The Rodent Research-3 (RR-3) mission was designed to study the effectiveness of a potential countermeasure for the loss of muscle and bone mass that occurs during spaceflight. Myostatin is a protein secreted by myoblasts that inhibits muscle cell growth and differentiation. Mutations in myostatin, or drugs that block myostatin cause increases in muscle mass. The RR-3 experiment was sponsored by pharmaceutical company Eli Lilly and Co., and the Center for the Advancement of Science in Space and assessed the efficacy of myostatin inhibition to prevent skeletal muscle atrophy and weakness. Twenty BALB/c, 18-weeks-old female mice (10 controls and 10 treated) were flown to the ISS and housed in the AEM-X 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, and 10 mice for basal controls (initial time after launch). At the end of the experiment, spaceflight, ground controls, and basal groups had blood collected, were euthanized and one hind limb removed and whole carcasses were stored at -80°C until Biospecimen Sharing Program (BSP) dissection. GeneLab received various tissues for multi-omic analysis from flight, ground, and basal (pre-flight) mice that received only the control injection. All myostatin-inhibitor injected mice were processed exclusively by Eli-Lily.
This investigation was funded by the ISS National Lab, the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division, and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
Microbes interact with humans in complex ways and understanding how they respond to the spaceflight environment is important to the success of future manned spaceflight missions. The BRIC-23 mission was designed to measure the response of Bacillus subtilis and Staphylococcus aureus to the spaceflight environment. This experiment aimed to produce high-quality omics data from B. subtilis and S. aureus grown aboard the International Space Station (ISS) to allow comparison to matched ground controls. These datasets will be publically released through the GeneLab data system to allow widespread distribution. There were two primary objectives for this experiment: (1) Demonstrate all post-flight processes and operations required for successful completion of GeneLab Reference Missions conducted on ISS, and (2) Generate high-quality GeneLab Reference Mission omics data sets for two prokaryotic model organisms, Bacillis subtilis and Staphylococcus aureus. Freezing Control Experiment: The BRIC hardware has significant thermal inertia, thus the freezing rate of samples placed at -80°C is quite slow. This could affect RNA-sequencing, proteomic and metabolic data sets. In an effort to understand how slow freezing could affect these data sets, a control experiment was designed in which B. subtilis and S. aureus were grown in petri plates and either slow frozen to -80°C at a rate matching the BRIC-23 spaceflight samples or processed immediately to harvest RNA and protein.
This investigation was funded by the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division, and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
Spaceflight is known to affect immune cell populations. In particular, splenic B-cell numbers decrease during spaceflight and in ground-based physiological models. Although antibody isotype changes have been assessed during and after spaceflight, an extensive characterization of the impact of spaceflight on antibody composition has not been conducted in mice. Next Generation Sequencing and bioinformatic tools are now available to assess antibody repertoires. We can now identify immunoglobulin gene-segment usage, junctional regions, and modifications that contribute to specificity and diversity. Due to limitations on the International Space Station, alternate sample collection and storage methods must be employed. Our group compared Illumina MiSeq® sequencing data from multiple sample preparation methods in normal C57Bl/6J mice to validate that sample preparation and storage would not bias the outcome of antibody repertoire characterization. In this report, we also compared sequencing techniques and a bioinformatic workflow on the data output when we assessed the IgH and Igκ variable gene usage. Our bioinformatic workflow has been optimized for Illumina HiSeq® and MiSeq® datasets, and is designed specifically to reduce bias, capture the most information from Ig sequences, and produce a data set that provides other data mining options.
This work was supported by NASA grants NNX13AN34G and NNX15AB45G, NIH grant GM103418, the Molecular Biology Core supported by the College of Veterinary Medicine at Kansas State University, and the Kansas State University Johnson Cancer Center.
Interplanetary human spaceflight represents a formidable medical challenge, but also provides a unique platform for investigating human adaptation to extreme environmental changes. Understanding the long-term effects of isolation has relevance in a range of scenarios, and it is well recognized that a better understanding of the relationship between environmental exposure and the epigenome can lead to more effective preventive measures. Here we conduct a longitudinal epigenetic, mood state, and biochemical profiling of 6 crew members in an experiment simulating a 520-day mission to Mars. Illumina HumanMethylation450 BeadChip was used to obtain DNA methylation profiles. Firstly, we found that long-term isolation can induce global DNA methylation remodeling, and this change seems to be an active adaptation (rather than a random process or a by-product of the isolation). This study is the first to demonstrate the dynamic relationship between global epigenetic remodeling and isolation-induced mood state and biochemical changes. Secondly, by considering the location of methylation sites within the genome and using gene-pathway annotation, we were able to identify pathways that were significantly enriched in methylation events and consider their association with specific function and the timeline of the mission. Thirdly, via our definition of epi-entropy, a measure of entropy adapted to methylation events, we observed that the methylation remodeling produced a marked reduction in epi-entropy. Results suggest that DNA methylation change is an indicator of change rather than its by-product, i.e., there is a psychology-epigenome-metabolism model of long-term depression; DNA methylation programs the environment signal into the epigenome, which is subsequently transformed into the biochemical output and health outcome. Thus, longitudinal epigenetic profiling could code the effect of isolation and act as early indicators of latent health outcome. A longitudinal epigenetic, mood state and biochemical profiling of 6 crew members in an experiment simulating a 520-day mission to Mars. 36 samples of blood cell DNA methylation profiling were obtained by Illumina HumanMethylation450 BeadChip, across 6 sampling points during the 520 days mission for all of the 6 crew members.
