Ethyl pyruvate preserves cardiac function and attenuates oxidative injury after prolonged myocardial ischemia

OBJECTIVE: Myocardial injury and dysfunction following ischemia are mediated in part by reactive oxygen species. Pyruvate, a key glycolytic intermediary, is an effective free radical scavenger but unfortunately is limited by aqueous instability. The ester derivative, ethyl pyruvate, is stable in solution and should function as an antioxidant and energy precursor. This study sought to evaluate ethyl pyruvate as a myocardial protective agent in a rat model of ischemia-reperfusion injury. METHODS: Rats underwent 30-minute ischemia and 30-minute reperfusion of the left anterior descending coronary artery territory. Immediately prior to both ischemia and reperfusion, animals received an intravenous bolus of either ethyl pyruvate (n = 26) or vehicle control (n = 26). Myocardial high-energy phosphate levels were determined by adenosine triphosphate assay, oxidative injury was measured by lipid peroxidation assay, infarct size was quantified by triphenyltetrazolium chloride staining, and cardiac function was assessed in vivo. RESULTS: Ethyl pyruvate administration significantly increased myocardial adenosine triphosphate levels compared with control (87.6 +/- 29.2 nmol/g vs 10.0 +/- 2.4 nmol/g, P =.03). In ischemic myocardium, ethyl pyruvate reduced oxidative injury compared with control (63.8 +/- 3.3 nmol/g vs 89.5 +/- 3.0 nmol/g, P <.001). Ethyl pyruvate diminished infarct size as a percentage of area at risk (25.3% +/- 1.5% vs 33.6% +/- 2.1%, P =.005). Ethyl pyruvate improved myocardial function compared with control (maximum pressure: 86.6 +/- 2.9 mm Hg vs 73.5 +/- 2.5 mm Hg, P <.001; maximum rate of pressure rise: 3518 +/- 243 mm Hg/s vs 2703 +/- 175 mm Hg/s, P =.005; maximal rate of ventricular systolic volume ejection: 3097 +/- 479 microL/s vs 2120 +/- 287 microL/s, P =.04; ejection fraction: 41.9% +/- 3.8% vs 31.4% +/- 4.1%, P =.03; cardiac output: 26.7 +/- 0.9 mL/min vs 22.7 +/- 1.3 mL/min, P =.01; and end-systolic pressure-volume relationship slope: 1.09 +/- 0.22 vs 0.59 +/- 0.2, P =.02). CONCLUSIONS: In this study of myocardial ischemia-reperfusion injury, ethyl pyruvate enhanced myocardial adenosine triphosphate levels, attenuated myocardial oxidative injury, decreased infarct size, and preserved cardiac function.     

Reduction of myocardial reperfusion injury by aprotinin after regional ischemia and cardioplegic arrest

BACKGROUND: Surgical coronary revascularization with cardiopulmonary bypass and cardioplegia has been associated with reperfusion injury. The serine protease inhibitor aprotinin has been suggested to reduce reperfusion injury, yet a clinically relevant study examining regional ischemia under conditions of cardiopulmonary bypass and cardioplegia has not been performed. METHODS: Pigs were subjected to 30 minutes of regional myocardial ischemia by distal left anterior descending coronary artery occlusion, followed by 60 minutes of cardiopulmonary bypass with 45 minutes of cardioplegic arrest and 90 minutes of post-cardiopulmonary bypass reperfusion. The treatment group (n = 6) was administered aprotinin systemically (40,000 kallikrein-inhibiting units [KIU]/kg intravenous loading dose, 40,000 KIU/kg pump prime, and 10,000 KIU x kg(-1) x h(-1) intravenous continuous infusion). Control animals (n = 6) received crystalloid solution. Global and regional myocardial functions were analyzed by the left ventricular+dP/dt and the percentage segment shortening, respectively. Left ventricular infarct size was measured by tetrazolium staining. Tissue myeloperoxidase activity was measured. Myocardial sections were immunohistochemically stained for nitrotyrosine. Coronary microvessel function was studied by videomicroscopy. RESULTS: Myocardial infarct size was decreased with aprotinin treatment (27.0% +/- 3.5% vs 45.3% +/- 3.0%, aprotinin vs control; P <.05). Myocardium from the ischemic territory showed diminished nitrotyrosine staining in aprotinin-treated animals versus controls, and this was significant by grade (1.3 +/- 0.2 vs 3.2 +/- 0.2, aprotinin vs control; P <.01). In the aprotinin group, coronary microvessel relaxation improved most in response to the endothelium-dependent agonist adenosine diphosphate (44.7% +/- 3.2% vs 19.7% +/- 1.7%, aprotinin vs control; P <.01). No significant improvements in myocardial function were observed with aprotinin treatment. CONCLUSIONS: Aprotinin reduces reperfusion injury after regional ischemia and cardioplegic arrest. Protease inhibition may represent a molecular strategy to prevent postoperative myocardial injury after surgical revascularization with cardiopulmonary bypass.     

