Goal Directed Perfusion Is Not Associated with a Decrease in Acute Kidney Injury in Patients Predicted to Be at High Risk for Acute Renal Failure after Cardiac Surgery.

Small increases in serum creatinine postoperatively reflect an acute kidney injury (AKI) that likely occurred during cardiopulmonary bypass (CPB). Maintaining adequate oxygen delivery (DO2) during CPB, known as GDP (goal-directed perfusion), improves outcomes. Whether GDP improves outcomes of patients at high risk for acute renal failure (ARF) is unknown. Forty-seven adult patients undergoing cardiac surgery with CPB utilizing GDP with Cleveland Clinic Acute Renal Failure Score of 3 or greater were compared with a matched cohort of patients operated upon using a flow-directed strategy. CPB flow in the GDP cohort was based on a DO2 goal of 260 mL/min/m2. Serum creatinine values were used to determine whether postoperative AKI occurred according to AKIN (Acute Kidney Injury Network) guidelines. We examined the distribution of all variables using proportions for categorical variables and means (standard deviations) for continuous variables and compared treatment groups using t tests for categorical variables and tests for differences in distributions for continuous and count variables. We used inverse probability of treatment weighting to adjust for treatment selection bias. In adjusted models, GDP was not associated with a decrease in AKI (odds ratio [OR]: .97; confidence interval [CI]: .62, 1.52), but was associated with higher odds of ARF (OR: 3.13; CI: 1.26, 7.79), mortality (OR: 3.35; CI: 1.14, 9.89), intensive care unit readmission (OR: 2.59; CI: 1.31, 5.15), need for intraoperative red blood cell transfusion (OR: 2.02; CI: 1.26, 3.25), and postoperative platelet transfusion (OR: 1.78; CI: 1.05, 3.01) when compared with the historic cohort. In patients who are at high risk for postoperative renal failure, GDP was not associated with a decrease in AKI when compared to the historical cohort managed traditionally by determining CPB flows based on body surface area. Surprisingly, the GDP cohort performed significantly worse than the retrospective control group in terms of ARF, mortality, intensive care unit readmission, and RBC and platelet transfusions.

Tranexamic acid: Beyond antifibrinolysis.

Tranexamic acid (TXA) is a popular antifibrinolytic drug widely used in hemorrhagic trauma patients and cardiovascular, orthopedic, and gynecological surgical patients. TXA binds plasminogen and prevents its maturation to the fibrinolytic enzyme plasmin. A number of studies have demonstrated the broad life-saving effects of TXA in trauma, superior to those of other antifibrinolytic agents. Besides preventing fibrinolysis and blood loss, TXA has been reported to suppress posttraumatic inflammation and edema. Although the efficiency of TXA transcends simple inhibition of fibrinolysis, little is known about its mechanisms of action besides the suppression of plasmin maturation. Understanding the broader effects of TXA at the cell, organ, and organism levels are required to elucidate its potential mechanisms of action transcending antifibrinolytic activity. In this article, we provide a brief review of the current clinical use of TXA and then focus on the effects of TXA beyond antifibrinolytics such as its anti-inflammatory activity, protection of the endothelial and epithelial monolayers, stimulation of mitochondrial respiration, and suppression of melanogenesis.

Prolonged Cardiopulmonary Bypass is Associated With Endothelial Glycocalyx Degradation.

BACKGROUND: The endothelial glycocalyx (EG) is involved in critical regulatory mechanisms that maintain endothelial vascular integrity. We hypothesized that prolonged cardiopulmonary bypass (CPB) may be associated with EG degradation. We performed an analysis of soluble syndecan-1 levels in relation to duration of CPB, as well as factors associated with cell stress and damage, such as mitochondrial DNA (mtDNA) and inflammation. METHODS: Blood samples from subjects undergoing cardiac surgery with CPB (n = 54) were obtained before and during surgery, 4-8 h and 24 h after completion of CPB, and on postoperative day 4. Flow cytometry was used to determine subpopulations of white blood cells. Plasma levels of mtDNA were determined using quantitative polymerase chain reaction and plasma content of shed syndecan-1 was measured. To determine whether syndecan-1 was signaling white blood cells, the effect of recombinant syndecan-1 on mobilization of neutrophils from bone marrow was tested in mice. RESULTS: CPB is associated with increased mtDNA during surgery, increased syndecan-1 blood levels at 4-8 h, and increased white blood cell count at 4-8 h and 24 h. Correlation analysis revealed significant positive associations between time on CPB and syndecan-1 (rs = 0.488, P < 0.001) and level of syndecan-1 and neutrophil count (rs = 0.351, P = 0.038) at 4-8 h. Intravenous administration of recombinant syndecan-1 in mice resulted in a 2.5-fold increase in the number of circulating neutrophils, concurrent with decreased bone marrow neutrophil number. CONCLUSIONS: Longer duration of CPB is associated with increased plasma levels of soluble syndecan-1, a signal for EG degradation, which can induce neutrophil egress from the bone marrow. Development of therapy targeting EG shedding may be beneficial in patients with prolonged CPB.

