermia treatment, all these effects were reversed in Akt1+/- mice. Taken together, drugs that target Akt1 and its effectors may have the potential to mimic hypothermia-like protection to improve sudden cardiac arrest survival when administered during CPR.
Asunto(s)
Reanimación Cardiopulmonar , Paro Cardíaco/metabolismo , Paro Cardíaco/terapia , Hipotermia Inducida , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Presión Sanguínea/fisiología , Reanimación Cardiopulmonar/métodos , Femenino , Glucosa/metabolismo , Hipotermia Inducida/métodos , Inflamación/metabolismo , Inflamación/terapia , Ratones Endogámicos C57BL , Ratones Transgénicos , Contracción Muscular/fisiología , Proteínas Proto-Oncogénicas c-akt/genética , Distribución AleatoriaRESUMEN
OBJECTIVE: To design, implement, and evaluate the effectiveness of an enhanced peer mentoring program (EPMP) for faculty in emergency medicine aimed at overcoming traditional mentoring challenges. METHODS: Full time faculty (Clinical Instructor, Assistant, and Associate levels) were placed into peer groups (based upon their primary academic roles) led by senior faculty advisors at the Professor level. Peer groups met at least quarterly from 2012 to 2017. In lieu of a structured curriculum, session topics were informed by individual faculty surveys and peer group consensus. Areas of focus included work-life balance, prioritizing academic commitments, identification of mentors (both within and external to the department and university), networking opportunities, promotions goals, and career satisfaction. RESULTS: Effectiveness of the EPMP was evaluated by academic productivity and advancement over a 5- year period. A total of 22 faculty members participated in the program. There was an increase in promotions to the next academic level, from 3 promotions in the five years before the program to 7 promotions in the five years of the program. Total grant funding increased 3-fold from $500,000 to $1,706,479 from the first year to the last year of the evaluation period. CONCLUSIONS: This enhanced peer mentoring program was effective in mitigating many of the traditional mentoring challenges faced by faculty in academia and was successful in improving both academic productivity and advancement.
Asunto(s)
Movilidad Laboral , Docentes Médicos , Tutoría , Grupo Paritario , Apoyo a la Investigación como Asunto/tendencias , Centros Médicos Académicos , Chicago , Eficiencia Organizacional/tendencias , Medicina de Emergencia , Femenino , Humanos , Masculino , Evaluación de Programas y Proyectos de SaludRESUMEN
Baicalein is a natural flavonoid with anti-oxidant activities protecting against ischemia/reperfusion (I/R) injury. Previous studies suggest that oxidative burst early after reperfusion accelerates cell death. We therefore investigated the critical therapeutic window of baicalein by examining the timing of baicalein treatment in relation to its oxidant modulating and cytoprotective effects. Using an established chick cardiomyocyte model of I/R, we administered baicalein at various time points after reperfusion and assessed cell viability and the profiles of reactive oxygen species (ROS), nitric oxide (NO), and Akt phosphorylation. Baicalein administered at the onset of reperfusion resulted in a concentration-dependent reduction of cell death (25 µM 48.2±1.9%, 50µM 43.8±1.5%, 100µM 36.6±2.1%, vs. I/R control 57.3±1.4%, all p<0.05). Baicalein (100µM) timely and effectively scavenged ROS burst and enhanced NO production in the early reperfusion phase. Cotreatment with NO synthase (NOS) inhibitor l-NAME (200µM) partially abrogated the cytoprotective effect. Baicalein (100µM) given after reperfusion lost protective effect in a time-dependent manner with cytoprotection completely lost if >60min. Even with only 15-min delay after reperfusion, the ROS scavenging effect was abolished and the NO enhancing effect markedly reduced. The phosphorylation of Akt, an upstream regulator of eNOS, also diminished as the delay lengthened. In conclusion, baicalein treatment after reperfusion confers cardioprotection in a concentration- and time-dependent manner. The critical therapeutic window lies in the early reperfusion phase, during which ROS scavenging and Akt-eNOS mediated NO signaling are most effective.
