Cardioprotection by Poloxamer 188 is Mediated through Increased Endothelial Nitric Oxide Production.

Ischemia/reperfusion (I/R) injury significantly contributes to the morbidity and mortality associated with cardiac events. Poloxamer 188 (P188), a nonionic triblock copolymer, has been proposed to mitigate I/R injury by stabilizing cell membranes. However, the underlying mechanisms remain incompletely understood, particularly concerning endothelial cell function and nitric oxide (NO) production. We employed human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) and endothelial cells (ECs) to elucidate the effects of P188 on cellular survival, function, and NO secretion under simulated I/R conditions. iPSC-CMs contractility and iPSC-ECs' NO production were assessed following exposure to P188. Further, an isolated heart model using Brown Norway rats subjected to I/R injury was utilized to evaluate the ex-vivo cardioprotective effects of P188, examining cardiac function and NO production, with and without the administration of a NO inhibitor. In iPSC-derived models, P188 significantly preserved CM contractile function and enhanced cell viability after hypoxia/reoxygenation. Remarkably, P188 treatment led to a pronounced increase in NO secretion in iPSC-ECs, a novel finding demonstrating endothelial protective effects beyond membrane stabilization. In the rat isolated heart model, administration of P188 during reperfusion notably improved cardiac function and reduced I/R injury markers. This cardioprotective effect was abrogated by NO inhibition, underscoring the pivotal role of NO. Additionally, a dose-dependent increase in NO production was observed in non-ischemic rat hearts treated with P188, further establishing the critical function of NO in P188 induced cardioprotection. In conclusion, our comprehensive study unveils a novel role of NO in mediating the protective effects of P188 against I/R injury. This mechanism is evident in both cellular models and intact rat hearts, highlighting the potential of P188 as a therapeutic agent against I/R injury. Our findings pave the way for further investigation into P188's therapeutic mechanisms and its potential application in clinical settings to mitigate I/R-related cardiac dysfunction.

Ischemic Post-Conditioning in a Rat Model of Asphyxial Cardiac Arrest.

BACKGROUND: Ischemic post-conditioning (IPoC) has been shown to improve outcomes in limited pre-clinical models. As down-time is often unknown, this technique needs to be investigated over a range of scenarios. As this tool limits reperfusion injury, there may be limited benefit or even harm after short arrest and limited ischemia-reperfusion injury. METHODS: Eighteen male Wistar rats underwent 7 min of asphyxial arrest. Animals randomized to IPoC received a 20 s pause followed by 20 s of compressions, repeated four times, initiated 40 s into cardiopulmonary resuscitation. If return of spontaneous circulation (ROSC) was achieved, epinephrine was titrated to mean arterial pressure (MAP) of 70 mmHg. Data were analyzed using t-test or Mann-Whitney test. Significance set at p ≤ 0.05. RESULTS: The rate of ROSC was equivalent in both groups, 88%. There was no statistically significant difference in time to ROSC, epinephrine required post ROSC, carotid flow, or peak lactate at any timepoint. There was a significantly elevated MAP with IPoC, 90.7 mmHg (SD 13.9), as compared to standard CPR, 76.7 mmHg (8.5), 2 h after ROSC, p = 0.03. CONCLUSIONS: IPoC demonstrated no harm in a model of short arrest using a new arrest etiology for CPR based IPoC intervention in a rat model.

Newly Developed Di-Block Copolymer-Based Cell Membrane Stabilizers Protect Mouse Coronary Artery Endothelial Cells against Hypoxia/Reoxygenation Injury.

Reperfusion after ischemia causes additional cellular damage, known as reperfusion injury, for which there is still no effective remedy. Poloxamer (P)188, a tri-block copolymer-based cell membrane stabilizer (CCMS), has been shown to provide protection against hypoxia/reoxygenation (HR) injury in various models by reducing membrane leakage and apoptosis and improving mitochondrial function. Interestingly, substituting one of its hydrophilic poly-ethylene oxide (PEO) blocks with a (t)ert-butyl terminus added to the hydrophobic poly-propylene oxide (PPO) block yields a di-block compound (PEO-PPOt) that interacts better with the cell membrane lipid bi-layer and exhibits greater cellular protection than the gold standard tri-block P188 (PEO75-PPO30-PEO75). For this study, we custom-made three different new di-blocks (PEO113-PPO10t, PEO226-PPO18t and PEO113-PPO20t) to systemically examine the effects of the length of each polymer block on cellular protection in comparison to P188. Cellular protection was assessed by cell viability, lactate dehydrogenase release, and uptake of FM1-43 in mouse artery endothelial cells (ECs) following HR injury. We found that di-block CCMS were able to provide the same or better EC protection than P188. Our study provides the first direct evidence that custom-made di-block CCMS can be superior to P188 in improving EC membrane protection, raising their potential in treating cardiac reperfusion injury.

