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Comparative Transcriptomic Analysis of Adult Medaka Tissues Sampled after Adaptation to a Space Environment

To understand how humans adapt to space environments many experiments can be conducted on astronauts while they work aboard the Space Shuttle or the International Space Station (ISS). We also need animal experiments that can apply to human models and help prevent or solve the physical issues we face in space travel. The medaka is a suitable model fish for studying space adaptation because in the second International Microgravity Laboratory mission in 1994 adult fish mated successfully in space during 15 days of flight. In 2012 another space experiment Medaka Osteoclast was performed. Male and female fish (6 weeks old at launching) were maintained in the Aquatic Habitat system for 2 months in the ISS. The RNA-seq analysis of tissues from these fish will revealed tissue-specific responsiveness and common stress responses during space adaptation.

Rodent Research-1 (RR1) NASA Validation Flight: Mouse tibialis anterior muscle transcriptomic proteomic and epigenomic data

NASA s Rodent Research (RR) project is playing a critical role in advancing biomedical research on the physiological effects of space environments. Due to the limited resources for conducting biological experiments aboard the International Space Station (ISS) it is imperative to use crew time efficiently while maximizing high-quality science return. NASA s GeneLab project has as its primary objectives to 1) further increase the value of these experiments using a multi-omics systems biology-based approach and 2) disseminate these data without restrictions to the scientific community. The current investigation assessed viability of RNA DNA and protein extracted from archived RR-1 tissue samples for epigenomic transcriptomic and proteomic assays. During the first RR spaceflight experiment a variety of tissue types were harvested from subjects snap-frozen or RNAlater-preserved and then stored at least a year at -80C after return to Earth. They were then prioritized for this investigation based on likelihood of significant scientific value for spaceflight research. All tissues were made available to GeneLab through the bio-specimen sharing program managed by the Ames Life Science Data Archive and included mouse adrenal glands quadriceps gastrocnemius tibialis anterior extensor digitorum longus soleus eye and kidney. We report here protocols for and results of these tissue extractions and thus the feasibility and value of these kinds of omics analyses. In addition to providing additional opportunities for investigation of spaceflight effects on the mouse transcriptome and proteome in new kinds of tissues our results may also be of value to program managers for the prioritization of ISS crew time for rodent research activities.

Rodent Research-1 (RR1) National Lab Validation Flight: Mouse liver transcriptomic proteomic and epigenomic data

The Rodent Reasearch-1 National Lab (RR-1 CASIS) experiment was performed to study the effect of microgravity on muscle wasting. RNA DNA and protein were purified from RR-1 CASIS (the Center for the Advancement of Science in Space) liver samples. Groups included: Flight (FLT) dissected on-orbit (21 or 22 days after launch); age-matched Ground Controls (GC); and Basal Controls (BC euthanized at time of launch). RNA-Seq whole genome BS-Seq (bisulfite sequencing) and proteomic expression profiling were performed.

Transcriptomic analysis of liver from mice subjected to simulated spaceflight euthanasia freezing and tissue preservation protocols

To understand the molecular mechanisms affected by spaceflight it is essential to achieve high quality sample preservation on-orbit for downstream gene expression analysis. However sample preservation protocols must also be compatible with available equipment and crew time. NASA s Rodent Research (RR) missions have used various methods for euthanasia carcass preservation and tissue preservation. This study extends the sample preservation study performed by GeneLab in GLDS-49 which examined conditions used for the RR-1 mission to include conditions used for multiple RR missions and is designed to help determine factors which may confound data analysis. To determine whether these various factors affect changes in gene expression this ground-based study generated gene expression profiles measured by RNAseq from the livers of 20-21 week-old female C57BL/6J mice. Multiple interacting factors were investigated: 1) To understand how euthanasia protocols affect gene expression when mouse carcasses are slow frozen mice were euthanized by either euthasol injection ketamine/xylazine injection or CO2 inhalation and carcasses slow frozen on dry-ice mimicking carcass preservation in the MELFI on the ISS. Carcasses were thawed and RNA extracted from livers; 2) To understand how carcass preservation protocols affect gene expression mice were euthanized with euthasol and carcasses preserved by flash freezing in liquid nitrogen slow freezing on dry ice or immersion in RNAlater following three-way segmentation. Carcasses were thawed and RNA extracted from livers; 3) To understand how tissue preservation protocols affect gene expression mice were euthanized with euthasol and livers dissected and processed immediately or preserved by flash freezing in liquid nitrogen slow freezing on dry ice or immersion in RNAlater. Liver samples that were processed immediately were homogenized in RLT buffer and then either immediately further processed for RNA extraction or were stored for 70 days at -80C post-homogenization in sample RLT buffer prior to RNA extraction.

