Transcriptional profiling, protein expression profiling, and phosphoprotein profiling of kidneys from mice flown on the RR-10 mission

The objective of the Rodent Research-10 Mission (RR-10) was to investigate how spaceflight affects the cellular and molecular mechanisms of normal bone tissue regeneration in space. To this end, ten (10) 14-15 weeks-old female B6129SF2/J Wild Type (WT), and ten (10) 14-15 weeks-old female B6;129S2-Cdkn1atm1Tyj/J (p21-null) mice received a pre-flight subcutaneous injection of the bone marker (Alizarin Red), and were then delivered to the ISS aboard SpaceX-21. At 7 days before euthanasia, all 20 mice received an intraperitoneal (IP) injection with a bone formation marker (Calcein). At 48 +/- 2 hours before euthanasia, all 20 mice received an IP injection with a second dose of Calcein as well as a cell proliferation marker (BrdU). Then, following 28-29 days in microgravity, the Flight mice were euthanized. Following removal of hindlimbs, carcasses were wrapped in aluminum foil, preserved in the CryoChiller, and stored at -80 C or colder until return to Earth. In addition to the Flight group, three ground control groups were also part of the study: Basal (representing the pre-launch state), Vivarium (standard vivarium housing for the same duration of time as flight), and Ground (flight habitat in the International Space Station Environment Simulator, ISSES). Twenty mice (10 of each strain) were included in each of these control groups (except Vivarium which included 12 of each strain). These were treated, euthanized and processed on the same schedule and in the same manner as the flight samples. At the end of RR-10 experiment, all frozen carcasses were partially thawed and kidney tissues were removed and preserved by flash freezing in LN2. Kidneys were kept at -80 C freezer until processing. To ensure that both transcriptional profiling and protein expression profiling datasets are representative of the entire kidney, whole kidneys were first pulverized into a fine powder on dry ice. This powder was then split into two equal fractions with half being used for RNA isolation and half being used for protein isolation. RNA was used to generate three different transcriptional profiling datasets: a 3' tag-Seq datasets (20 M clusters at SE 93 bp), a polyA enriched dataset (60 M clusters at PE 150 bp), and a ribodepleted dataset (60 M clusters at PE 150 bp). Protein was used to generate protein expression profiling, and phosphoprotein profiling using the iTRAQ method (Isobaric tags for relative and absolute quantitation). Transcriptional profiling dataset features WT samples from the Flight, Ground, Basal and Vivarium groups. Protein expression profiling and phosphoprotein profiling datasets exclude the Vivarium group.

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  "026:00"
]

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  "026:000"
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[
  "Biological and Physical Sciences"
]