TAT delivery of a PTEN peptide inhibitor has direct cardioprotective effects and improves outcomes in rodent models of cardiac arrest.

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.

A novel pharmacological strategy by PTEN inhibition for improving metabolic resuscitation and survival after mouse cardiac arrest.

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.

Lipid emulsion rapidly restores contractility in stunned mouse cardiomyocytes: a comparison with therapeutic hypothermia.

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.

Survival and Neurologic Outcomes From Pharmacologic Peptide Administration During Cardiopulmonary Resuscitation of Pulseless Electrical Activity.

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.

Extracorporeal cardiopulmonary resuscitation (eCPR) and cerebral perfusion: A narrative review.

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.

Intra-ischemic hypothermia cardioprotection involves modulation of PTEN/Akt/ERK signaling and fatty acid oxidation.

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.

Nicotinamide restores tissue NAD+ and improves survival in rodent models of cardiac arrest.

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.

A cell-penetrating PHLPP peptide improves cardiac arrest survival in murine and swine models.

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.

Mitochondrial oxidant stress triggers cell death in simulated ischemia-reperfusion.

To clarify the relationship between reactive oxygen species (ROS) and cell death during ischemia-reperfusion (I/R), we studied cell death mechanisms in a cellular model of I/R. Oxidant stress during simulated ischemia was detected in the mitochondrial matrix using mito-roGFP, a ratiometric redox sensor, and by Mito-Sox Red oxidation. Reperfusion-induced death was attenuated by over-expression of Mn-superoxide dismutase (Mn-SOD) or mitochondrial phospholipid hydroperoxide glutathione peroxidase (mito-PHGPx), but not by catalase, mitochondria-targeted catalase, or Cu,Zn-SOD. Protection was also conferred by chemically distinct antioxidant compounds, and mito-roGFP oxidation was attenuated by NAC, or by scavenging of residual O(2) during the ischemia (anoxic ischemia). Mitochondrial permeability transition pore (mPTP) oscillation/opening was monitored by real-time imaging of mitochondrial calcein fluorescence. Oxidant stress caused release of calcein to the cytosol during ischemia, a response that was inhibited by chemically diverse antioxidants, anoxia, or over-expression of Mn-SOD or mito-PHGPx. These findings suggest that mitochondrial oxidant stress causes oscillation of the mPTP prior to reperfusion. Cytochrome c release from mitochondria to the cytosol was not detected until after reperfusion, and was inhibited by anoxic ischemia or antioxidant administration during ischemia. Although DNA fragmentation was detected after I/R, no evidence of Bax activation was detected. Over-expression of the anti-apoptotic protein Bcl-X(L) in cardiomyocytes did not confer protection against I/R-induced cell death. Moreover, murine embryonic fibroblasts with genetic depletion of Bax and Bak, or over-expression of Bcl-X(L), failed to show protection against I/R. These findings indicate that mitochondrial ROS during ischemia triggers mPTP activation, mitochondrial depolarization, and cell death during reperfusion through a Bax/Bak-independent cell death pathway. Therefore, mitochondrial apoptosis appears to represent a redundant death pathway in this model of simulated I/R. This article is part of a Special Issue entitled: Mitochondria and Cardioprotection.

Baicalein Preconditioning Cardioprotection Involves Pro-Oxidant Signaling and Activation of Pyruvate Dehydrogenase.

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.

Part 9: post-cardiac arrest care: 2010 American Heart Association Guidelines for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care.

The goal of immediate post-cardiac arrest care is to optimize systemic perfusion, restore metabolic homeostasis, and support organ system function to increase the likelihood of intact neurological survival. The post-cardiac arrest period is often marked by hemodynamic instability as well as metabolic abnormalities. Support and treatment of acute myocardial dysfunction and acute myocardial ischemia can increase the probability of survival. Interventions to reduce secondary brain injury, such as therapeutic hypothermia, can improve survival and neurological recovery. Every organ system is at risk during this period, and patients are at risk of developing multiorgan dysfunction. The comprehensive treatment of diverse problems after cardiac arrest involves multidisciplinary aspects of critical care, cardiology, and neurology. For this reason, it is important to admit patients to appropriate critical-care units with a prospective plan of care to anticipate, monitor, and treat each of these diverse problems. It is also important to appreciate the relative strengths and weaknesses of different tools for estimating the prognosis of patients after cardiac arrest.

