Monoamine Oxidase-Related Vascular Oxidative Stress in Diseases Associated with Inflammatory Burden.

Monoamine oxidases (MAO) with 2 isoforms, A and B, located at the outer mitochondrial membrane are flavoenzyme membranes with a major role in the metabolism of monoaminergic neurotransmitters and biogenic amines in the central nervous system and peripheral tissues, respectively. In the process of oxidative deamination, aldehydes, hydrogen peroxide, and ammonia are constantly generated as potential deleterious by-products. While being systematically studied for decades as sources of reactive oxygen species in brain diseases, compelling evidence nowadays supports the role of MAO-related oxidative stress in cardiovascular and metabolic pathologies. Indeed, oxidative stress and chronic inflammation are the most common pathomechanisms of the main noncommunicable diseases of our century. MAO inhibition with the new generation of reversible and selective drugs has recently emerged as a pharmacological strategy aimed at mitigating both processes. The aim of this minireview is to summarize available information regarding the contribution of MAO to the vascular oxidative stress and endothelial dysfunction in hypertension, metabolic disorders, and chronic kidney disease, all conditions associated with increased inflammatory burden.

Monoamine oxidase is a source of oxidative stress in obese patients with chronic inflammation 1.

Obesity is an important preventable risk factor for morbidity and mortality from cardiometabolic disease. Oxidative stress (including in visceral adipose tissue) and chronic low-grade inflammation are the major underlying pathomechanisms. Monoamine oxidase (MAO) has recently emerged as an important source of cardiovascular oxidative stress. The present study was conducted to evaluate the role of MAO as contributor to reactive oxygen species (ROS) production in white adipose tissue and vessels harvested from patients undergoing elective abdominal surgery. To this aim, visceral adipose tissue and mesenteric artery branches were isolated from obese patients with chronic inflammation and used for organ bath, ROS production, quantitative real-time PCR, and immunohistology studies. The human visceral adipose tissue and mesenteric artery branches contain mainly the MAO-A isoform, as shown by the quantitative real-time PCR and immunohistology experiments. A significant upregulation of MAO-A, the impairment in vascular reactivity, and increase in ROS production were found in obese vs. non-obese patients. Incubation of the adipose tissue samples and vascular rings with the MAO-A inhibitor (clorgyline, 30 min) improved vascular reactivity and decreased ROS generation. In conclusion, MAO-A is the predominant isoform in human abdominal adipose and vascular tissues, is overexpressed in the setting of inflammation, and contributes to the endothelial dysfunction.

A Comprehensive Assessment of Apigenin as an Antiproliferative, Proapoptotic, Antiangiogenic and Immunomodulatory Phytocompound.

Apigenin (4',5,7-trihydroxyflavone) (Api) is an important component of the human diet, being distributed in a wide number of fruits, vegetables and herbs with the most important sources being represented by chamomile, celery, celeriac and parsley. This study was designed for a comprehensive evaluation of Api as an antiproliferative, proapoptotic, antiangiogenic and immunomodulatory phytocompound. In the set experimental conditions, Api presents antiproliferative activity against the A375 human melanoma cell line, a G2/M arrest of the cell cycle and cytotoxic events as revealed by the lactate dehydrogenase release. Caspase 3 activity was inversely proportional to the Api tested doses, namely 30 µM and 60 µM. Phenomena of early apoptosis, late apoptosis and necrosis following incubation with Api were detected by Annexin V-PI double staining. The flavone interfered with the mitochondrial respiration by modulating both glycolytic and mitochondrial pathways for ATP production. The metabolic activity of human dendritic cells (DCs) under LPS-activation was clearly attenuated by stimulation with high concentrations of Api. Il-6 and IL-10 secretion was almost completely blocked while TNF alpha secretion was reduced by about 60%. Api elicited antiangiogenic properties in a dose-dependent manner. Both concentrations of Api influenced tumour cell growth and migration, inducing a limited tumour area inside the application ring, associated with a low number of capillaries.

Vitamin D improves vascular function and decreases monoamine oxidase A expression in experimental diabetes.

