Supplemental Material Sommer et al Cox4i2

Hemodynamic measurements in isolated, perfused, and ventilated mouse lungs Lungs of either WT or Cox4i2 mice were isolated, perfused and ventilated as described previously. Briefly, mice were deeply anesthetized intraperitoneally with xylazine (20 mg/kg body weight, Bayer Healthcare, Leverkusen, Germany) and ketamine (100 mg/kg body weight, Pfizer, Karlsruhe, Germany), and anticoagulated with heparin (50,000 I.E./kg body weight). After reaching deep anesthesia mice were intubated via a tracheostoma and ventilated with room air with a tidal volume of 300 μl, at a frequency of 90 breaths/min and a positive end-expiratory pressure of 3 cm H2O using a piston pump (Minivent Type 845, Hugo Sachs Elektronik, March-Hugstetten, Germany). Subsequently, midsternal thoracotomy was performed and catheters were inserted into the pulmonary artery and left atrium. Perfusion with sterile Krebs–Henseleit buffer (Serag-Wiessner, Naila, Germany) was initiated through the catheter in the pulmonary artery using a peristaltic pump (ISM834A V2.10, Ismatec, Glattbrugg, Switzerland), with a buffer temperature of 4°C and a flow of 0.2 ml/min. In parallel with the onset of artificial perfusion, ventilation was changed from room air to a pre-mixed gas (21% O2 5.3% CO2, balanced with N2). The lungs were removed from the thorax while continuously ventilated and perfused and were freely suspended from a force transducer for the monitoring of organ weight in a temperatureequilibrated, humidified chamber. After rinsing the lungs, the perfusion circuit was closed for recirculation, left atrial pressure was set at 2.0 mm Hg, the flow was slowly increased to 2 ml/min and the entire system heated to 37°C. Pressures in the pulmonary artery and the left atrium were registered via small diameter catheters with pressure transducers.