Preparation of Homogenate

All steps of the preparation were performed on ice. Fresh left ventricular tissue was rapidly but gently dissected and immediately transferred into 5 mL of ice‐cold buffer solution (K medium), stored on ice. Then tissue samples were transported to mitochondrial laboratory in the same building within 5 minutes and processed immediately. According to modified protocol for human skeletal muscle homogenates 13 connective and adipose tissue were removed using pre‐cold scissors and anatomic forceps to ensure only heart muscle tissue was homogenized. After gentle blotting by a piece of sterile gauze, heart muscle tissue was weighted using analytical scale (1‐μg readability) and cut into fine fragments. Then tissue fragments were scraped into a pre‐chilled Dounce Tissue Grinder (glass construction; Wheaton^, Millville, USA), 1 mL of K medium was added for each 100 mg of muscle to obtain 10% homogenate. Minced cardiac tissues were then manually homogenized by 8 to 10 initial strokes with a glass pestle. Raw homogenate was then filtered through cheesecloth. The whole procedure took 5 to 10 minutes. Notably, the same operator prepared all the samples for measurement in all animals to minimize variability.