The active form of vitamin D, 1,25-dihydroxycholecalciferol (1,25(OH)2D3), was reported to improve vascular function in patients with diabetes, yet the underlying mechanisms remain to be fully elucidated. Monoamine oxidase (MAO), a mitochondrial enzyme, with two isoforms (A and B) that generates hydrogen peroxide (H2O2) as by-product, has been recently reported to contribute to the pathogenesis of endothelial dysfunction in diabetes. The present study assessed the interaction between vitamin D and MAO in the vascular wall in the setting of type 1 experimental diabetes. To this aim, diabetes was induced in male Wistar rats via a single injection of streptozotocin (STZ, 50 mg/kg, IP) and 1 month later thoracic aortas were harvested and used for organ bath studies and H2O2 measurements. MAO expression was assessed by immunohistochemistry and RT-PCR. Endothelial function was evaluated in isolated aortic rings in the absence vs. presence of 1,25(OH)2D3 (100 nM, 24 h incubation). In diabetic animals, we found a significant reduction in the endothelial-dependent relaxation to acetylcholine and an increased expression of the MAO-A isoform, respectively. Vitamin D significantly improved vascular function, mitigated oxidative stress and decreased MAO-A expression in diabetic vascular preparations. In conclusion, MAO-A is induced in diabetic aortas and vitamin D can improve diabetes-induced endothelial dysfunction by modulating the MAO-A expression.

Monoamine oxidase inhibition improves vascular function and reduces oxidative stress in rats with lipopolysaccharide-induced inflammation.

Oxidative stress and vascular inflammation are the two major pathomechanisms that contribute to the progression of both cardiovascular and metabolic diseases. We have previously demonstrated that monoamine oxidases (MAOs), mitochondrial enzymes with two isoforms (A and B), are contributors to the endothelial dysfunction associated with inflammation in mice. The present study was purported to assess the effects of MAOs on endothelial dysfunction in rats with lipopolysaccharide (LPS)-induced acute inflammation. To this aim, aortas harvested from rats treated or not with a single dose of LPS were used for organ-bath studies of vascular reactivity and H2O2 production assessment in the presence vs. absence of MAO inhibitors. Our results demonstrate that MAO-A and B isoforms are induced in the rat vascular system after LPS administration. Both reversible and irreversible MAOs inhibition improved vascular function and reduced oxidative stress. In conclusion, MAOs are contributors to the occurrence of endothelial dysfunction in the rat model of LPS-induced acute inflammation. MAO inhibition may become a viable therapeutic strategy for the treatment of cardiometabolic disease.

Methylene blue alleviates endothelial dysfunction and reduces oxidative stress in aortas from diabetic rats.

Endothelial dysfunction and the related increase in reactive oxygen species (ROS) production are important events in the pathophysiology of diabetes mellitus (DM). Methylene blue (MB) has been systematically investigated for its protective effects against refractory hypotension and mitochondrial dysfunction. We have previously demonstrated that MB improved mitochondrial respiration and partially decreased oxidative stress in diabetic rat hearts. The present study was aimed to investigate whether MB modulates vascular function and ROS production in thoracic aortic rings isolated from rats with streptozotocin-induced DM (after 4 weeks of hyperglycemia). The effects of MB (0.1 µM, 30 min ex vivo incubation) on vascular reactivity in organ chamber (phenylephrine-induced contraction, acetylcholine-induced relaxation) and H2O2 production (assessed by ferrous iron xylenol orange oxidation assay) were investigated in vascular preparations with intact endothelium and after denudation. DM elicited a significant alteration of vascular function: increased contractility to phenylephrine, attenuation of acetylcholine-dependent relaxation, and augmented H2O2 generation. Ex vivo incubation with MB partially reversed all these changes (by approximately 70%) in vascular segments with intact endothelial layer (but not in denuded vessels). In conclusion, MB might be useful in alleviating endothelial dysfunction and mitigating endothelial oxidative stress, observations that clearly require further investigation in the setting of cardiometabolic disease.

Quercetin exerts an inhibitory effect on cellular bioenergetics of the B164A5 murine melanoma cell line.

