755 ATRIAL NATRIURETIC PEPTIDE STIMULATES AUTOPHAGY/MITOPHAGY AND IMPROVES MITOCHONDRIAL DISFUNCTION IN CHRONIC HEART FAILURE


 
 
 Mitochondrial dysfunction, causing increased reactive oxygen species production, is a molecular feature of heart failure (HF) and it has been reported in circulating leucocytes of patients with HF with reduced ejection fraction (HFrEF). Atrial natriuretic peptide (ANP) exerts many cardiac beneficial effects, including the ability to protect cardiomyocytes by promoting autophagy.
 
 
 
 To test the impact of ANP on autophagy/mitophagy responses, altered mitochondrial structure and function and increased oxidative stress in chronic HFrEF patients.
 Experimental design. The present study used both ex-vivo and in-vivo approaches. We examined sixteen consecutive chronic HFrEF patients referring to the outpatient clinic of the Cardiology unit of Sant’Andrea Hospital in Rome. Out of them, for the ex-vivo study we enrolled 10 patients matching the following inclusion criteria: age under 75 years and left ventricle ejection fraction <40%. For the in-vivo study we recruited six patients and evaluated them before and after 2-month treatment with the Angiotensin Receptor Neprilysin inhibitor (ARNi) sacubitril/valsartan started at the dosage of 49/51 mg twice daily, rapidly uptitrated at the dosage of 97/103 mg twice daily, while maintaining unchanged the remaining therapy. Patients with recent hospitalizations for acute HF or other acute conditions within the last 3 months before the enrollment, with malignancy, inflammatory or infectious diseases, diabetes mellitus, history of cigarette smoking and alcohol abuse were excluded.
 
 
 
 The ex-vivo direct exposure to αANP caused an improvement of mitochondrial membrane potential with a restoration of the mitochondrial protonmotive force (p<0.05) and promoted the recovery of the IMM extension with a significant improvement in the IMM/OMM index (p<0.05). The production of ROS was significantly decreased after the exposure to αANP (p<0.05). Analyzing the mitochondrial stress, we observed that the treatment with αANP induced a down-modulation of p66shc mRNA (p<0.01). The αANP treatment induced a rapid and significant autophagic response. The cytofluorimetric analysis showed a significant increase of LysoTracker levels highlighting a rise of intracellular acidic compartments (p<0.05), with the up modulation of the LC3 and Beclin mRNA levels (p<0.01 and p<0.05 respectively). The significant reduction in mitochondrial mass observed in treated PBMCs evoked an increase of the mitophagic process related to αANP (p<0.01). In vivo, the efficacy of the treatment with sacubitril/valsartan was proved by the increase in αANP levels (p<0.01) whereas levels of NT-proBNP decreased (p<0.05). PBMCs collected after treatment were characterized by lower mitochondrial oxidative stress levels and were provided of structurally more intact mitochondria characterized by better functional performances (p<0.05). PBMCs of the treated patients showed an increased level of acid compartments associated with an up-modulation of LC3 and Beclin mRNA levels and a reduction of mitochondrial mass (p<0.05). Also in this experimental context, the activation of autophagy was confirmed by ultrastructural morphometric evaluation (p<0.05).
 
 
 
 αANP stimulates both autophagy and mitophagy responses, counteracts mitochondrial dysfunction and damage ultimately reducing mitochondrial oxidative stress generation in PBMCs from chronic HF patients. These properties were confirmed upon sacubitril/valsartan administration, a therapeutic approach indicated for HFrEF treatment.
