Effect of doxycycline treatment on germ-free mice

To characterize the mitochondrial stress response induced by Doxycycline in vivo in liver and kidneys, we administered doxycycline (Dox) at 500 mg/kg/day (mpkd) in the drinking water to 9 weeks-old germ-free C57BL/6J mice for 16 days, hence eliminating the potential confounding impacts of Doxycycline on the microbiome. Male 9 weeks-old C57BL/6J mice were treated for 16 days with 500 mg/kg/day doxycycline hyclate (Sigma) in drinking water and sacrified afterwards. All animals used in the experiments were randomly assigned to experimental or control groups. Mice were housed with ad libitum access to water and food and kept under a 12-hour dark/12-hour light cycle. As doxycycline is bitter, we supplemented the water for both conditions (treatments and controls) with 50 g/L sucrose. Drinking water was changed every 48 hours. Germ-free C57BL/6J mice were obtained from the Clean Mouse Facility, University of Bern (Bern, Switzerland). All animal experiments were carried out according to the institutional, national Swiss and EU ethical guidelines and were approved by the local animal experimentation committee of the Canton de Vaud.