Part 1: executive summary: 2010 American Heart Association Guidelines for Cardiopulmonary Resuscitation and Emergency Cardiovascular Care.

The goal of therapy for bradycardia or tachycardia is to rapidly identify and treat patients who are hemodynamically unstable or symptomatic due to the arrhythmia. Drugs or, when appropriate, pacing may be used to control unstable or symptomatic bradycardia. Cardioversion or drugs or both may be used to control unstable or symptomatic tachycardia. ACLS providers should closely monitor stable patients pending expert consultation and should be prepared to aggressively treat those with evidence of decompensation.

Therapeutic hypothermia cardioprotection in murine hemorrhagic shock/resuscitation differentially affects p38α/p38γ, Akt, and HspB1.

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.

Akt1 genetic deficiency limits hypothermia cardioprotection following murine cardiac arrest.

Therapeutic hypothermia (TH) cardioprotection has recently been associated with increased Akt signaling in a rat model of cardiac arrest. However, it is not known whether Akt is required for this beneficial effect of TH. We used a mouse model of cardiac arrest demonstrating TH cardioprotection to study the response of mice deficient in an Akt1 allele. We hypothesized that Akt1 mediates TH cardioprotection and that decreases in Akt1 content would diminish such protection. Adult C57BL/6 wild-type (WT) mice underwent an 8-min cardiac arrest. After 6 min, the mice were randomized to normothermia (WT(NT), 37 degrees C) or TH (WT(TH), 30 degrees C). Following cardiopulmonary resuscitation and the return of spontaneous circulation (ROSC), the animals were hemodynamically monitored for 240 min (R240). At R240, cardiac tissue Akt content and phosphorylation were assayed. Studies were repeated in Akt1 heterozygous (Akt1(+/-)) mice. As a result, baseline characteristics and ROSC rates were equivalent across groups. At R240, WT(TH) mice exhibited lower heart rate, larger stroke volume, and higher cardiac output than WT(NT) animals (P < 0.05). Cardioprotection in WT(TH) at R240 was associated with increased cardiac Akt phosphorylation at Ser473 and Thr308 compared with that in WT(NT) (P < 0.05). TH-associated alterations in Akt phosphorylation, stroke volume, heart rate, and cardiac output were abrogated in Akt1(+/-) animals. In conclusion, TH improves post-ROSC cardiac function and increases Akt phosphorylation in WT, but not Akt1(+/-), mice. The Akt1 isoform appears necessary for TH-mediated cardioprotection.

Therapeutic hypothermia cardioprotection via Akt- and nitric oxide-mediated attenuation of mitochondrial oxidants.

Therapeutic hypothermia (TH) is a promising cardioprotective treatment for cardiac arrest and acute myocardial infarction, but its cytoprotective mechanisms remain unknown. In this study, we developed a murine cardiomyocyte model of ischemia-reperfusion injury to better determine the mechanisms of TH cardioprotection. We hypothesized that TH manipulates Akt, a survival kinase that mediates mitochondrial protection by modulating reactive oxygen species (ROS) and nitric oxide (NO) generation. Cardiomyocytes, isolated from 1- to 2-day-old C57BL6/J mice, were exposed to 90 min simulated ischemia and 3 h reperfusion. For TH, cells were cooled to 32 degrees C during the last 20 min of ischemia and the first hour of reperfusion. Cell viability was evaluated by propidium iodide and lactate dehydrogenase release. ROS production was measured by 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate and mitochondrial membrane potential (DeltaPsim) by 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazoly-carbocyanine iodide (JC-1). Phospho (p)-Akt (Thr308), p-Akt (Ser473), and phosphorylated heat shock protein 27 (p-HSP27) (Ser82) were analyzed by Western blot analysis. TH attenuated reperfusion ROS generation, increased NO, maintained DeltaPsim, and decreased cell death [19.3 + or - 3.3% (n = 11) vs. 44.7 + or - 2.7% (n = 10), P < 0.001]. TH also increased p-Akt during ischemia before reperfusion. TH protection and attenuation of ROS were blocked by the inhibition of Akt and NO synthase but not by a cGMP inhibitor. HSP27, a regulator of Akt, also exhibited increased phosphorylation (Ser82) during ischemia with TH. We conclude that TH cardioprotection is mediated by enhanced Akt/HSP27 phosphorylation and enhanced NO generation, resulting in the attenuation of ROS generation and the maintenance of DeltaPsim following ischemia-reperfusion.