Asunto(s)
Cardiotónicos/farmacología , Flavanonas/farmacología , Depuradores de Radicales Libres/farmacología , Daño por Reperfusión Miocárdica/metabolismo , Óxido Nítrico/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Células Cultivadas , Pollos , Humanos , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/genética , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Oxidantes/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas c-akt/genética , Especies Reactivas de Oxígeno/metabolismo , Factores de TiempoRESUMEN
Baicalein is a flavonoid with excellent oxidant scavenging capability. It has been reported to protect against a variety of oxidative injuries including ischemia/reperfusion (I/R). However, the optimal treatment strategy for I/R injury and the protective mechanisms are not fully understood. In this study we employed an established chick cardiomyocyte model of I/R and investigated the effects of three baicalein treatment strategies on reactive oxygen species (ROS) scavenging, nitric oxide (NO) production and cell viability. The molecular signaling pathways were also explored. Compared to the I/R control (cell death 52.2[Formula: see text][Formula: see text][Formula: see text]2.0%), baicalein preventive treatment (25[Formula: see text][Formula: see text]M, pretreated for 72[Formula: see text]h and continued through I/R) conferred the best protection (19.5[Formula: see text][Formula: see text][Formula: see text]3.9%, [Formula: see text]), followed by I/R treatment (treated during I/R) and reperfusion treatment (treated at reperfusion only). Preventive and I/R treatments almost completely abolished ROS generation during both ischemic and reperfusion phases, and increased NO production and Akt phosphorylation. Reperfusion treatment reduced the ROS burst in the early reperfusion phase only, and had no effect on NO production and Akt activation. Further, the phosphorylation of phosphatase and tensin homolog (PTEN), a phosphatase negatively regulating Akt activation, was significantly increased by baicalein preventive treatment and slightly by the I/R treatment. PTEN protein expression was reduced in the same trend accordingly. Baicalein reperfusion treatment had no effects on PTEN phosphorylation and expression. Our results indicate that baicalein preventive treatment confers optimal cardioprotection against I/R injury, and this protection involves effective oxidant scavenging and the activation of PTEN/Akt/NO pathway.
Asunto(s)
Cardiotónicos , Flavanonas/farmacología , Flavanonas/uso terapéutico , Depuradores de Radicales Libres , Daño por Reperfusión Miocárdica/prevención & control , Óxido Nítrico/biosíntesis , Fosfohidrolasa PTEN/metabolismo , Fitoterapia , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Embrión de Pollo , Miocitos Cardíacos , Fosforilación/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Sudden cardiac arrest (SCA) is a leading cause of death in the United States. Despite return of spontaneous circulation, patients die due to post-SCA syndrome that includes myocardial dysfunction, brain injury, impaired metabolism, and inflammation. No medications improve SCA survival. Our prior work suggests that optimal Akt activation is critical for cooling protection and SCA recovery. Here, we investigate a small inhibitor of PTEN, an Akt-related phosphatase present in heart and brain, as a potential therapy in improving cardiac and neurological recovery after SCA. Anesthetized adult female wild-type C57BL/6 mice were randomized to pretreatment of VO-OHpic (VO) 30 min before SCA or vehicle control. Mice underwent 8 min of KCl-induced asystolic arrest followed by CPR. Resuscitated animals were hemodynamically monitored for 2 h and observed for 72 h. Outcomes included heart pressure-volume loops, energetics (phosphocreatine and ATP from (31)P NMR), protein phosphorylation of Akt, GSK3ß, pyruvate dehydrogenase (PDH) and phospholamban, circulating inflammatory cytokines, plasma lactate, and glucose as measures of systemic metabolic recovery. VO reduced deterioration of left ventricular maximum pressure, maximum rate of change in the left ventricular pressure, and Petco2 and improved 72 h neurological intact survival (50% vs. 10%; P < 0.05). It reduced plasma lactate, glucose, IL-1ß, and Pre-B cell colony enhancing factor, while increasing IL-10. VO increased phosphorylation of Akt and GSK3ß in both heart and brain, and cardiac phospholamban phosphorylation while reducing p-PDH. Moreover, VO improved cardiac bioenergetic recovery. We concluded that pharmacologic PTEN inhibition enhances Akt activation, improving metabolic, cardiovascular, and neurologic recovery with increased survival after SCA. PTEN inhibitors may be a novel pharmacologic strategy for treating SCA.