Design and Implementation of a Rat Ex Vivo Lung Perfusion Model.

Ex vivo lung preparations are a useful model that can be translated to many different fields of research, complementing corresponding in vivo and in vitro models. Laboratories wishing to use isolated lungs need to be aware of important steps and inherent challenges to establish a setup that is affordable, reliable, and that can be easily adapted to fit the topic of interest. This paper describes a DIY (do it yourself) model for ex vivo rat lung ventilation and perfusion to study drug and gas effects on pulmonary vascular tone, independent of changes in cardiac output. Creating this model includes a) the design and construction of the apparatus, and b) the lung isolation procedure. This model results in a setup that is more cost-effective than commercial alternatives and yet modular enough to adapt to changes in specific research questions. Various obstacles had to be resolved to ensure a consistent model that is capable of being used for a variety of different research topics. Once established, this model has proven to be highly adaptable to different questions and can easily be altered for different fields of study.

Peripheral Intravenous Waveform Analysis Responsiveness to Subclinical Hemorrhage in a Rat Model.

BACKGROUND: Early detection and quantification of perioperative hemorrhage remains challenging. Peripheral intravenous waveform analysis (PIVA) is a novel method that uses a standard intravenous catheter to detect interval hemorrhage. We hypothesize that subclinical blood loss of 2% of the estimated blood volume (EBV) in a rat model of hemorrhage is associated with significant changes in PIVA. Secondarily, we will compare PIVA association with volume loss to other static, invasive, and dynamic markers. METHODS: Eleven male Sprague Dawley rats were anesthetized and mechanically ventilated. A total of 20% of the EBV was removed over ten 5 minute-intervals. The peripheral intravenous pressure waveform was continuously transduced via a 22-G angiocatheter in the saphenous vein and analyzed using MATLAB. Mean arterial pressure (MAP) and central venous pressure (CVP) were continuously monitored. Cardiac output (CO), right ventricular diameter (RVd), and left ventricular end-diastolic area (LVEDA) were evaluated via transthoracic echocardiogram using the short axis left ventricular view. Dynamic markers such as pulse pressure variation (PPV) were calculated from the arterial waveform. The primary outcome was change in the first fundamental frequency (F1) of the venous waveform, which was assessed using analysis of variance (ANOVA). Mean F1 at each blood loss interval was compared to the mean at the subsequent interval. Additionally, the strength of the association between blood loss and F1 and each other marker was quantified using the marginal R2 in a linear mixed-effects model. RESULTS: PIVA derived mean F1 decreased significantly after hemorrhage of only 2% of the EBV, from 0.17 to 0.11 mm Hg, P = .001, 95% confidence interval (CI) of difference in means 0.02 to 0.10, and decreased significantly from the prior hemorrhage interval at 4%, 6%, 8%, 10%, and 12%. Log F1 demonstrated a marginal R2 value of 0.57 (95% CI 0.40-0.73), followed by PPV 0.41 (0.28-0.56) and CO 0.39 (0.26-0.58). MAP, LVEDA, and systolic pressure variation displayed R2 values of 0.31, and the remaining predictors had R2 values ≤0.2. The difference in log F1 R2 was not significant when compared to PPV 0.16 (95% CI -0.07 to 0.38), CO 0.18 (-0.06 to 0.04), or MAP 0.25 (-0.01 to 0.49) but was significant for the remaining markers. CONCLUSIONS: The mean F1 amplitude of PIVA was significantly associated with subclinical blood loss and most strongly associated with blood volume among the markers considered. This study demonstrates feasibility of a minimally invasive, low-cost method for monitoring perioperative blood loss.

High central venous pressure amplitude predicts successful defibrillation in a porcine model of cardiac arrest.