Evaluation of an Enhanced Peer Mentoring Program on Scholarly Productivity and Promotion in Academic Emergency Medicine: A Five-Year Review.

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.

Baicalein Cardioprotection via Oxidant Scavenging and Akt-Nitric Oxide Signaling: Identification of Early Reperfusion Phase as the Critical Therapeutic Window.

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.

Akt1-mediated CPR cooling protection targets regulators of metabolism, inflammation and contractile function in mouse cardiac arrest.

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.

Cardiac catheterization is underutilized after in-hospital cardiac arrest.

BACKGROUND: Indications for immediate cardiac catheterization in cardiac arrest survivors without ST elevation myocardial infarction (STEMI) are uncertain as electrocardiographic and clinical criteria may be challenging to interpret in this population. We sought to evaluate rates of early catheterization after in-hospital ventricular fibrillation (VF) arrest and the association with survival. METHODS: Using a billing database we retrospectively identified cases with an ICD-9 code of cardiac arrest (427.5) or VF (427.41). Discharge summaries were reviewed to identify in-hospital VF arrests. Rates of catheterization on the day of arrest were determined by identifying billing charges. Unadjusted analyses were performed using Chi-square, and adjusted analyses were performed using logistic regression. RESULTS: One hundred and ten in-hospital VF arrest survivors were included in the analysis. Cardiac catheterization was performed immediately or within 1 day of arrest in 27% (30/110) of patients and of these patients, 57% (17/30) successfully received percutaneous coronary intervention. Of those who received cardiac catheterization the indication for the procedure was STEMI or new left bundle branch block (LBBB) in 43% (13/30). Therefore, in the absence of standard ECG data suggesting acute myocardial infarction, 57% (17/30) received angiography. Patients receiving cardiac catheterization were more likely to survive than those who did not receive catheterization (80% vs. 54%, p<.05). CONCLUSION: In patients receiving cardiac catheterization, more than half received this procedure for indications other than STEMI or new LBBB. Cardiac catheterization was associated with improved survival. Future recommendations need to be established to guide clinicians on which arrest survivors might benefit from immediate catheterization.

Nitric oxide during ischemia attenuates oxidant stress and cell death during ischemia and reperfusion in cardiomyocytes.

Nitric oxide (NO) has been implicated as a cardioprotective agent during ischemia/reperfusion (I/R), but the mechanism of protection is unknown. Oxidant stress contributes to cell death in I/R, so we tested whether NO protects by attenuating oxidant stress. Cardiomyocytes and murine embryonic fibroblasts were administered NO (10-1200 nM) during simulated ischemia, and cell death was assessed during reperfusion without NO. In each case, NO abrogated cell death during reperfusion. Cells overexpressing endothelial NO synthase (NOS) exhibited a similar protection, which was abolished by the NOS inhibitor N(omega)-nitro-l-arginine methyl ester. Protection was not mediated by guanylate cyclase or the mitochondrial K(ATP) channel, as inhibitors of these systems failed to abolish protection. NO did not prevent decreases in mitochondrial potential, but cells protected with NO demonstrated recovery of potential at reperfusion. Measurements using C11-BODIPY reveal that NO attenuates lipid peroxidation during ischemia and reperfusion. Measurements of oxidant stress using the ratiometric redox sensor HSP-FRET demonstrate that NO attenuates protein oxidation during ischemia. These findings reveal that physiological levels of NO during ischemia can attenuate oxidant stress both during ischemia and during reperfusion. This response is associated with a remarkable attenuation of cell death, suggesting that ischemic cell death may be a regulated event.