Upon weightlessness and microgravity, deleterious effects on the neurosensory and neurovestibular systems, haematological changes, and deconditioning of musculoskeletal, cardiovascular and cardiopulmonary functions have been reported. In particular, loss of muscle mass and strength are triggered by weightlessness in humans during space flights, that is similarly observed as a result of physical inactivity conditions and aging on Earth. However, skeletal muscle tissue is of paramount importance for health maintenance (e.g. being essential to locomotion, heat production, and metabolism). To better prevent or eventually treat microgravity-induced muscle atrophy, its underlying mechanisms have first to be characterized in detail. Using cutting-edge quantitative proteomics, the aim of the present study was therefore to get an in-depth view of the molecular regulations triggered by space conditions in skeletal muscles of mice during the 30-day flight of the BION-M1 biosatellite. As muscles differing in their fiber type composition appear to respond differently to microgravity (see above), we characterized here the differential response of the soleus, extensor digitorum longus and vastus lateralis muscles.
This work was supported by the French Space Agency, CNRS and Strasbourg University (H2E project), French Proteomic Infrastructure (ProFI; ANR-10-INSB-08-03), and the European Space Agency PRODEX programme with Enterprise Ireland. We thank Dr. B. Shenkman from the Institute for Biomedical Problems of the Russian Academy of Sciences (Moscow, Russia).
During spaceflight, bone mineral density is decreased by the influence of osteoclast activation, which molecular mechanism is expectantly investigated. In the study of medaka bone development, the system of vertebra formation was investigated, and firstly identified the presence of osteoclasts in medaka. Moreover, osteoclast resorbing activity was affected by hypergravity, indicating the possibility that we can investigate the effect of microgravity on osteoclasts in space. To find this effect, the alteration of osteoclast activity under microgravity was examined, with the histological analysis or the expression analysis by RNA in-situ hybridization. Furthermore, since we have succeeded the establishment of the medaka osteoclast-specific transgenic lines, we perform the in-vivo imaging analyses for gene expression and cell mobility. Finally, to examine the gravity sensing system, we employ tooth and bone as the high-density organs, which are highly sensitive to gravity, and perform the histological analysis and the gene expression analysis of such gravity-sensitive tissues at surrounding pharyngeal teeth and supporting bone.
RNAseq libraries constructions and analysis was partially supported by Russian Science Foundation grant for international groups (No. 14-44-00022) work was mainly by grants-in-aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan (grant no. 25514003, 16K15778, 16H01635) and partly by the Ground-based Research Program for Space Utilization promoted by the Japan Space Forum.
This study focuses on the hardiest microorganisms inhabiting the ISS, their diversity, and capabilities to resist certain stresses. Dust samples were specifically selected from the Russian modules that were obtained 8-10 years ago, and stored since then under sealed conditions on Earth. Targeting long-time survivors and spore-forming microorganisms, the cultivable microbial community of these samples was assessed to obtain model microbial strains that could help analyze specific adaptation toward environmental stresses such as desiccation and lack of nutrients. In this study, microorganisms were analyzed with respect to their resistance toward thermal stress, and exposure to clinically relevant antibiotics. In addition, the bacterial and archaeal community was assessed via molecular methods (NGS sequencing), and the new data compared with previously derived information from the ISS microbiome.
The ARBEX project received funding from of the Austrian Forschungsforderungsgesellschaft FFG (Project number: 847977).
These investigations studied the fundamentals of how plants perceive gravity and develop in microgravity. It informs how gene regulation is altered by spaceflight conditions.
This study was funded by NASA grant NNX10AF44G.