Asunto(s)
Metabolismo Energético , Inhibidores Enzimáticos/uso terapéutico , Paro Cardíaco/tratamiento farmacológico , Compuestos Organometálicos/uso terapéutico , Fosfohidrolasa PTEN/antagonistas & inhibidores , Animales , Citocinas/sangre , Femenino , Paro Cardíaco/metabolismo , Hemodinámica , Ratones , Ratones Endogámicos C57BL , Compuestos Organometálicos/farmacología , Resucitación/métodosRESUMEN
OBJECTIVES: Cooling following cardiac arrest can improve survival significantly. However, delays in achieving target temperature may decrease the overall benefits of cooling. Here, we test whether lipid emulsion, a clinically approved drug reported to exert cardioprotection, can rescue heart contractility in the setting of delayed cooling in stunned mouse cardiomyocytes. DESIGN: Cell culture study. SETTING: Academic research laboratory. SUBJECTS: Cardiomyocytes isolated from 1- to 2-day-old C57BL6 mice. INTERVENTIONS: Cardiomyocytes were exposed to 30 minutes of ischemia followed by 90 minutes of reperfusion and 10 minutes of isoproterenol with nine interventions: 1) no additional treatment; 2) intraischemic cooling at 32 °C initiated 10 minutes prior to reperfusion; 3) delayed cooling started 20 minutes after reperfusion; 4) lipid emulsion + delayed cooling; 5) lipid emulsion (0.25%) administered at reperfusion; 6) lipid emulsion + intraischemic cooling; 7) delayed lipid emulsion; 8) lipid emulsion + delayed cooling + Akt inhibitor (API-2, 10 µM); and 9) lipid emulsion + delayed cooling + Erk inhibitor (U0126, 10 µM). Inhibitors were given to cells 1 hour prior to ischemia. MEASUREMENTS AND MAIN RESULTS: Contractility was recorded by real-time phase-contrast imaging and analyzed with pulse image velocimetry in MATLAB (Mathworks, Natick, MA). Ischemia diminished cell contraction. The cardioprotective effect of cooling was diminished when delayed but was rescued by lipid emulsion. Further, lipid emulsion on its own improved recovery of the contractility to a greater extent as intraischemic cooling. However, cotreatment of lipid emulsion and intraischemic cooling did not further improve the recovery compared to either treatment alone. Furthermore, Akt and Erk inhibitors blocked lipid emulsion-induced protection. CONCLUSIONS: Lipid emulsion improved contractility and rescued contractility in the context of delayed cooling. This protective effect required Akt and Erk signaling. Lipid emulsion might serve as a treatment or adjunct to cooling in ameliorating myocardial ischemia/reperfusion injury.