The Arabidopsis spaceflight transcriptome: a comparison of whole plants to discrete root hypocotyl and shoot responses to the orbital environment

Arabidopsis thaliana was evaluated for its response to the spaceflight environment in three replicated experiments on the International Space Station. Two approaches were used; GFP reporter genes were used to collect gene expression data in real time within unique GFP imaging hardware and plants were harvested on orbit to RNAlater for subsequent analyses of gene expression with using Affymetrix and SAGE transcriptome analyses. Three tissue types were examined (leaves hypocotyls and roots) and compared to analyses conducted with whole plants. Transcriptome analyses with whole plants suggested that the spaceflight environment had little impact on the transcriptome of arabidopsis however closer examination of selected tissues revealed that there are a number of tissue-specific responses that arabidopsis employs to respond to this novel environment

Transcriptome analysis of murine spleen in space

Our study aims to comprehensively understand effects induced by the space environment on mammals. To achieve this aim we analyze the male mice housed under environments as the artificial gravity and the microgravity (space environment) in Japanese Experiment Module JEM) of the International Space Station (ISS) on orbit for 35 days. After recovered these mice on the ground transcriptome analysis by next-generation sequencing technology is performed about spleen to examine alteration of gene expression in the space.

RNA-Seq transcriptome analysis of reactive oxygen species gene network in Mizuna plants grown in long-term space flight

Space environment is suspected to generate reactive oxygen species (ROS) and induce oxidative stress in plants however little is known about the gene expression of ROS gene network in plants grown in long-term space flight. RNA-Seq was used to define the large-scale gene expression profiles of Mizuna harvested after 27 days cultivation in the international space station to understand the molecular response and adaptation to space environment.Results: Total reads of transcripts from the Mizuna grown in the international space station as well as on the ground by RNA-Seq using next generation sequencing technology showed 8,258 and 14,170 transcripts up- and down-regulated in the space-grown Mizuna respectively when compared with those from the ground-grown Mizuna. A total of 20 in 32 ROS oxidative marker genes were up-regulated including high expression of 4 hallmarks and preferentially expressed gene associated with ROS-scavenging genes was thioredoxin glutaredoxin and alternative oxidase genes. In the transcription factors of ROS gene network MEKK1-MKK4-MPK3 OXI1-MKK4-MPK3 and OXI1-MPK3 of MAP cascades induction of WRKY22 by MEKK1-MKK4-MPK3 cascade induction of WRKY25 and repression of ZAT7 by Zat12 were suggested. RbohD and RbohF genes were up-regulated preferentially in NADPH oxidase genes which produce ROS.Conclusions: Our large-scale transcriptome analysis demonstrated that the space environment induced oxidative stress and ROS gene network was activated in the space-grown Mizuna some of which were common genes up-regulated by abiotic and biotic stress and were preferentially up-regulated genes by the space environment even though Mizuna grew in the space as well as on the ground showing that plants could acclimate to the space environment by reprograming the expression of ROS gene network.

Rodent Research-3-CASIS: Mouse kidney transcriptomic proteomic and epigenomic data

The Rodent Research-3 (RR-3) mission was sponsored by the pharmaceutical company Eli Lilly and Co. and the Center for the Advancement of Science in Space to study the effectiveness of a potential countermeasure for the loss of muscle and bone mass that occurs during spaceflight. Twenty BALB/c 18-weeks old female mice (ten controls and ten treated) were flown to the ISS and housed in the Rodent Habitat for 39-42 days. Twenty mice of similar age sex and strain were used for ground controls housed in identical hardware and matching ISS environmental conditions. Basal controls were housed in standard vivarium cages. Spaceflight ground controls and basal groups had blood collected then were euthanized had one hind limb removed and finally whole carcasses were stored at -80 C until dissection. All mice in this data set received only the control/sham injection.

Low dose (0.04 Gy) irradiation (LDR) and hindlimb unloading (HLU) microgravity in mice: brain transcriptomic and epigenomic data

The purpose of the present study was to evaluate damage in brain and eye in a ground-based model for spaceflight which includes prolonged unloading and low-dose radiation. Low-dose/Low-dose-rate (LDR) gamma-radiation using 57Co plates (0.04 Gy) was delivered whole-body to mature 6-month old female C57BL/6 mice to simulate the radiation component. Anti-orthostatic tail suspension was used to model the unloading fluid shift and physiological stress aspects of the microgravity component. Mice were hindlimb suspended and/or irradiated for 21 days. Tissues were collected at 7 days 1 4 and 9 months following simulated microgravity. Herein we proposed to use omics-based molecular phenotyping approach for identification and characterization of genomic signatures in multiple organ system associated with low-dose radiation and simulated microgravity.