Comparison of intraoperative tranexamic acid and epsilon-aminocaproic acid in cardiopulmonary bypass patients.

Objective: To compare tranexamic acid (TXA) and epsilon-aminocaproic acid (EACA) in patients undergoing cardiac surgery with cardiopulmonary bypass. Methods: Over a consecutive 2-year period, 824 adult cardiac surgery patients who received TXA during an EACA shortage were compared with 778 patients who received EACA postshortage. Patient characteristics and process and outcome variables were collected through chart review and database queries. This retrospective analysis used inverse probability of treatment weighting to control for confounding by indication, and propensity scores were calculated using a logistic regression model. Results: In adjusted models, overall transfusion rates for the TXA cohort (odds ratio [OR], 0.94; 95% confidence interval [95% CI], 0.81-1.10) and administration of platelets (OR, 1.04; 95% CI, 0.85-1.27), red blood cells (OR, 0.93; 95% CI, 0.80-1.09), fresh frozen plasma (OR, 1.00; 95% CI, 0.79-1.25), and cryoprecipitate (OR, 1.08; 95% CI, 0.71-1.64) were equivalent to the EACA cohort. In addition, there was no statistical difference with respect to stroke, seizure, mortality, reoperation for bleeding, chest tube drainage, and acute kidney injury. Patients who received TXA had shorter ventilator times (difference in medians -1.33 hours [95% CI, -1.86 to -0.80]) and lower postsurgical charges (difference of medians -$2913 [95% CI, -5147 to -679]). Conclusions: Substituting TXA for EACA during cardiac surgery with cardiopulmonary bypass did not change transfusion rate or amount, nor was there a significant difference in chest tube drainage. Patients who received TXA had a statistically significant but not clinically significant lower postoperative ventilator times and charges without an increase in mortality, stroke, reoperation for bleeding, acute kidney injury, or seizures.

Tranexamic acid suppresses the release of mitochondrial DNA, protects the endothelial monolayer and enhances oxidative phosphorylation.

Damage-associated molecular patterns, including mitochondrial DNA (mtDNA) are released during hemorrhage resulting in the development of endotheliopathy. Tranexamic acid (TXA), an antifibrinolytic drug used in hemorrhaging patients, enhances their survival despite the lack of a comprehensive understanding of its cellular mechanisms of action. The present study is aimed to elucidate these mechanisms, with a focus on mitochondria. We found that TXA inhibits the release of endogenous mtDNA from granulocytes and endothelial cells. Furthermore, TXA attenuates the loss of the endothelial monolayer integrity induced by exogenous mtDNA. Using the Seahorse XF technology, it was demonstrated that TXA strongly stimulates mitochondrial respiration. Studies using Mitotracker dye, cells derived from mito-QC mice, and the ActivSignal IPAD assay, indicate that TXA stimulates biogenesis of mitochondria and inhibits mitophagy. These findings open the potential for improvement of the strategies of TXA applications in trauma patients and the development of more efficient TXA derivatives.

Tranexamic acid suppresses the release of mitochondrial DAMPs and reduces lung inflammation in a murine burn model.