Modulation of mitochondrial bioenergetics and glycolysis in malignancies has recently emerged a potential chemotherapeutic strategy since numerous malignant cells have overcome inhibition of the glycolytic pathway by increasing mitochondrial ATP production. Quercetin is a flavonoid with antioxidant, antiangiogenic, and chemoprotective properties but the mitochondrial effects are less characterized. The present study was purported to assess the effects of quercetin on the bioenergetic profile of B164A5 murine melanoma cell line. The oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured at 24, 48, and 72 h of treatment of B164A5 murine melanoma cells with increasing concentrations (25, 50, 100, and 150 µM) of quercetin using the extracellular flux analyzer Seahorse XF24e (Seahorse Agilent). Analysis of mitochondrial function was performed in the presence of the classic modulators of the electron transport chain: oligomycin, FCCP, and rotenone. 72-h treatment with quercetin induced a dose-dependent decrease of all OCR parameters (basal respiration, proton leak, ATP turnover, maximal respiration, reserve capacity) as well as of ECAR. At variance, 48-h treatment induced a decrease of OCR and ECAR when quercetin was applied at 50, 100, and 150 µM, while the 24-h treatment induced a decrease of bioenergetic parameters only for the highest concentrations (100 and 150 µM) of the compound. Our data clearly demonstrated that quercetin elicited dose-dependent inhibitory effect on examined parameters of cellular bioenergetics that was most potent at 72 h of treatment. Thereby quercetin, modulating both glycolytic and mitochondrial pathways for ATP production, might be an efficient approach in killing cancer cells.

Modulation of Cancer Metabolism by Phytochemicals - A Brief Overview.

Despite tremendous research efforts for effective therapies, cancer remains the plague of the century and its burden is expected to increase worldwide in the near future. Metabolic reprogramming is a firmly established hallmark of all cancers, regardless of their cellular or tissue origin, being a prerequisite for both tumor growth and invasion. Functional dependence of tumors on glycolysis and glutaminolysis and the crucial contribution of mitochondria to the tumor bioenergetic versatility are well recognized features and established therapeutic targets. The complex landscape of tumor metabolism in the context of the dynamic, bidirectional crosstalk with its stromal environment is a rapidly evolving field that increasingly supports the view of cancer both as metabolic disease and a disease of impaired cellular 'communication'. Many of the approved anticancer drugs are derived from natural sources and the search of novel drug candidates is still a priority view the rapid development of chemoresistance. Phytochemicals are biologically active plant compounds with preventive and/or curative anticancer properties able to potentiate the effects of standard therapies while decreasing their toxicity via multitarget modulatory effects. The present mini-review will briefly summarize the hallmarks of metabolic reprogramming in tumor cells and the phytochemicals that have been reported to modulate the dysregulated metabolism of tumor and its environment, with special emphasis on triterpenes.

Contribution of monoamine oxidases to vascular oxidative stress in patients with end-stage renal disease requiring hemodialysis.

Arteriovenous fistula (AVF) is the "lifeline" for patients with end-stage renal disease (ESRD) undergoing hemodialysis. AVF maturation failure is a poorly understood process, one of the contributors being endothelial dysfunction due to oxidative stress. Monoamine oxidases (MAOs) A and B were recently identified as novel sources of vascular oxidative stress. The aim of the present study was to assess the contribution of MAOs to the endothelial dysfunction in patients with ESDR with indication of hemodialysis. Fragments of brachial artery collaterals were harvested from ESRD patients during the surgical procedure aimed at creating the vascular access in the cubital fossa. The effect of increasing concentrations (10, 30, 100 µmol/L) of the irreversible MAO-A inhibitor, clorgyline, and MAO-B inhibitor, selegiline, on endothelial-dependent relaxation (EDR) in response to cumulative doses of acetylcholine was studied in isolated phenylephrine-preconstricted vascular rings. Hydrogen peroxide (H2O2) production was assessed using ferrous oxidation xylenol orange assay. We showed that incubation of brachial rings with MAO inhibitors significantly improved EDR and attenuated H2O2 generation in patients with ESRD. MAO-related oxidative stress might contribute to the primary dysfunction/non-maturation of the AVF and MAO inhibitors could improve maturation and long-term patency of the vascular access in dialysis patients.

Methylene blue improves mitochondrial respiration and decreases oxidative stress in a substrate-dependent manner in diabetic rat hearts.

