Transcriptional analysis of dorsal skin from mice flown on the RR-5 mission

The objective of the Rodent Research-5 (RR-5) study was to evaluate bone loss in mice during spaceflight and to determine if treatment with a modified version of NEL-like molecule-1 (NELL-1) can reduce or prevent bone loss that would otherwise occur during spaceflight. To this end, a cohort of forty 30-weeks-old female BALB/cAnNTac mice were flown to the ISS and housed in the Rodent Habitat. Six days after launch half of the mice were treated with NELL-1 (10 mg/kg in 0.3 ml PBS), while the other half were treated with vehicle control (0.3 mls PBS). Fourteen days after launch animals were again treated with NELL-1 or vehicle control as before, except that all animals were also injected with the bone marker, calcein green (20 mg/kg in 0.1 ml). Injections of vehicle, NELL-1, and bone markers were intraperitoneal. After all forty mice on orbit received two treatments; ten control mice and ten experimental mice were randomly selected for live animal return (LAR). At approximately 30 days after launch the twenty LAR mice were transported live back to Earth. Animals were allowed to recover for 30 days in standard habitats before euthanasia via intraperitoneal injection with ketamine/xylazine. During the recovery, the animals received another two treatments. GeneLab received RNA later preserved dorsal skin from ten live animal return and ten matching ground control mice. These were from the vehicle control animals only. RNA was extracted, libraries generated (stranded, ribodepleted) and sequenced (target 60 M clusters at PE 150 bp).

啮齿类动物研究5号（Rodent Research-5，RR-5）的研究目标为评估太空飞行期间小鼠的骨丢失状况，并探究经修饰的NEL样分子1（NEL-like molecule-1，NELL-1）治疗能否减少或预防太空飞行中本应出现的骨丢失。
为此，研究团队选取40只30周龄的雌性BALB/cAnNTac小鼠，将其运送至国际空间站（International Space Station，ISS）并饲养于啮齿类动物栖息地舱（Rodent Habitat）内。发射后6天，半数小鼠接受NELL-1治疗（剂量为10 mg/kg，溶于0.3 ml磷酸盐缓冲液（Phosphate Buffered Saline，PBS）中），另一半则接受赋形剂对照处理（仅注射0.3 ml PBS）。
发射后14天，所有小鼠再次接受NELL-1或赋形剂对照治疗，与此前不同的是，所有小鼠同时注射了骨标记物绿钙素（calcein green，剂量为20 mg/kg，溶于0.1 ml溶剂中）。上述赋形剂、NELL-1及骨标记物均通过腹腔注射给药。
在轨的40只小鼠均完成两次治疗后，研究人员随机选取10只对照小鼠与10只实验组小鼠进行活体动物回收（live animal return，LAR）。发射后约30天，这20只完成活体回收的小鼠被活体运送回地球。
小鼠在标准饲养舱中恢复30天后，通过腹腔注射氯胺酮/甲苯噻嗪实施安乐死。恢复期间，小鼠额外接受了两次治疗。
GeneLab获取了10只活体回收小鼠与10只匹配的地面对照小鼠的RNA later保存的背部皮肤组织，且所有样本均来自赋形剂对照小鼠。
研究人员对样本进行RNA提取、文库构建（链特异性、核糖体RNA去除）及测序，测序参数为PE 150 bp，目标生成60 M个测序簇。