BACKGROUND: Severe burn injuries are known to initiate a profound systemic inflammatory response (SIRS) that may lead to burn shock and other SIRS-related complications. Damage-associated molecular patterns (DAMPs) are important early signaling molecules that initiate SIRS after burn injury. Previous work in a rodent model has shown that application of a topical immune modulator (p38MAPK inhibitor) applied directly to the burn wound decreases cytokine expression, reduces pulmonary inflammation and edema. Our group has demonstrated that tranexamic acid (TXA)-in addition to its use as an antifibrinolytic-has cell protective in vitro effects. We hypothesized that administration of TXA after burn injury would attenuate DAMP release and reduce lung inflammation. METHODS: C57/BL6 male mice underwent a 40% Total Body Surface Area (TBSA) scald burn. Sham animals underwent the same procedure in room temperature water. One treatment group received the topical application of p38MAPK inhibitor after burn injury. The other treatment group received an intraperitoneal administration of TXA after burn injury. Animals were sacrificed at 5 hours. Plasma was collected by cardiac puncture. MtDNA levels in plasma were determined by quantitative Polymerase Chain Reaction (qPCR). Syndecan-1 levels in plasma were measured by ELISA. Lungs were harvested, fixed, and paraffin-embedded. Sections of lungs were stained for antigen to detect macrophages. RESULTS: Topical p38MAPK inhibitor and TXA significantly attenuated mtDNA release. Both TXA and the topical p38MAPK inhibitor reduced lung inflammation as represented by decreased macrophage infiltration. Syndecan-1 levels showed no difference between burn and treatment groups. CONCLUSION: Both p38 MAPK inhibitor and TXA demonstrated the ability to attenuate burn-induced DAMP release and lung inflammation. Beyond its role as an antifibrinolytic, TXA may have significant anti-inflammatory effects pertinent to burn resuscitation. Further study is required; however, TXA may be a useful adjunct in burn resuscitation.

AMPK is critical for mitochondrial function during reperfusion after myocardial ischemia.

AMP-activated kinase (AMPK) is a stress responsive kinase that regulates cellular metabolism and protects against cardiomyocyte injury during ischemia-reperfusion (IR). Mitochondria play an important role in cell survival, but the specific actions of activated AMPK in maintaining mitochondrial integrity and function during reperfusion are unknown. Thus, we assessed the consequences of AMPK inactivation on heart mitochondrial function during reperfusion. Mouse hearts expressing wild type (WT) or kinase-dead (KD) AMPK were studied. Mitochondria isolated from KD hearts during reperfusion had intact membrane integrity, but demonstrated reduced oxidative capacity, increased hydrogen peroxide production and decreased resistance to mitochondrial permeability transition pore opening compared to WT. KD hearts showed increased activation of the mitogen activated protein kinase kinase 4 (MKK4) and downstream c-Jun terminal kinase (JNK) and greater necrosis during reperfusion after coronary occlusion. Transgenic expression of mitochondrial catalase (MCAT) prevented the excessive cardiac JNK activation and attenuated the increased myocardial necrosis observed during reperfusion in KD mice. Inhibition of JNK increased the resistance of KD hearts to mPTP opening, contractile dysfunction and necrosis during IR. Thus, intrinsic activation of AMPK is critical to prevent excess mitochondrial reactive oxygen production and consequent JNK signaling during reperfusion, thereby protecting against mPTP opening, irreversible mitochondrial damage and myocardial injury.

LKB1 deletion causes early changes in atrial channel expression and electrophysiology prior to atrial fibrillation.

AIMS: Liver kinase B1 (LKB1) is a protein kinase that activates the metabolic regulator AMP-activated protein kinase (AMPK) and other related kinases. Deletion of LKB1 in mice leads to cardiomyopathy and atrial fibrillation (AF). However, the specific role of the LKB1 pathway in early atrial biology remains unknown. Thus, we investigated whether LKB1 deletion altered atrial channel expression and electrophysiological function in a cardiomyocyte-specific knockout mouse model. METHODS AND RESULTS: We performed a systematic comparison of αMHC-Cre LKB1(fl/fl) and littermate LKB1(fl/fl) male mice. This included analysis of gene expression, histology, and echocardiography, as well as cellular and tissue-level electrophysiology using patch-clamp recordings in vitro, optical mapping ex vivo, and ECG recordings in vivo. At postnatal day 1, atrial depolarization was prolonged, and Nav1.5 and Cx40 expression were markedly down-regulated in MHC-Cre LKB1(fl/fl) mice. Inward sodium current density was significantly decreased in MHC-Cre LKB1(fl/fl) neonatal atrial myocytes. Subsequently, additional alterations in atrial channel expression, atrial fibrosis, and spontaneous onset of AF developed by 2 weeks of age. In adult mice, abnormalities of interatrial conduction and bi-atrial electrical coupling were observed, likely promoting the perpetuation of AF. Mice with AMPK-inactivated hearts demonstrated modest overlap in channel expression with MHC-Cre LKB1(fl/fl) hearts, but retained normal structure, electrophysiological function and contractility. CONCLUSIONS: Deletion of LKB1 causes early defects in atrial channel expression, action potential generation and conduction, which precede widespread atrial remodelling, fibrosis and AF. LKB1 is critical for normal atrial growth and electrophysiological function.