Transcriptomic analysis of skin from mice subjected to chronic low-dose radiation hindlimb unloading or a combination of both

The purpose of this study was to evaluate transcriptional changes in mouse skin using a ground-based model for spaceflight. This model includes prolonged unloading and low-dose irradiation. Low-dose-rate gamma-radiation was delivered to 6-month old female C57BL/6J mice using 57Co plates (0.04 Gy) to simulate the radiation environment of spaceflight. Anti-orthostatic tail suspension was used to model the unloading fluid shift and physiological stress aspects of the microgravity component of spaceflight. Mice were hindlimb suspended and/or irradiated for 21 days. Mice were euthanized and dorsal skin was collected 7 days following treatment. RNA sequencing data was generated to assess transcriptional changes in these skin samples.

Identifying radiation exposure biomarkers from mouse blood transcriptome

Here we present a whole-genome survey of the murine transcriptomic response to physiologically-relevant radiation doses 2 and 8 Gy. There are 18 distinct biological samples here. Mice were exposed to ionizing radiation (Cesium-138 source) and whole blood was collected by cardiac puncture 6 hours post treatment. Doses were 0 (7 samples) 2 (5 samples) and 8 (6 samples) gy.

Low dose (0.04 Gy) irradiation (LDR) and hindlimb unloading (HLU) microgravity in mice: retina transcriptomic and epigenomic data

The purpose of the present study was to evaluate damage in brain and eye in a ground-based model for spaceflight which includes prolonged unloading and low-dose radiation. Low-dose/Low-dose-rate (LDR) gamma-radiation using 57Co plates (0.04 Gy) was delivered whole-body to mature 6-month old female C57BL/6 mice to simulate the radiation component. Anti-orthostatic tail suspension was used to model the unloading fluid shift and physiological stress aspects of the microgravity component. Mice were hindlimb suspended and/or irradiated for 21 days. Tissues were collected at 7 days 1 and 4 months following simulated microgravity. Herein we proposed to use omics-based molecular phenotyping approach for identification and characterization of genomic signatures in multiple organ system associated with low-dose radiation and simulated microgravity.

Rodent Research-1 (RR1) NASA Validation Flight: Mouse liver transcriptomic proteomic and epigenomic data

RR-1 is a validation flight to evaluate the hardware operational and science capabilities of the Rodent Research Project on the ISS. RNA DNA and protein were purified from liver tissues from RR-1 mice (female C57BL/6J 16wk old at time of launch). From each group two liver samples were collected and frozen immediately after euthanasia. The rest of the liver samples from each group were collected from frozen carcasses dissected post-flight. RNA-Seq whole genome and RNA BS-Seq (bisulfite sequencing) and proteomic expression profiling were performed. RNA extracted from these tissues was re-sequenced; these data are available as part of GLDS-168 (https://genelab-data.ndc.nasa.gov/genelab/accession/GLDS-168).

Transcriptomic analysis of quadriceps from mice subjected to simulated spaceflight euthanasia freezing and tissue preservation protocols

To understand the molecular mechanisms affected by spaceflight it is essential to achieve high quality sample preservation on-orbit for downstream gene expression analysis. However sample preservation protocols must also be compatible with available equipment and crew time. NASA s Rodent Research (RR) missions have used various methods for euthanasia carcass preservation and tissue preservation. This study extends the sample preservation study performed by GeneLab in GLDS-49 which examined conditions used for the RR-1 mission to include conditions used for multiple RR missions and is designed to help determine factors which may confound data analysis. To determine whether these various factors affect changes in gene expression this ground-based study generated gene expression profiles measured by RNAseq from the quadriceps of 20-21 week-old female C57BL/6J mice. Multiple interacting factors were investigated: 1) To understand how euthanasia protocols affect gene expression when mouse carcasses are slow frozen mice were euthanized by either euthasol injection ketamine/xylazine injection or CO2 inhalation and carcasses slow frozen on dry-ice mimicking carcass preservation in the MELFI on the ISS. Carcasses were thawed and RNA extracted from quadriceps; 2) To understand how carcass preservation protocols affect gene expression mice were euthanized with euthasol and carcasses preserved by flash freezing in liquid nitrogen slow freezing on dry ice or immersion in RNAlater following three-way segmentation. Carcasses were thawed and RNA extracted from quadriceps; 3) To understand how tissue preservation protocols affect gene expression mice were euthanized with euthasol and quadriceps immediately dissected and preserved by flash freezing in liquid nitrogen slow freezing on dry ice or immersion in RNAlater.