Warm blood cardioplegic arrest induces mitochondrial-mediated cardiomyocyte apoptosis associated with increased urocortin expression in viable cells

OBJECTIVES: This study assesses the mechanisms of apoptosis in patients after on-pump coronary artery bypass graft surgery and the potential involvement of the endogenous cardiac peptide urocortin as a cardiomyocyte salvage mechanism. We have previously described the mechanisms of apoptosis after ischemia-reperfusion injury in the rat heart and shown that endogenous urocortin is cardioprotective. Here we extend these findings to the human heart exposed to ischemic-reperfusion injury. METHODS: Two sequential biopsy specimens were obtained from the right atriums of 24 patients undergoing coronary artery bypass grafting at the start of grafting and 10 minutes after release of the aortic clamp. Apoptosis was identified by means of immunocytochemical colocalization between terminal deoxynucleotidyl transferase-mediated nick end-labeling positivity and active caspase-3. Immunostaining for active caspase-9 and caspase-8 was performed to identify the pathways of apoptosis induction. Urocortin and adenosine triphosphate-dependent potassium channel expression was also assessed by means of immunocytochemistry. RESULTS: Myocyte apoptosis (<0.1% before coronary artery bypass grafting) was increased after coronary artery bypass grafting and reperfusion and was greater in patients with longer periods of cardioplegic arrest (3.3% +/- 0.5% with <55 minutes and 5.1% +/- 0.9% with 85-100 minutes, P <.001). Processing of caspase-9 was always more pronounced than that of caspase-8 (P <.05). Cardioplegic arrest was also associated with increased urocortin expression (up to 29% +/- 3.5% vs <3% in samples obtained before coronary artery bypass grafting, P <.001) but only in nonapoptotic myocytes. These and surrounding viable myocytes also showed increased Kir6.1 adenosine triphosphate-dependent potassium channel expression. CONCLUSIONS: Cardioplegic arrest and subsequent reperfusion result in cardiomyocyte apoptosis, largely through mitochondrial injury, as well as exclusive urocortin expression in viable cells. This finding might suggest a cardioprotective role for endogenous urocortin in human subjects.     

Sialyl LewisX oligosaccharide preserves myocardial and endothelial function during cardioplegic ischemia

BACKGROUND: Neutrophil adhesion to endothelium contributes to myocardial reperfusion injury after cardiac operation. Initial neutrophil-endothelial interactions involve selectins, which bind Sialyl-LewisX on neutrophils. Blockade of selectin-mediated neutrophil-endothelial interactions with CY-1503, a synthetic analogue of Sialyl-LewisX, might reduce reperfusion injury after myocardial ischemia. METHODS: The efficacy of CY-1503 to attenuate global myocardial reperfusion injury was assessed in isolated blood-perfused neonatal lamb hearts that had 2 hours of cold cardioplegic ischemia. CY-1503 (40 mg/L) or saline vehicle was added to blood perfusate before ischemia. Contractile function (developed pressure, dP/dt) and coronary vascular endothelial function (acetylcholine response) were assessed at base line and during reperfusion. Myocardial neutrophil accumulation was assessed by myeloperoxidase quantification. RESULTS: Compared to controls, treatment with CY-1503 improved recovery of all indices of contractile function, preserved coronary vascular endothelial function, and reduced myocardial neutrophil accumulation. CONCLUSIONS: In isolated neonatal lamb hearts that underwent hypothermic cardioplegic ischemia, CY-1503 administration reduced myocardial neutrophil accumulation and preserved endothelial and contractile function. Selectin blockade of leukocyte-endothelial interactions might attenuate reperfusion injury and enhance myocardial protection during cardiac surgical procedures.     