Diabetic cardiomyopathy has been systematically associated with compromised mitochondrial energetics and increased generation of reactive oxygen species (ROS) that underlie its progression to heart failure. Methylene blue is a redox drug with reported protective effects mainly on brain mitochondria. The purpose of the present study was to characterize the effects of acute administration of methylene blue on mitochondrial respiration, H2O2 production, and calcium sensitivity in rat heart mitochondria isolated from healthy and 2 months (streptozotocin-induced) diabetic rats. Mitochondrial respiratory function was assessed by high-resolution respirometry. H2O2 production and calcium retention capacity were measured spectrofluorimetrically. The addition of methylene blue (0.1 µmol·L-1) elicited an increase in oxygen consumption of mitochondria energized with complex I and II substrates in both normal and diseased mitochondria. Interestingly, methylene blue elicited a significant increase in H2O2 release in the presence of complex I substrates (glutamate and malate), but had an opposite effect in mitochondria energized with complex II substrate (succinate). No changes in the calcium retention capacity of healthy or diabetic mitochondria were found in the presence of methylene blue. In conclusion, in cardiac mitochondria isolated from diabetic and nondiabetic rat hearts, methylene blue improved respiratory function and elicited a dichotomic, substrate-dependent effect on ROS production.

The effect of purinergic signaling via the P2Y11 receptor on vascular function in a rat model of acute inflammation.

There is a growing body of evidence pointing to the role of purinergic signaling in the development and progression of various conditions that have inflammation as a common pathogenetic denominator. The aim of the present study was to assess the involvement of P2Y11 purinergic receptors in the regulation of vascular function in aortic segments obtained using an experimental model of acute inflammation, the lipopolysaccharide (LPS, 8 mg/kg, i.p)-treated rats. Twelve hours after LPS administration, thoracic aortas were isolated and used for studies of vascular reactivity in the organ bath and for the measurement of reactive oxygen species (ROS) generation, respectively. LPS treatment significantly increased contractility to phenylephrine and attenuated the endothelium-dependent relaxation of the vascular segments in response to acetylcholine; an increased production of hydrogen peroxide (H2O2) was also recorded. The P2Y11 activator, NF546, decreased the LPS-induced aortic H2O2 release and partially normalized the vasomotor function, namely reduced contractility and improved relaxation. The effect was abolished by co-treatment with the P2Y11 inhibitor, NF340, and also after endothelium denudation. Importantly, NF546 did not elicit an antioxidant effect by acting as a H2O2 scavenger, suggesting that the beneficial outcome of this treatment on the vasculature is the consequence of P2Y11 stimulation. In conclusion, purinergic P2Y11 receptors stimulation improves vascular function and mitigates oxidative stress in the setting of acute systemic inflammation, revealing salutary effects and therapeutic potential in pathologies associated with endothelial dysfunction.

Monoamine oxidase inhibition improves vascular function in mammary arteries from nondiabetic and diabetic patients with coronary heart disease.

Monoamine oxidases (MAOs) are mitochondrial enzymes with 2 isoforms that have emerged as important contributors to cardiovascular oxidative stress via the constant generation of hydrogen peroxide. The present study was purported to assess whether MAO-derived H2O2 contributes to the endothelial dysfunction in mammary arteries harvested from coronary heart disease patients with and without diabetes mellitus subjected to coronary artery bypass grafting. To this aim, the effects of MAO inhibition on vascular contractility to phenylephrine and endothelial-dependent relaxation (EDR) in response to acetylcholine were studied in vascular segments. Clorgyline (irreversible MAO-A inhibitor), selegiline (irreversible MAO-B inhibitor), and moclobemide (reversible MAO-A inhibitor) were applied in the organ bath (10 µmol/L). MAO expression was assessed by immunohistochemistry. We found a constant impairment of EDR that has been significantly attenuated in the presence of the MAO-A and MAO-B inhibitors in both groups of coronary heart disease patients. MAO-B was the dominant isoform in all human diseased vessels. In conclusion, in vitro inhibition of MAO significantly improved EDR in human mammary arteries, regardless of the presence of diabetes. These data suggest that MAO inhibitors might be useful in restoring endothelial response in clinical conditions associated with increased oxidative stress, such as coronary artery disease and diabetes.

The Role of Mitochondrial Reactive Oxygen Species in Cardiovascular Injury and Protective Strategies.

Ischaemia/reperfusion (I/R) injury of the heart represents a major health burden mainly associated with acute coronary syndromes. While timely coronary reperfusion has become the established routine therapy in patients with ST-elevation myocardial infarction, the restoration of blood flow into the previously ischaemic area is always accompanied by myocardial injury. The central mechanism involved in this phenomenon is represented by the excessive generation of reactive oxygen species (ROS). Besides their harmful role when highly generated during early reperfusion, minimal ROS formation during ischaemia and/or at reperfusion is critical for the redox signaling of cardioprotection. In the past decades, mitochondria have emerged as the major source of ROS as well as a critical target for cardioprotective strategies at reperfusion. Mitochondria dysfunction associated with I/R myocardial injury is further described and ultimately analyzed with respect to its role as source of both deleterious and beneficial ROS. Furthermore, the contribution of ROS in the highly investigated field of conditioning strategies is analyzed. In the end, the vascular sources of mitochondria-derived ROS are briefly reviewed.