Transcriptome Profiles in Normal Human Bronchial Epithelial Cells after Exposure to gamma-rays and different HZE particles

Distinct transcriptome profiles in response to low-LET and high-LET and different radiation qualities of HZE particles. Total RNA obtained from HBEC3KT cells after 1 4 12 and 24 hours of radiation. Mock-irradiated samples at each time point and control samples before radiation (0 hour) were also collected.

Genetic Dissection of the Spaceflight Transcriptome Responses in Plants: are some responses unnecessary?

Experimentation on the International Space Station has reached the stage where repeated and nuanced transcriptome studies are beginning to illuminate the structural and metabolic differences between plants grown in space compared to plants on the Earth. Genes that are important in setting up the spaceflight responses are being identified; their role in spaceflight physiological adaptation are increasingly understood and the fact that different genotypes adapt differently is recognized. However the basic question of whether these spaceflight responses are required for survival has yet to be posed and the fundamental notion that spaceflight responses may be non-adaptive has yet to be explored. Therefore the experiments presented here were designed to ask if portions of the plant spaceflight response can be genetically removed without causing loss of spaceflight survival and without causing increased stress responses. The CARA experiment compared the spaceflight transcriptome responses of two Arabidopsis ecotypes Col-0 and WS as well as that of a PhyD mutant of Col-0. When grown with the ambient light of the ISS phyD displayed a significantly reduced spaceflight transcriptome response compared to Col-0 suggesting that altering the activity of a single gene can actually improve spaceflight adaptation by reducing the transcriptome cost of physiological adaptation. The WS genotype showed an even simpler spaceflight transcriptome response in the ambient light of the ISS more broadly indicating that the plant genotype can be manipulated to reduce the transcriptome cost of plant physiological adaptation to spaceflight and suggesting that genetic manipulation might further reduce or perhaps eliminate the metabolic cost of spaceflight adaptation. When plants were germinated and then left in the dark on the ISS the WS genotype actually mounted a larger transcriptome response than Col-0 suggesting that the in-space light environment affects physiological adaptation which further implies that manipulating the local habitat can also substantially impact the metabolic cost of spaceflight adaptation.

Rodent Research-1 (RR1) NASA Validation Flight: Mouse kidney transcriptomic proteomic and epigenomic data

Effect of microgravity on HUVECs (Human Umbilical vein Endothelial cells) cells and its transcriptome analysis.

Adaptation of humans in low gravity conditions is a matter of utmost importance when efforts are on to a gigantic leap in human space expeditions for tourism and formation of space colonies. In this connection cardiovascular adaptation in low gravity is a critical component of human space exploration. Deep high-throughput sequencing approach allowed us to analyze the miRNA and mRNA expression profiles in human umbilical cord vein endothelial cells (HUVEC) cultured under gravity (G) and stimulated microgravity (MG) achieved with a clinostat. The present study identified totally 1870 miRNAs differentially expressed in HUVEC under MG condition when compared to the cells subjected to unitary G conditions. The functional association of identified miRNAs targeting specific mRNAs revealed that miRNAs hsa-mir-496 hsa-mir-151a hsa-miR-296-3p hsa-mir-148a hsa-miR-365b-5p hsa-miR-3687 hsa-mir-454 hsa-miR-155-5p and hsa-miR-145-5p differentially regulated the genes involved in cell adhesion angiogenesis cell cycle JAK-STAT signaling MAPK signaling nitric oxide signaling VEGF signaling and wound healing pathways. Further the q-PCR based experimental studies of upregulated and downregulated miRNA and mRNAs demonstrate that the above reported miRNAs influence the cell proliferation and vascular functions of the HUVEC in MG conditions effectively. Consensus on the interactome results indicates restricted fluctuations in the transcriptome of the HUVEC exposed to short-term MG that could lead to higher levels of endothelial functions like angiogenesis and vascular patterning.

Rodent Research-1 (RR1) NASA Validation Flight: Mouse quadriceps muscle transcriptomic proteomic and epigenomic data

Rodent Research-3-CASIS: Mouse liver transcriptomic proteomic and epigenomic data