AIM: Increasing venous return during cardiopulmonary resuscitation (CPR) has been shown to improve hemodynamics during CPR and outcomes following cardiac arrest (CA). We hypothesized that a high central venous pressure amplitude (CVP-A), the difference between the maximum and minimum central venous pressure during chest compressions, could serve as a robust predictor of return of spontaneous circulation (ROSC) in addition to traditional measurements of coronary perfusion pressure (CPP) and end-tidal CO2 (etCO2) in a porcine model of CA. METHODS: After 10 min of ventricular fibrillation, 9 anesthetized and intubated female pigs received mechanical chest compressions with active compression/decompression (ACD) and an impedance threshold device (ITD). CPP, CVP-A and etCO2 were measured continuously. All groups received biphasic defibrillation (200 J) at minute 4 of CPR and were classified into two groups (ROSC, NO ROSC). Mean values were analyzed over 3 min before defibrillation by repeated-measures Analysis of Variance and receiver operating characteristic (ROC). RESULTS: Five animals out of 9 experienced ROSC. CVP-A showed a statistically significant difference (p = 0.003) between the two groups during 3 min of CPR before defibrillation compared to CPP (p = 0.056) and etCO2 (p = 0.064). Areas-under-the-curve in ROC analysis for CVP-A, CPP and etCO2 were 0.94 (95% Confidence Interval 0.86, 1.00), 0.74 (0.54, 0.95) and 0.78 (0.50, 1.00), respectively. CONCLUSION: In our study, CVP-A was a potentially useful predictor of successful defibrillation and return of spontaneous circulation. Overall, CVP-A could serve as a marker for prediction of ROSC with increased venous return and thereby monitoring the beneficial effects of ACD and ITD.

P188 Therapy in In Vitro Models of Traumatic Brain Injury.

Traumatic brain injury (TBI) is a significant cause of morbidity and mortality worldwide. Varied mechanisms of injury contribute to the heterogeneity of this patient population as demonstrated by the multiple published grading scales and diverse required criteria leading to diagnoses from mild to severe. TBI pathophysiology is classically separated into a primary injury that is characterized by local tissue destruction as a result of the initial blow, followed by a secondary phase of injury constituted by a score of incompletely understood cellular processes including reperfusion injury, disruption to the blood-brain barrier, excitotoxicity, and metabolic dysregulation. There are currently no effective pharmacological treatments in the wide-spread use for TBI, in large part due to challenges associated with the development of clinically representative in vitro and in vivo models. Poloxamer 188 (P188), a Food and Drug Administration-approved amphiphilic triblock copolymer embeds itself into the plasma membrane of damaged cells. P188 has been shown to have neuroprotective properties on various cell types. The objective of this review is to provide a summary of the current literature on in vitro models of TBI treated with P188.

Perioperative implications of patients with alpha gal allergies.

Alpha Gal Syndrome (AGS) is an emerging immune response to mammalian products (MP) containing the oligosaccharide galactose-α-1,3 galactose (α-Gal) which includes meats and inactive ingredients in certain medications. This becomes clinically important in the perioperative realm as MPs are commonly found in the operating room, and pre- and post-operative settings, and can trigger responses as severe as anaphylaxis. In this review, authors discuss the epidemiology, diagnosis and treatment of AGS reactions. Additionally, strategies are explored in order to screen and prevent exposure to MP with a multidisciplinary approach. While this emerging allergy is still not fully understood, it is of paramount importance that all anesthesia providers recognize the implications of MP exposure in AGS patients and ultimately prevent harm in this highly vulnerable population.

Venous waveform analysis detects acute right ventricular failure in a rat respiratory arrest model.