Improving myocardial metabolic and functional recovery after cardioplegic arrest

The myocardial oxidation of fatty acids and glucose, the predominant substrates for aerobic metabolism, is impaired after cardioplegic arrest for coronary revascularization. Because lactate can be readily metabolized to pyruvate, it may be the preferred substrate for aerobic metabolism after cardioplegic arrest when arterial concentrations are elevated. Nineteen patients undergoing elective coronary revascularization with blood cardioplegia were randomized to receive LOW (nine patients, no exogenous lactate) or HIGH (10 patients, a perioperative infusion of Ringer's lactate) arterial lactate concentrations. Coronary sinus catheterization and lactate labeled with carbon 14 permitted calculation of myocardial oxygen consumption and lactate oxidation which were significantly increased during reperfusion in the group with HIGH arterial lactate concentrations. Atrial pacing at 110 beats/min on cardiopulmonary bypass resulted in myocardial lactate production (suggesting ischemic anaerobic metabolism) in the LOW lactate group, but atrial pacing increased lactate consumption and oxidation in the HIGH lactate group (suggesting increased aerobic metabolism). Systolic function (the relation between end-systolic pressure and volume) as assessed by nuclear ventriculography 3 hours postoperatively was significantly better (p less than 0.05 by analysis of covariance) in the HIGH lactate group. Postoperative myocardial creatine kinase release was significantly lower in the HIGH lactate group, which suggested less perioperative ischemic injury. Lactate was the preferred substrate for myocardial oxidative metabolism after cardioplegic arrest, and the higher arterial lactate concentrations improved myocardial metabolic and functional recovery and reduced perioperative ischemic injury.     

N-acetylcysteine prevents reactive oxygen species-mediated myocardial stress in patients undergoing cardiac surgery: results of a randomized, double-blind, placebo-controlled clinical trial

OBJECTIVE: Reactive oxygen species have been shown to contribute to myocardial stress in patients undergoing cardiac surgery, as demonstrated by myocardial 8-iso-prostaglandin-F(2)alpha and nitrotyrosine formation. We hypothesized that the reactive oxygen species scavenger N-acetylcysteine attenuates reactive oxygen species-mediated myocardial stress in patients undergoing cardiac surgery. METHODS: Forty patients undergoing coronary artery surgery (mean age +/- SD, 66 +/- 9 years; 9 women and 31 men) were randomized to receive either N-acetylcysteine (100 mg/kg into cardiopulmonary bypass prime followed by infusion at 20 mg.kg(-1).h(-1), n = 20) or placebo (n = 20). Patients and clinical staff were blinded to group assignment. Transmural left ventricular biopsy specimens collected before and at the end of cardiopulmonary bypass were subjected to immunocytochemical staining against 8-iso-prostaglandin-F(2)alpha (primary measure) as an indicator for reactive oxygen species-mediated lipid peroxidation and nitrotyrosine (coprimary measure) as a marker for peroxynitrite-mediated tissue injury. Cardiomyocyte staining was quantitatively determined by using densitometry (in gray units). Global left ventricular function was measured on the basis of fractional area of contraction by using transesophageal echocardiography. RESULTS: Patient characteristics in both groups were comparable. The change in left ventricular cardiomyocyte staining (end of cardiopulmonary bypass--before cardiopulmonary bypass) differed significantly between groups for both primary measures: 8-iso-prostaglandin-F(2)alpha, -1.8 +/- 7.5 gray units (mean +/- SD, N-acetylcysteine group) versus 5.0 +/- 4.1 gray units (placebo group; 95% confidence interval, 2.6-11.0, P =.003); nitrotyrosine, -6.4 +/- 10.0 gray units (N-acetylcysteine group) versus 9.2 +/- 8.4 gray units (placebo group; 95% confidence interval, 9.4-21.7, P <.001). Hemodynamics and clinical outcomes were comparable in both groups. CONCLUSIONS: Reactive oxygen species scavenging with N-acetylcysteine attenuates myocardial oxidative stress in the hearts of patients subjected to cardiopulmonary bypass and cardioplegic arrest.     