Role of uncoupling protein 3 in ischemia-reperfusion injury, arrhythmias, and preconditioning.

Overexpression of mitochondrial uncoupling proteins (UCPs) attenuates ischemia-reperfusion (I/R) injury in cultured cardiomyocytes. However, it is not known whether UCPs play an essential role in cardioprotection in the intact heart. This study evaluated the cardioprotective efficacy of UCPs against I/R injury and characterized the mechanism of UCP-mediated protection in addition to the role of UCPs in ischemic preconditioning (IPC). Cardiac UCP3 knockout (UCP3(-/-)) and wild-type (WT) mice hearts were subjected to ex vivo and in vivo models of I/R injury and IPC. Isolated UCP3(-/-) mouse hearts were retrogradely perfused and found to have poorer recovery of left ventricular function compared with WT hearts under I/R conditions. In vivo occlusion of the left coronary artery resulted in twofold larger infarcts in UCP3(-/-) mice compared with WT mice. Moreover, the incidence of in vivo I/R arrhythmias was higher in UCP3(-/-) mice. Myocardial energetics were significantly impaired with I/R, as reflected by a decreased ATP content and an increase in the AMP-to-ATP ratio. UCP3(-/-) hearts generated more reactive oxygen species (ROS) than WT hearts during I/R. Pretreatment of UCP3(-/-) hearts with the pharmacological uncoupling agent carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone improved postischemic functional recovery. Also the protective efficacy of IPC was abolished in UCP3(-/-) mice. We conclude that UCP3 plays a critical role in cardioprotection against I/R injury and the IPC phenomenon. There is increased myocardial vulnerability to I/R injury in hearts lacking UCP3. The mechanisms of UCP3-mediated cardioprotection include regulation of myocardial energetics and ROS generation by UCP3 during I/R.

Endothelium-derived neuregulin protects the heart against ischemic injury.

BACKGROUND: Removal of cardiac endothelial cells (EC) has been shown to produce significant detrimental effects on the function of adjacent cardiac myocytes, suggesting that EC play a critical role in autocrine/paracrine regulation of the heart. Despite this important observation, the mediators of the protective function of EC remain obscure. Neuregulin (NRG, a member of the epidermal growth factor family) is produced by EC and cardiac myocytes contain receptors (erbB) for this ligand. We hypothesized that NRG is an essential factor produced by EC, which promotes cardioprotection against ischemic injury. METHODS AND RESULTS: We demonstrate that human cardiac EC express and release NRG in response to hypoxia-reoxygenation. Under conditions where hypoxia--reoxygenation causes significant cardiac myocyte cell death, NRG can significantly decrease apoptosis of isolated adult ventricular myocytes. Coculturing adult murine myocytes with human umbilical vein, murine lung microvascular, or human coronary artery EC can also protect myocytes against hypoxia--reoxygenation--induced apoptosis. These protective effects are abolished by NRG gene deletion or silencing of NRG expression in EC. Finally, endothelium-selective deletion of NRG in vivo leads to significantly decreased tolerance to ischemic insult, as demonstrated by impaired postischemic contractile recovery in a perfused whole-organ preparation and larger infarct sizes after coronary artery ligation. CONCLUSION: Together, these data demonstrate that EC-derived NRG plays an important role in cardiac myocyte protection against ischemic injury in the heart and supports the idea that manipulation of this signaling pathway may be an important clinical target in this setting.

Activation of the aryl hydrocarbon receptor by doxorubicin mediates cytoprotective effects in the heart.