BACKGROUND: Peripheral intravenous analysis (PIVA) has been shown to be more sensitive than central venous pressure (CVP) for detecting hemorrhage and volume overload. We hypothesized that PIVA is superior to CVP for detecting right ventricular (RV) failure in a rat model of respiratory arrest. METHODS: Eight Wistar rats were studied in accordance with the ARRIVE guidelines. CVP, mean arterial pressure (MAP), and PIVA were recorded. Respiratory arrest was achieved with IV Rocuronium. PIVA utilizes Fourier transform to quantify the amplitude of the peripheral venous waveform, expressed as the "f1 amplitude". RV diameter was measured with transthoracic echocardiography. RESULTS: RV diameter increased from 0.34 to 0.54 cm during arrest, p = 0.001, and returned to 0.33 cm post arrest, p = 0.97. There was an increase in f1 amplitude from 0.07 to 0.38 mmHg, p = 0.01 and returned to 0.08 mmHg, p = 1.0. MAP decreased from 119 to 67 mmHg, p = 0.004 and returned to 136 mmHg, p = 0.50. There was no significant increase in CVP from 9.3 mmHg at baseline to 10.5 mmHg during respiratory arrest, p = 0.91, and recovery to 8.6 mmHg, p = 0.81. CONCLUSIONS: This study highlights the utility of PIVA to detect RV failure in small-caliber vessels, comparable to peripheral veins in the human pediatric population. IMPACT: Right ventricular failure remains a diagnostic challenge, particularly in pediatric patients with small vessel sizes limiting invasive intravascular monitor use. Intravenous analysis has shown promise in detecting hypovolemia and volume overload. Intravenous analysis successfully detects right ventricular failure in a rat respiratory arrest model. Intravenous analysis showed utility despite utilizing small peripheral venous access and therefore may be applicable to a pediatric population. Intravenous analysis may be helpful in differentiating various types of shock.

Buffer glucose adjustment affects myocardial function after ischemia-reperfusion in long-term diabetic rat isolated hearts.

Due to its comorbidities type 2 diabetes mellitus (T2DM) and hypertension, the Zucker Spontaneous Hypertensive Fatty (ZSF1) rat is a clinically relevant animal model when assessing ischemia-reperfusion (IR) injury. Most IR studies in hearts isolated from diabetic animals have been conducted at normal glucose concentrations, providing a different environment compared to in-vivo. We hypothesized IR injury to be attenuated in isolated hearts of diabetic ZSF1 rats when adjusting the Krebs-buffer (KB) to their in-vivo, i.e., elevated blood glucose (BG) levels. Diabetic and non-diabetic ZSF1 rats were anesthetized, hearts isolated and Langendorff-prepared. While standard KB was used for the non-diabetic and diabetic unadjusted groups, KB with glucose levels increased to each rat's prior BG level was used for the adjusted diabetic group. All hearts underwent 30 min ischemia and 120 min reperfusion. Diastolic contracture during ischemia and early reperfusion was delayed and temporarily attenuated in the adjusted compared to the unadjusted diabetic and the non-diabetic groups. The decrease in coronary flow on reperfusion was attenuated in diabetic animals. Left ventricular developed pressure and contractility were not different among the three groups. Infarct size was significantly lower in non-diabetic animals; buffer adjustment made no difference in diabetic animals. In our study, T2DM did not worsen myocardial function in ZSF1 rat isolated hearts. Since our results reveal that hearts with an adjusted glucose level exhibit an at least temporary improvement of function following IR, further studies should consider adapting glucose levels to create more realistic conditions in isolated, perfused hearts.

Differential Effects of Reperfusion on Cardiac Mitochondrial Subpopulations in a Preclinical Porcine Model of Acute Myocardial Infarction.

Acute myocardial infarction (AMI) leads to localized cardiac ischemia and can be fatal if untreated. Despite being treatable, the threat of ischemia-reperfusion (IR) injury remains high. Mitochondria are central to both propagation and mitigation of IR injury, and cardiac mitochondria are categorized into two major subtypes-subsarcolemmal and interfibrillar mitochondria (SSM and IFM, respectively). We hypothesized that, in our pre-clinical porcine model of AMI, SSM and IFM are differentially affected by reperfusion. AMI was induced in female pigs by balloon occlusion of the left anterior descending artery for 45 min, followed by 4 h of reperfusion. At the end of reperfusion, animals were euthanized. Cardiac SSM and IFM from the affected ischemic area and a nearby non-ischemic area were isolated to compare mitochondrial function using substrates targeting mitochondrial electron transport chain complexes I and II. Despite detecting overall significant differences in mitochondrial function including yield, mitochondrial S3 and S4 respirations, and calcium retention, consistent individual functional differences in the two mitochondrial subpopulations were not observed, both between the two mitochondrial subtypes, as well as between the ischemic and non-ischemic tissue. Nonetheless, this study describes the mitochondrial subtype response within the initial few hours of reperfusion in a clinically relevant model of AMI, which provides valuable information needed to develop novel mitochondrially targeted therapies for AMI.

Simulated traumatic brain injury in in-vitro mouse neuronal and brain endothelial cell culture models.