Controlled hypobaria presents biology with an environment that is never encountered in terrestrial ecology, yet the apparent components of hypobaria are stresses typical of terrestrial ecosystems. High altitude, for example, presents terrestrial hypobaria always with hypoxia as a component stress, since the relative partial pressure of O2 is constant in the atmosphere. Laboratory-controlled hypobaria, however, allows the dissection of pressure effects away from the effects typically associated with altitude, in particular hypoxia, as the partial pressure of O2 can be varied. In this study, whole transcriptomes of plants grown in ambient (97 kPa/pO2 = 21 kPa) atmospheric conditions were compared to those of plants transferred to five different atmospheres of varying pressure and oxygen composition for 24 h: 50 kPa/pO2 = 10 kPa, 25 kPa/pO2 = 5 kPa, 50 kPa/pO2 = 21 kPa, 25 kPa/pO2 = 21 kPa, or 97 kPa/pO2 = 5 kPa. The plants exposed to these environments were 10-day-old Arabidopsis seedlings grown vertically on hydrated nutrient plates. In addition, 5-day-old plants were also exposed for 24 h to the 50 kPa and ambient environments to evaluate age-dependent responses. The gene expression profiles from roots and shoots showed that the hypobaric response contained more complex gene regulation than simple hypoxia, and that adding back oxygen to normoxic conditions did not completely alleviate gene expression changes in hypobaric responses.
Funding source: This work was funded by grants NNX13AM46G and NNA04CC6 to RF and AP from NASA Space Life and Physical Sciences managed through Kennedy Space Center.
Microgravity as well as chronic muscle disuse are two causes of low back pain originated at least in part from paraspinal muscle deconditioning. At present, no study has investigated the complexity of the molecular changes in human or mouse paraspinal muscles exposed to microgravity. The aim of this study was to evaluate longissimus dorsi and tongue (as a new potential in-flight negative control) adaptation to microgravity at global gene expression level. C57BL/N6 male mice were flown aboard the BION-M1 biosatellite for 30 days (BF) or housed in a replicate flight habitat on ground (BG). Global gene expression analysis identified 89 transcripts differentially regulated in longissimus dorsi of BF vs. BG mice (False Discovery Rrate < 0,05 and fold change < -2 and > +2), while only a small number of genes were found differentially regulated in tongue muscle (BF vs. BG = 27 genes). C57BL/N6 mice were randomly divided into three groups: Bion Flown (BF), mice flown aboard the Bion M1 biosatellite in microgravity environment for 30 days; Bion Ground (BG), mice housed in the same habitat of flown animals but exposed to Earth gravity; and Flight Control (FC), mice housed in a standard animal facility.
This work was supported from grants of the Department of Economics and Technology of the German Government (BMWi) through the German AeroSpace Board, Deutsches Zentrum fur Luft- und Raumfahrt (DLR), e.V. Bonn, Germany (grant # 50 WB821, 1121, and 1421 to DB).
A combination therapy of electromagnetic fields (EMF) and simulated microgravity (SMG) has not been examined in regenerative medicine of cartilage. In the present study, a bioreactor system using extremely low-frequency EMF and SMG was applied during the chondrogenic differentiation of human mesenchymal stem cells (hMSCs). It was hypothesized that a beneficial effect of EMF regarding chondrogenesis (COL2A) could be combined with an avoiding effect of SMG regarding hypertrophy (COLXA1) of cartilage. Pellet cultures of hMSCs formed cartilaginous tissue under the addition of growth factors (FGF; TGF-beta3). Pure SMG reduced COLXA1 expression but also COL2A expression of hMSCs. Pure EMF showed no gene expression changes of hMSCs during chondrogenic differentiation. Combining EMF/SMG resulted in a re-increase of COL2A but did not reach control levels. The COL2A to COLXA1 ratio of combined EMF/SMG was not significantly different from control levels. The combination therapy of EMF/SMG did not significantly improve the chondrogenic potential of hMSCs chondrogenic differentiation, electromagnetic stimulation-control, or 1 timepoint with/without stimulation.
Ionizing radiation (IR) can be extremely harmful for human cells since an improper DNA-damage response (DDR) to IR can contribute to carcinogenesis initiation. Perturbations in the DDR pathway can originate from alteration in the functionality of the microRNA-mediated gene regulation, being microRNAs (miRNAs) small noncoding RNA that act as post-transcriptional regulators of gene expression. In this study, insight was gained into the role of miRNAs in the regulation of DDR to IR under microgravity, a condition of weightlessness experienced by astronauts during space missions, which could have a synergistic action on cells, increasing the risk of radiation exposure. The miRNA expression profile of human peripheral blood lymphocytes (PBL) was analyzed and incubated for 4 and 24 h in normal gravity (1 g), and in modeled microgravity (MMG) during the repair time after irradiation with 0.2 and 2Gy of gamma-rays. Results show that MMG alters miRNA expression signature of irradiated PBL by decreasing the number of radio-responsive miRNAs. Moreover, let-7i*, miR-7, miR-7-1*, miR-27a, miR-144, miR-200a, miR-598, miR-650 are deregulated by the combined action of radiation and MMG. Integrated analyses of miRNA and mRNA expression profiles, carried out on PBL of the same donors, identified significant miRNA-mRNA anti-correlations of DDR pathway. Gene ontology analysis reports that the biological category of response to DNA damage is enriched when PBL are incubated in 1 g but not in MMG. Moreover, some anti-correlated genes of p53-pathway show a different expression level between 1 g and MMG. