Transcriptional analysis of liver from mice flown on the RR-6 mission

The objective of the Rodent Research-6 (RR-6) study was to evaluate muscle atrophy in mice during spaceflight and to test the efficacy of a novel therapeutic to mitigate muscle wasting. The experiment involved an implantable subcutaneous nanochannel delivery system (nDS; between scapula), which delivered the drug formoterol (FMT; a selective Beta-2 adrenoceptor agonist), over the course of time. To this end, a cohort of forty 32-weeks-old female C57BL/6NTac mice were either sham operated or implanted with vehicle or treatment-filled nDS, launched in two Transporters (20 mice per Transporter) on SpaceX-13 on December 15, 2017. They were transferred to Rodent Habitats onboard the International Space Station (ISS), and maintained in microgravity for 29 days (N=20, Live Animal Return [LAR]), or >50 days (N=20, ISS Terminal). After 29 days, the 20 LAR animals were returned live to back to Earth on January 13, 2018,. After splashdown, the animals were ambulatory on-ground for ~4 days, until all subjects were processed during one day of dissections. There were two Baseline groups of animals sacrificed (LAR Baseline & FLT Baseline; N=20; 40 animals; ~36 weeks old) at Kennedy Space Center (KSC; 12/9/17). A Ground Control group mimicked the Flight LAR group, which was housed at KSC, then shipped alive to Novartis' Facilities, where both the LAR and LAR Ground Control groups were processed (~41 weeks old; 1/16/18). All were anesthetized with isoflurane, blood samples were obtained by closed-chest cardiac puncture, and the animals were euthanized by exsanguination and thoracotomy. The 20 ISS Terminal mice were anesthetized via intraperitoneal injection of ketamine/xylazine/acepromazine over the course of a four days of dissections (2/6/18 until 2/9/18; 53-56 days after launch; 44 weeks old at time of on-orbit dissections). Blood samples and euthanasia were conducted the same as LAR and Baseline. Following blood draw and hind limb dissection, the ISS-terminal animal carcasses were wrapped in aluminum foil, placed in a ziploc bag and placed in storage at -80C or colder until return. The ISS-terminal Ground Controls (at KSC) followed the same euthanasia timeline, methods, and preservation. The final processing of frozen ISS-terminal, frozen ISS-terminal Ground Controls and frozen 0-day FLT baseline animals were completed at Houston Methodist Research Institute, in Houston, TX (5/21/18 until 5/24/18). GeneLab received samples of liver from only sham treated animals (no drug treated animals) from the following groups Flight: LAR (n=10), ISS Terminal (n= 10); Ground Controls: LAR GC (N=9), ISS Terminal GC (N=10), LAR Baseline (n=10) ISS Terminal Baseline (n=10). Total RNA was extracted and sequenced at a target depth of 60 M clusters per sample (ribodepleted, paired end 150).

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