Traumatic brain injury can lead to fatal outcomes such as disability and death. Every year, it affects many patients all over the world. Not only the primary ischemic event, but also the subsequent reperfusion can cause severe brain injury. This so-called ischemia/reperfusion injury combined with mechanical forces lead to cellular disruption. Hence, this paper describes a special in-vitro model, mimicking traumatic brain injury by combining both hypoxia/reoxygenation and compression to simulate ischemia/reperfusion injury as well as the mechanical effects that occur concurrently when suffering traumatic brain injury. Through this approach, stroke, concussion, and traumatic brain injury can be studied on different cell lines in a simplified way. We used two primary mouse brain cell cultures, namely neurons and endothelial cells. Our results show that for the different cell types, different timelines of hypoxia and compression need to be explored to achieve the optimal amount of cellular damage in order to effectively mimic traumatic brain injury. Thus, this model will be useful to test potential treatments of brain injury in future in-vitro studies.

Development of a Cell Co-Culture Model to Mimic Cardiac Ischemia/Reperfusion In Vitro.

Ischemic heart disease is the leading cause of death and disability worldwide. Reperfusion causes additional injury beyond ischemia. Endothelial cells (ECs) can protect cardiomyocytes (CMs) from reperfusion injury through cell-cell interactions. Co-cultures can help investigate the role of cell-cell interactions. A mixed co-culture is the simplest approach but is limited as isolated treatments and downstream analyses of single cell types are not feasible. To investigate whether ECs can dose-dependently attenuate CM cell damage and whether this protection can be further optimized by varying the contact distance between the two cell lines, we used Mouse Primary Coronary Artery Endothelial Cells and Adult Mouse Cardiomyocytes to test three types of cell culture inserts which varied in their inter-cell layer distance at 0.5, 1.0, and 2.0 mm, respectively. In CMs-only, cellular injury as assessed by lactate dehydrogenase (LDH) release increased significantly during hypoxia and further upon reoxygenation when the distance was 2.0 mm compared to 0.5 and 1.0 mm. When ECs and CMs were in nearly direct contact (0.5 mm), there was only a mild attenuation of the reoxygenation injury of CMs following hypoxia. This attenuation was significantly increased when the spatial distance was 1.0 mm. With 2.0 mm distance, ECs attenuated CM injury during both hypoxia and hypoxia/reoxygenation, indicating that sufficient culture distancing is necessary for ECs to crosstalk with CMs, so that secreted signal molecules can circulate and fully stimulate protective pathways. Our findings suggest, for the first time, that optimizing the EC/CM co-culture spatial environment is necessary to provide a favorable in vitro model for testing the role of ECs in CM-protection against simulated ischemia/reperfusion injury. The goal of this report is to provide a step-by-step approach for investigators to use this important model to their advantage.

Poloxamer 188 Exerts Direct Protective Effects on Mouse Brain Microvascular Endothelial Cells in an In Vitro Traumatic Brain Injury Model.

Traumatic Brain Injury (TBI), the main contributor to morbidity and mortality worldwide, can disrupt the cell membrane integrity of the vascular endothelial system, endangering blood-brain barrier function and threatening cellular subsistence. Protection of the vascular endothelial system might enhance clinical outcomes after TBI. Poloxamer 188 (P188) has been shown to improve neuronal function after ischemia/reperfusion (I/R) injury as well as after TBI. We aimed to establish an in vitro compression-type TBI model, comparing mild-to-moderate and severe injury, to observe the direct effects of P188 on Mouse Brain Microvascular Endothelial Cells (MBEC). Confluent MBEC were exposed to normoxic or hypoxic conditions for either 5 or 15 h (hours). 1 h compression was added, and P188 was administered during 2 h reoxygenation. A direct effect of P188 on MBEC was tested by assessing cell number/viability, cytotoxicity/membrane damage, metabolic activity, and total nitric oxide production (tNOp). While P188 enhanced cell number/viability, metabolic activity, and tNOp, an increase in cytotoxicity/membrane damage after mild-to-moderate injury was prevented. In severely injured MBEC, P188 improved metabolic activity only. P188, present during reoxygenation, influenced MBEC function directly in simulated I/R and compression-type TBI.

No Direct Postconditioning Effect of Poloxamer 188 on Mitochondrial Function after Ischemia Reperfusion Injury in Rat Isolated Hearts.