AIMS: Doxorubicin (DOX) is a highly effective chemotherapeutic agent; however, cumulative dose-dependent cardiotoxicity is a significant side effect of this therapy. Because DOX is a polyaromatic hydrocarbon, we hypothesized that it will be metabolized by the activation of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that is involved in the metabolism of numerous xenobiotic agents. These studies were performed to determine whether DOX activates AhR and whether this activation modulates the toxicity of DOX in cardiomyocytes. METHODS AND RESULTS: Treatment with DOX induced AhR migration to the nucleus, increased AhR binding with its co-factor, aryl hydrocarbon receptor nuclear translocator-1 (ARNT1), and increased the expression of AhR-regulated phase I (CYP1A1) and phase II (GSTA1) drug-metabolizing enzymes in both cardiomyocytes and in the intact heart. Knockdown of AhR in H9C2 cells abolished DOX-induced increases in CYP1A1 and GSTA1 expression. Similar results were obtained by treating adult rat ventricular myocytes with the AhR antagonist, CH-223191. Taken together, these findings indicate that DOX-induced upregulation of CYP1A1 and GSTA1 expression is AhR dependent. AhR null mice treated with 10 mg/kg DOX did not show any activation of CYP1A1 or GSTA1 expression. Moreover, lack of AhR in vivo resulted in a significant decrease in left ventricular function compared with wild-type animals, and increased p53 activation and apoptosis in the heart after treatment with DOX. CONCLUSIONS: These findings indicate that AhR plays an important role in DOX metabolism by the heart and further demonstrate that AhR is cardioprotective against DOX-induced cardiotoxicity.

Uncoupling protein downregulation in doxorubicin-induced heart failure improves mitochondrial coupling but increases reactive oxygen species generation.

PURPOSE: Doxorubicin-based chemotherapy is limited by the development of dose-dependent left ventricular dysfunction and congestive heart failure caused by reactive oxygen species (ROS). Uncoupling proteins (UCP) can inhibit mitochondrial ROS production as well as decrease myocyte damage from exogenous ROS. Prior studies have shown that cardiac UCP2 and UCP3 mRNA expression is decreased with acute doxorubicin treatment. However, the expression of UCP protein in hearts with doxorubicin cardiotoxicity and the resultant changes in mitochondrial function and oxidant stress have not been determined. METHODS: Heart failure was induced in Sprague-Dawley rats with intraperitoneal injections of doxorubicin (2 mg/kg t.i.w., total dose: 18 mg/kg). Mitochondria were isolated from mice receiving doxorubicin or saline injections for determination of UCP2 and UCP3 expression. In addition, mitochondrial respiration, ATP synthesis and ROS production were determined. RESULTS: Doxorubicin-induced heart failure was associated with significant decreases in UCP2 and UCP3 protein expression compared with nonfailing hearts (P < 0.05). While the rates of state 3 and state 4 respiration and ATP synthesis were lower in mitochondria isolated from failing hearts, the respiratory control ratio was 15% higher (P < 0.05), and the ratio of ATP production to oxygen consumption was 25% higher (P < 0.05) in mitochondria from failing hearts, indicating greater coupling between citric acid cycle flux and mitochondrial ATP synthesis. However, the decrease in UCP expression was associated with 50% greater mitochondrial ROS generation (P < 0.05). CONCLUSIONS: Downregulation of myocardial UCP2 and UCP3 in the setting of doxorubicin-induced heart failure is associated with improved efficiency of ATP synthesis, which might compensate for abnormal energy metabolism. However, this beneficial effect is counterbalanced by greater oxidant stress.

AMP-activated protein kinase mediates ischemic glucose uptake and prevents postischemic cardiac dysfunction, apoptosis, and injury.

AMP-activated protein kinase (AMPK) is an important regulator of diverse cellular pathways in the setting of energetic stress. Whether AMPK plays a critical role in the metabolic and functional responses to myocardial ischemia and reperfusion remains uncertain. We examined the cardiac consequences of long-term inhibition of AMPK activity in transgenic mice expressing a kinase dead (KD) form of the enzyme. The KD mice had normal fractional shortening and no heart failure, cardiac hypertrophy, or fibrosis, although the in vivo left ventricular (LV) dP/dt was lower than that in WT hearts. During low-flow ischemia and postischemic reperfusion in vitro, KD hearts failed to augment glucose uptake and glycolysis, although glucose transporter content and insulin-stimulated glucose uptake were normal. KD hearts also failed to increase fatty acid oxidation during reperfusion. Furthermore, KD hearts demonstrated significantly impaired recovery of LV contractile function during postischemic reperfusion that was associated with a lower ATP content and increased injury compared with WT hearts. Caspase-3 activity and TUNEL-staining were increased in KD hearts after ischemia and reperfusion. Thus, AMPK is responsible for activation of glucose uptake and glycolysis during low-flow ischemia and plays an important protective role in limiting damage and apoptotic activity associated with ischemia and reperfusion in the heart.