Asunto(s)
Butadienos/farmacología , Cardiotónicos/farmacología , Clorpropamida/análogos & derivados , Hipotermia Inducida/métodos , Daño por Reperfusión Miocárdica/prevención & control , Miocitos Cardíacos/patología , Nitrilos/farmacología , Animales , Clorpropamida/farmacología , Modelos Animales de Enfermedad , Isquemia/fisiopatología , Ratones , Ratones Endogámicos C57BL , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Contracción Muscular/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Factores de TiempoRESUMEN
Recent work shows that cooling protection after mouse cardiac arrest and cardiomyocyte ischemia is mediated by Akt activation. The PI3K p85 subunit can either augment or inhibit Akt activation depending on its binding to p110 or PTEN respectively. To further clarify the role of PI3K p85 in cardioprotection, we studied novel TAT-p85 fusion proteins that selectively inhibit PI3K p85 binding. We hypothesized that TAT fused p85 lacking the PTEN binding site (TAT-ΔPTEN p85) would enhance Akt phosphorylation to afford cardioprotection. Conversely, TAT fused p85 lacking the p110 binding site (TAT-Δp110p85) would decrease Akt phosphorylation and abrogate cardioprotection. Microscopy and Western blot analysis demonstrated that TAT fusion protein was transduced into cardiomyocytes within 5 min and remained more than 2 h. Inhibition of PI3K/Akt by TAT-Δp110 p85 significantly increased cell death from 44.6±2.7% to 92.5±3.4% after simulated ischemia and reperfusion. By contrast, PTEN inhibition using TAT-ΔPTEN p85 decreased cell death to 11.9±5.3%, a similar level of cardioprotection seen with past cooling studies. Additional studies with the small molecule PTEN inhibitor VO-OHpic confirmed that PTEN inhibition was highly protective against cell death induced by ischemia and reperfusion. We conclude that blockade of p85-PTEN interaction and PTEN inhibition may be promising strategies for rescuing the heart from ischemia and reperfusion injury.
Asunto(s)
Productos del Gen tat/metabolismo , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/patología , Fosfohidrolasa PTEN/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Secuencia de Aminoácidos , Animales , Muerte Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Peróxido de Hidrógeno/farmacología , Ratones Endogámicos C57BL , Modelos Biológicos , Datos de Secuencia Molecular , Daño por Reperfusión Miocárdica/enzimología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/enzimología , Fosfohidrolasa PTEN/química , Fosforilación/efectos de los fármacos , Unión Proteica/efectos de los fármacosRESUMEN
Baicalein, a flavonoid derived from Scutellaria baicalensis Georgi, possesses cardioprotection against oxidant injury by scavenging reactive oxygen species (ROS). Few studies investigate whether baicalein protection is mediated by attenuating mitochondrial ROS and modulating the prosurvival and proapoptotic signaling. Primary cultured chick cardiomyocytes were used to study the role of baicalein in mitochondrial superoxide [Formula: see text] generation and signaling of Akt and JNK. Cells were exposed to H 2 O 2 for 2 h and baicalein was given 2 h prior to and during 2 h of H 2 O 2 exposure. Cell viability was assessed by propidium iodide and DNA fragmentation. H 2 O 2 (500 µM) significantly induced 45.3 ± 6.2% of cell death compared to the control (p < 0.001) and resulted in DNA laddering. Baicalein (10, 25 or 50 µM) dose-dependently reduced the cell death to 38.7 ± 5.6% (p = 0.226); 31.2 ± 3.9% (p < 0.01); 30.3 ± 5.3% (p < 0.01), respectively. It also attenuated DNA laddering. Further, baicalein decreased intracellular ROS and mitochondrial [Formula: see text] generation that was confirmed by superoxide dismutase PEG-SOD and mitochondria electron transport chain complex III inhibitor stigmatellin. In addition, baicalein increased Akt phosphorylation and decreased JNK phosphorylation in H 2 O 2-exposed cells. Moreover, baicalein augmented mitochondrial phosphorylation of Akt Thr308 and GSK3ß Ser9, and prevented mitochondrial cytochrome c release assessed by cellular fractionation. Our results suggest that baicalein cardioprotection may involve an attenuation of mitochondrial [Formula: see text] and an increase in mitochondrial phosphorylation of Akt and GSK3ß while decreasing JNK activation.