Acute inhibition of monoamine oxidase and ischemic preconditioning in isolated rat hearts: interference with postischemic functional recovery but no effect on infarct size reduction.

Monoamine oxidases (MAOs) have recently emerged as important mitochondrial sources of oxidative stress in the cardiovascular system. Generation of reactive oxygen species during the brief episodes of ischemic preconditioning (IPC) is responsible for the cardioprotection at reperfusion. The aim of this study was to assess the effects of two MAO inhibitors (clorgyline and pargyline) on the IPC-related protection in isolated rat hearts. Animals subjected to 30 min global ischemia and 120 min reperfusion were assigned to the following groups: (i) Control, no additional intervention; (ii) IPC, 3 cycles of 5 min ischemia and 5 min reperfusion before the index ischemia; (iii) IPC-clorgyline, IPC protocol bracketed for 5 min with clorgyline (50 µmol/L); (iv) IPC-pargyline, IPC protocol bracketed for 5 min with pargyline (0.5 mmol/L). The postischemic functional recovery was assessed by the left ventricular developed pressure (LVDP) and the indices of contractility (+dLVP/dt max) and relaxation (-dLVP/dt max). Infarct size (IS) was quantified by TTC staining. In both genders, IPC significantly improved functional recovery that was further enhanced in the presence of either clorgyline or pargyline. IS reduction was comparable among all the preconditioned groups, regardless of the presence of MAO inhibitors. In isolated rat hearts, acute inhibition of MAOs potentiates the IPC-induced postischemic functional recovery without interfering with the anti-necrotic protection.

Modulation of mitochondrial respiratory function and ROS production by novel benzopyran analogues.

A substantial body of evidence indicates that pharmacological activation of mitochondrial ATP-sensitive potassium channels (mKATP) in the heart is protective in conditions associated with ischemia/reperfusion injury. Several mechanisms have been postulated to be responsible for cardioprotection, including the modulation of mitochondrial respiratory function. The aim of the present study was to characterize the dose-dependent effects of novel synthetic benzopyran analogues, derived from a BMS-191095, a selective mKATP opener, on mitochondrial respiration and reactive oxygen species (ROS) production in isolated rat heart mitochondria. Mitochondrial respiratory function was assessed by high-resolution respirometry, and H2O2 production was measured by the Amplex Red fluorescence assay. Four compounds, namely KL-1487, KL-1492, KL-1495, and KL-1507, applied in increasing concentrations (50, 75, 100, and 150 µmol/L, respectively) were investigated. When added in the last two concentrations, all compounds significantly increased State 2 and 4 respiratory rates, an effect that was not abolished by 5-hydroxydecanoate (5-HD, 100 µmol/L), the classic mKATP inhibitor. The highest concentration also elicited an important decrease of the oxidative phosphorylation in a K(+) independent manner. Both concentrations of 100 and 150 µmol/L for KL-1487, KL-1492, and KL-1495, and the concentration of 150 µmol/L for KL-1507, respectively, mitigated the mitochondrial H2O2 release. In isolated rat heart mitochondria, the novel benzopyran analogues act as protonophoric uncouplers of oxidative phosphorylation and decrease the generation of reactive oxygen species in a dose-dependent manner.

BIOENERGETIC CHARACTERIZATION OF H9C2 CELLS USING THE EXTRACELLULAR FLUX ANALYZER.

UNLABELLED: The aim of the present work was to standardize the working methodology for assessing the bioenergetic profile of H9c2 cardiomyoblasts cells, with reference to the optimization of cell seeding number and the establishment of favorable concentrations for the classic modulators of mitochondrial respiratory function, in particular the one of a classical uncoupler, FCCP. MATERIAL AND METHODS: The extracellular flux analyzer (XF, Seahorse Bioscience) is a novel high-throughput instrument able to monitor the metabolism of living cells by simultaneously measuring mitochondrial respiration and glycolysis. The in vitro platform will be further used to better understand the pathophysiology and the unrecognized side effects of drugs currently used in the therapy of major cardiovascular diseases. CONCLUSIONS: In the long run, characterization of novel pharmacological agents' effects on other cell lines, including tumoral ones, will be also considered.