Pinacidil improves contractile function and intracellular calcium handling in isolated cardiac myocytes exposed to simulated cardioplegic arrest

Background

We examined the effects of pinacidil on contractile function and intracellular calcium in isolated rat cardiomyocytes exposed to cardioplegic solution.

Methods

Rat myocytes were incubated at 24°C for 2 hours in cardioplegic solution with or without pinacidil (50 μmol/L), then they were perfused with Krebs-Henseleit solution with a gas phase of 95% O₂/5% CO₂ at the same temperature. Contraction and intracellular calcium transients were then measured by video tracking and spectrofluorometry.

Results

During 20 minutes of perfusion after 2 hours in cardioplegic solution with pinacidil, (1) the recovery of contractile function was significantly increased in terms of both amplitude of contraction (98.30% ± 9.90% versus 81.00% ± 11.25%; p < 0.05) and peak velocity of cell shortening (100.90% ± 13.79% versus 76.89% ± 18.14%; p < 0.01) when compared with myocytes in cardioplegic solution without pinacidil; (2) the amplitudes of the intracellular calcium transients evoked by electrical stimulation and caffeine (10 mmol/L) increased by 23.31% to approximately 40.72% and 61.73%, respectively, compared with those in cardioplegic solution without pinacidil; and (3) the decay time of the caffeine-induced intracellular calcium transient decreased by 36.64% ± 15.10% relative to that measured in cardioplegic solution without pinacidil. The effects induced by supplementing the cardioplegic solution with pinacidil were diminished in the presence of glibenclamide (10 μmol/L).

Conclusions

Addition of the adenosine triphosphate-sensitive potassium-channel opener, pinacidil, to a high potassium cardioplegic solution improves recovery of contractile properties and cytosolic calcium in isolated rat cardiac myocytes.     

Impact of sodium-hydrogen exchange inhibition by cariporide on death or myocardial infarction in high-risk CABG surgery patients: results of the CABG surgery cohort of the GUARDIAN study

OBJECTIVES: To evaluate the effects of cariporide on all-cause mortality or myocardial infarction at 36 days in patients at risk of myocardial necrosis after coronary artery bypass graft surgery. METHODS: In the coronary artery bypass graft cohort of the GUARD During Ischemia Against Necrosis trial, patients > or =18 years who required urgent coronary artery bypass graft, repeat coronary artery bypass graft, or had a history of unstable angina and > or =2 risk factors (age >65 years, female gender, diabetes mellitus, ejection fraction <35%, or left main or 3-vessel disease) were randomized to placebo (n = 743) or cariporide 20 mg (n = 736), 80 mg (n = 705), or 120 mg (n = 734). A 1-hour intravenous infusion was initiated shortly before surgery and administered every 8 hours for 2 to 7 days. Patients were followed up for 6 months. A nonparametric covariance analysis was used to calculate the primary efficacy endpoint. RESULTS: Baseline characteristics were similar between treatment groups. The cariporide 20- and 80-mg groups had event rates similar to placebo. The endpoint of all-cause mortality or myocardial infarction at day 36 was significant with cariporide 120 mg versus placebo (event rate 12.2% vs 16.2%; P =.027). The risk reduction was evident on postoperative day 1 (3.3% vs 6.5%; P =.005) and was maintained at 6 months (event rate 15.0% vs 18.6%; P =.033). Cariporide was well tolerated, and most adverse events were mild and transient in this high-risk population. CONCLUSIONS: Clinical benefit with cariporide 120 mg was observed early after treatment initiation and continued for 6 months postsurgery, suggesting that sodium-hydrogen exchange inhibition with cariporide is cardioprotective in patients undergoing high-risk coronary artery bypass graft surgery.