Asunto(s)
Flavanonas/farmacología , MAP Quinasa Quinasa 4/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Mitocondrias Cardíacas/metabolismo , Miocitos Cardíacos/metabolismo , Proteína Oncogénica v-akt/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Superóxidos/metabolismo , Animales , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Cardiotónicos/farmacología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Células Cultivadas , Embrión de Pollo , Flavanonas/aislamiento & purificación , Glucógeno Sintasa Quinasa 3/metabolismo , Glucógeno Sintasa Quinasa 3 beta , Miocitos Cardíacos/citología , Fosforilación/efectos de los fármacos , Scutellaria baicalensis/químicaRESUMEN
AIMS: To test the role of sphingosine-1-phosphate (S1P) signaling system in the in vivo setting of resuscitation and survival after cardiac arrest. MAIN METHODS: A mouse model of potassium-induced cardiac arrest and resuscitation was used to test the importance of S1P homeostasis in resuscitation and survival. C57BL/6 and sphingosine kinase-1 knockout (SphK1-KO) female mice were arrested for 8 min then subjected to 5 minute CPR with epinephrine bolus given at 90s after the beginning of CPR. Animal survival was monitored for 4h post-resuscitation. Upregulation of tissue and circulatory S1P levels were achieved via inhibition of S1P lyase by 2-acetyl-5-tetrahydroxybutyl imidazole (THI). Plasma and heart tissue S1P and ceramide levels were quantified by targeted ESI-LC/MS/MS. KEY FINDINGS: Lack of SphK1 and low tissue/circulatory S1P levels in SphK1-KO mice led to poor animal resuscitation after cardiac arrest and to impaired survival post-resuscitation. Inhibition of S1P lyase in SphK1-KO mice drastically improved animal resuscitation and survival. Improved resuscitation and survival of THI-treated SphK1-KO mice were better correlated with cardiac dihydro-S1P (DHS1P) than S1P levels. The lack of SphK1 and the inhibition of S1P lyase by THI were accompanied by modulation in cardiac S1PR1 and S1PR2 expression and by selective changes in plasma N-palmitoyl- and N-behenoyl-ceramide levels. SIGNIFICANCE: Our data provide evidence for the crucial role for SphK1 and S1P signaling system in resuscitation and survival after cardiac arrest, which may form the basis for development of novel therapeutic strategy to support resuscitation and long-term survival of cardiac arrest patients.
Asunto(s)
Aldehído-Liasas/antagonistas & inhibidores , Paro Cardíaco/terapia , Lisofosfolípidos/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Esfingosina/análogos & derivados , Animales , Reanimación Cardiopulmonar/métodos , Ceramidas/sangre , Cromatografía Liquida/métodos , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica , Paro Cardíaco/fisiopatología , Imidazoles/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Lisoesfingolípidos/genética , Transducción de Señal , Espectrometría de Masa por Ionización de Electrospray/métodos , Esfingosina/metabolismo , Receptores de Esfingosina-1-Fosfato , Tasa de Supervivencia , Espectrometría de Masas en Tándem/métodosRESUMEN
Previous studies suggest baicalein, in addition to its antioxidant effects, protects against hypoxia/reoxygenation injury via its pro-oxidant properties. We hypothesize that a brief period of baicalein treatment prior to ischemia/reperfusion (I/R) may trigger preconditioning protection via a mitochondrial pro-oxidant mechanism. Using an established chick cardiomyocyte model of I/R, cells were preconditioned with baicalein (10 µM) for 10 min followed by 10-min wash prior to I/R. Intracellular oxidants were measured using 2', 7'-dichlorofluorescin diacetate (DCFH/DA). Cell viability was assessed by propidium iodide and apoptosis determined by DNA fragmentation. Baicalein induced a transient but significant increase of DCF fluorescence within the 10-min preconditioning period, and led to significant reduction of cell death (38.9 ± 1.8% vs. 58.7 ± 1.2% in I/R control, n = 6, p < 0.001) and DNA fragmentation after I/R. Cotreatment with N-acetylcysteine (500 µM), mitochondrial complex III electron transport chain inhibitor myxothiazol (1 µM), mitochondrial KATP channel blocker 5-hydroxydecanoate-Na (5-HD, 500 µM) or anion channel inhibitor 4', 4'-diisothiocyanato-stilbene-2, 2'-disulfonic acid (DIDS, 200 µM) resulted in significant abrogation of oxidant increase during induction as well as the protection conferred by baicalein preconditioning. These results suggest that baicalein preconditioning exhibits significant anti-apoptotic protection against cardiomyocyte I/R injury by mitochondrial oxidant signaling, which was in part mediated by mitochondrial KATP channel and anion channel opening.