Myocardial infarction is a leading cause for morbidity and mortality worldwide. The only viable treatment for the ischemic insult is timely reperfusion, which further exacerbates myocardial injury. Maintaining mitochondrial function is crucial in preserving cardiomyocyte function in ischemia reperfusion (IR) injury. Poloxamer (P) 188 has been shown to improve cardiac IR injury by improving cellular and mitochondrial function. The aim of this study was to show if P188 postconditioning has direct protective effects on mitochondrial function in the heart. Langendorff prepared rat hearts were subjected to IR injury ex-vivo and reperfused for 10 min with 1 mM P188 vs. vehicle. Cardiac mitochondria were isolated with 1 mM P188 vs. 1 mM polyethylene glycol (PEG) vs. vehicle by differential centrifugation. Mitochondrial function was assessed by adenosine triphosphate synthesis, oxygen consumption, and calcium retention capacity. Mitochondrial function decreased significantly after ischemia and showed mild improvement with reperfusion. P188 did not improve mitochondrial function in the ex-vivo heart, and neither further P188 nor PEG induced direct mitochondrial protection after IR injury in this model.

Evaluation of In Vitro Neuronal Protection by Postconditioning with Poloxamer 188 Following Simulated Traumatic Brain Injury.

Traumatic brain injury (TBI) leads to morbidity and mortality worldwide. Reperfusion after ischemia adds detrimental injury to cells. Ischemia/reperfusion (I/R) injures cells in a variety of ways including cell membrane disruption. Hence, methods to improve endogenous membrane resealing capacity are crucial. Poloxamer (P) 188, an amphiphilic triblock copolymer, was found to be effective against I/R and mechanical injury in various experimental settings. The aim of this study was to establish an in vitro mouse neuronal TBI model and, further, to investigate if postconditioning with P188 directly interacts with neurons after compression and simulated I/R injury, when administered at the start of reoxygenation. Cellular function was assessed by cell number/viability, mitochondrial viability, membrane damage by lactated dehydrogenase (LDH) release and FM1-43 incorporation as well as apoptosis-activation by Caspase 3. Five hours hypoxia ± compression with 2 h reoxygenation proved to be a suitable model for TBI. Compared to normoxic cells not exposed to compression, cell number and mitochondrial viability decreased, whereas membrane injury by LDH release/FM1-43 dye incorporation and Caspase 3 activity increased in cells exposed to hypoxic conditions with compression followed by reoxygenation. P188 did not protect neurons from simulated I/R and/or compression injury. Future research is indicated.

Liver and Biliary Disease of Pregnancy and Anesthetic Implications: A Review.

Liver and biliary disease complicates pregnancy in varying degrees of severity to the mother and fetus, and anesthesiologists may be asked to assist in caring for these patients before, during, and after birth of the fetus. Therefore, it is important to be familiar with how different liver diseases impact the pregnancy state. In addition, knowing symptoms, signs, and laboratory markers in the context of a pregnant patient will lead to faster diagnosis and treatment of such patients. This review article discusses changes in physiology of parturients, patients with liver disease, and parturients with liver disease. Next, general treatment of parturients with acute and chronic liver dysfunction is presented. The article progresses to specific liver diseases with treatments as they relate to pregnancy. And finally, important aspects to consider when anesthetizing parturients with liver disease are discussed.

Potential Effects of Poloxamer 188 on Rat Isolated Brain Mitochondria after Oxidative Stress In Vivo and In Vitro.

Outcome after cerebral ischemia is often dismal. Reperfusion adds significantly to the ischemic injury itself. Therefore, new strategies targeting ischemia/reperfusion (I/R) injury are critically needed. Poloxamer (P)188, an amphiphilic triblock copolymer, is a highly promising pharmacological therapeutic as its capability to insert into injured cell membranes has been reported to protect against I/R injury in various models. Although mitochondrial function particularly profits from P188 treatment after I/R, it remains unclear if this beneficial effect occurs directly or indirectly. Here, rat isolated brain mitochondria underwent oxidative stress in vivo by asphyxial cardiac arrest or in vitro by the addition of hydrogen peroxide (H2O2) after isolation. Mitochondrial function was assessed by adenosine triphosphate synthesis, oxygen consumption, and calcium retention capacity. Both asphyxia and H2O2 exposure significantly impaired mitochondrial function. P188 did not preserve mitochondrial function after either injury mechanism. Further research is indicated.