Remote ischemic preconditioning preserves mitochondrial function and activates pro-survival protein kinase Akt in the left ventricle during cardiac surgery: A randomized trial

Highlights

• We investigated effects of remote ischemic preconditioning (RIPC) in the ventricle of patients undergoing CABG surgery.

• RIPC preserved left ventricular mitochondrial function through surgery.

• RIPC induced phosphorylation of prosurvival kinase Akt before cardiac ischemia.

• Phosphorylation of Akt substrates increased peroperatively within RIPC only.

Abstract

Background

Understanding the intracellular mechanisms induced by remote ischemic preconditioning (RIPC) in the human left ventricle opens new possibilities for development of pharmacological cardioprotection against ischemia and reperfusion injury. In this study we investigated the effects of RIPC on mitochondrial function, activation of pro-survival protein kinase Akt and microRNA expression in left ventricular biopsies from patients undergoing coronary artery bypass surgery (CABG).

Methods

Sixty patients were randomized to control (n = 30) or RIPC (n = 30). A blood pressure cuff was applied to the arm of all patients preoperatively. The cuff remained deflated in control group, whereas RIPC was performed by 3 cycles of cuff inflation to 200 mm Hg for 5 min, separated by 5 min deflation intervals. Left ventricular biopsies were obtained before and 15 min after aortic declamping. The primary outcome was mitochondrial respiration measured in situ. Secondary outcomes were activation of protein kinase Akt, assessed by western immunoblotting, and expression of microRNAs assessed by array and real-time polymerase chain reaction.

Results

Mitochondrial respiration was preserved during surgery in patients receiving RIPC (+ 0.2 μmol O₂/min/g, p = 0.69), and reduced by 15% in controls (− 1.5 μmol O₂/min/g, p = 0.02). Furthermore, RIPC activated protein kinase Akt before aortic clamping (difference from control + 43.3%, p = 0.04), followed by increased phosphorylation of Akt substrates at reperfusion (+ 26.8%, p < 0.01). No differences were observed in microRNA expression.

Conclusions

RIPC preserves mitochondrial function and activates pro-survival protein kinase Akt in left ventricle of patients undergoing CABG. Modulation of mitochondrial function and Akt activation should be further explored as cardioprotective drug targets.

Clinical Trial Registration: http://www.clinicaltrials.gov, unique identifier: NCT01308138.

Introduction

Remote ischemic preconditioning (RIPC) has emerged as a promising strategy to reduce myocardial reperfusion injury after cardiac surgery [1]. RIPC involves exposing a tissue to brief, non-harmful periods of ischemia to induce protection against subsequent ischemic challenge in another organ. The cardioprotective effects of RIPC have been extensively investigated in animal models, revealing potential molecular targets for pharmacological cardioprotection. Previous studies have shown that regulation of mitochondrial function and activation of pro-survival protein kinase Akt are involved in the cardioprotection induced by RIPC [2], [3], and that these cellular mechanisms interact [4].

Conserved mitochondrial function is required for optimal cardiac function, since it directly influences physiological processes that are essential for cardiomyocyte survival and proper contractile activity, including maintenance of energy substrates (ATP), pH control and scavenging of reactive oxygen species [5]. Previous studies have shown that both local ischemic preconditioning and RIPC prevent impairment of mitochondrial respiration induced by ischemia in rat skeletal muscle [6], and that maintaining an optimal mitochondrial function plays an important role in protecting the heart against ischemia [3], [7]. Moreover, mitochondrial damage has been unequivocally demonstrated as a trigger of apoptotic cardiomyocyte death [8], [9]. Preclinical studies showed that local ischemic preconditioning and RIPC reduce ischemic cardiac damage by blocking apoptosis through activation of pro-survival protein kinase Akt [10]. Accordingly, inhibition of Akt signaling completely blocks the effects of RIPC in a porcine model [11], while targeted activation of Akt renders potent cardioprotection in vivo [12].

Experimental studies also reported a causal involvement of microRNAs (e.g. microRNAs 199a and 320) [13], [14] in ischemia–reperfusion injury and mitochondrial physiology [15]. However, the effects of RIPC on left ventricular mitochondrial function and microRNA expression have never been explored in humans.

Despite robust preclinical evidence, intracellular mechanisms induced by RIPC in the human left ventricle are nearly unexplored. Therefore, we investigated the effects of RIPC on left ventricular mitochondrial function, microRNA expression and activation of protein kinase Akt in patients undergoing coronary artery bypass graft (CABG) surgery.