Asunto(s)
Flavanonas/administración & dosificación , Mitocondrias Cardíacas/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Oxidantes/metabolismo , Daño por Reperfusión/metabolismo , Daño por Reperfusión/prevención & control , Transducción de Señal , Animales , Células Cultivadas , Embrión de Pollo , Humanos , Precondicionamiento Isquémico , Mitocondrias Cardíacas/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Daño por Reperfusión/tratamiento farmacológicoRESUMEN
OBJECTIVE: Shallow chest compressions and incomplete recoil are common during cardiopulmonary resuscitation (CPR) and negatively affect outcomes. A step stool has the potential to alter these parameters when performing CPR in a bed but the impact has not been quantified. METHODS: We conducted a cross-over design, simulated study of in-hospital cardiac arrest. Rescuers performed a total of four 2-min segments of uninterrupted chest compressions, half of which were on a step stool. Compression characteristics were measured using a CPR-sensing defibrillator and subjective impressions were obtained from rescuer surveys. Paired analyses were performed to measure the impact of the step stool, taking into account rescuer characteristics, including height. RESULTS: Fifty subjects, of whom 36% were men, with a median height of 169.8 cm (range 148.6-190.5) volunteered to participate. Use of a step stool resulted in an average increase in compression depth of 4 mm (p<0.001) and 18% increase in incomplete recoil (p<0.001). However, unlike with incomplete recoil, the effect was more pronounced in rescuers in the lowest height tertile (9±9 mm vs 2±6 mm for those rescuers taller than 167 cm, p=0.006). CONCLUSIONS: Using a step stool when performing CPR in a bed results in a trade-off between increased compression depth and increased incomplete recoil. Given the nonlinear relationship between the increase in compression depth and rescuer height, the benefit of a step stool may outweigh the risks of incomplete release for rescuers ≤167 cm in height. The benefit is less clear in taller rescuers.
Asunto(s)
Reanimación Cardiopulmonar/métodos , Paro Cardíaco/terapia , Maniquíes , Reanimación Cardiopulmonar/educación , Estudios Cruzados , Desfibriladores , Femenino , Humanos , Masculino , Estudios ProspectivosRESUMEN
BACKGROUND: Therapeutic hypothermia (TH) has demonstrated great potential for forestalling cardiovascular collapse and improving outcomes in the setting of severe hemorrhagic shock (HS). We used an established mouse model of severe HS to study the response of interrelated cardiac-signaling proteins p38, HspB1, and Akt to shock, resuscitation, and cardioprotective TH. METHODS: Adult female C57BL6/J mice were bled and maintained at a mean arterial pressure of 35 mm Hg. After 30 minutes, mice were randomized to 120 minutes of TH (33°C ± 0.5°C) or continued normothermia at 37°C. After 90 minutes, animals were resuscitated and monitored for 180 minutes. Cardiac p38, Akt, and HspB1 phosphorylation (p-p38, p-Akt, and p-HspB1), expression, and Akt/HspB1 interactions were measured at serial time points during HS and resuscitation. Markers of mitochondrial damage (plasma cytochrome c), inflammation (myeloperoxidase), and apoptosis (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling) were analyzed. RESULTS: By 15 minutes HS, p-p38 and p-HspB1 significantly increased while p-Akt(T308) decreased (p < 0.05). TH attenuated phosphorylation of the p38α isoform during HS and increased phosphorylation of the p38γ isoform during both HS and early resuscitation (p < 0.05). TH increased Akt/HspB1 coimmunoprecipitation during early resuscitation and increased p-Akt and HspB1 expression during late resuscitation (p < 0.05). Finally, TH attenuated the myocardial myeloperoxidase and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining and plasma cytochrome c during late resuscitation. CONCLUSIONS: TH increases phosphorylation of p38γ during both HS and early resuscitation, but attenuates phosphorylation of p38α, increases Akt/HspB1 interaction, and modulates Akt phosphorylation during HS and resuscitation. Such TH-related signaling events are associated with reduced cardiac inflammation, apoptosis, and mitochondrial injury.