Monoamine Oxidases as Potential Contributors to Oxidative Stress in Diabetes: Time for a Study in Patients Undergoing Heart Surgery.

Oxidative stress is a pathomechanism causally linked to the progression of chronic cardiovascular diseases and diabetes. Mitochondria have emerged as the most relevant source of reactive oxygen species, the major culprit being classically considered the respiratory chain at the inner mitochondrial membrane. In the past decade, several experimental studies unequivocally demonstrated the contribution of monoamine oxidases (MAOs) at the outer mitochondrial membrane to the maladaptative ventricular hypertrophy and endothelial dysfunction. This paper addresses the contribution of mitochondrial dysfunction to the pathogenesis of heart failure and diabetes together with the mounting evidence for an emerging role of MAO inhibition as putative cardioprotective strategy in both conditions.

Monoamine oxidases are novel sources of cardiovascular oxidative stress in experimental diabetes.

Diabetes mellitus (DM) is widely recognized as the most severe metabolic disease associated with increased cardiovascular morbidity and mortality. The generation of reactive oxygen species (ROS) is a major event causally linked to the development of cardiovascular complications throughout the evolution of DM. Recently, monoamine oxidases (MAOs) at the outer mitochondrial membrane, with 2 isoforms, MAO-A and MAO-B, have emerged as novel sources of constant hydrogen peroxide (H2O2) production in the cardiovascular system via the oxidative deamination of biogenic amines and neurotransmitters. Whether MAOs are mediators of endothelial dysfunction in DM is unknown, and so we studied this in a streptozotocin-induced rat model of diabetes. MAO expression (mRNA and protein) was increased in both arterial samples and hearts isolated from the diabetic animals. Also, H2O2 production (ferrous oxidation - xylenol orange assay) in aortic samples was significantly increased, together with an impairment of endothelium-dependent relaxation (organ-bath studies). MAO inhibitors (clorgyline and selegiline) attenuated ROS production by 50% and partially normalized the endothelium-dependent relaxation in diseased vessels. In conclusion, MAOs, in particular the MAO-B isoform, are induced in aortas and hearts in the streptozotocin-induced diabetic rat model and contribute, via the generation of H2O2, to the endothelial dysfunction associated with experimental diabetes.

Betulinic acid as a potent and complex antitumor phytochemical: a minireview.

Betulinic acid (BA), a natural compound with a lupan skeleton, has been highly investigated in the past decade for a plethora of beneficial properties, including anti-cancer, anti-inflammatory, anti-angiogenic, immune-modulatory, and anti-HIV effects. In particular, BA has been reported to be effective in vitro against tumor cell lines of different origins, and also in vivo, in animal models of cancer. The best characterized mechanism of its antitumor effect consists of triggering apoptosis via the mitochondrial pathway. BA has also an anti-metastatic effect via the prevention of the epithelial-to-mesencymal transition in highly aggressive melanoma cells. Furthermore, in the same model, BA is able to counteract the pro-invasive potential of the pro-tumoral protein neutrophil gelatinaseassociated lipocalin. The present review addresses the current state of knowledge regarding the anti-tumor effects of betulinic acid, a potent chemotherapeutic agent.

Activation of prosurvival signaling pathways during the memory phase of volatile anesthetic preconditioning in human myocardium: a pilot study.

According to a compelling body of evidence anesthetic preconditioning (APC) attenuates the deleterious consequences of ischemia-reperfusion and protects the heart through a mechanism similar to ischemic preconditioning. The present study was purported to investigate the intracellular signaling pathways activated in human myocardium in response to a preconditioning protocol with two different volatile anesthetics, namely isoflurane and sevoflurane. To this aim, phosphorylation of PKCα and -δ, ERK1/2, Akt, and GSK3ß was determined at the end of the APC protocol, in human atrial samples harvested from patients undergoing open-heart surgery. The results demonstrate that preconditioning with volatile anesthetics triggers the activation of PKCδ and -α isoforms and of prosurvival kinases, ERK1/2, and Akt, while inhibiting their downstream target GSK3ß during the memory phase.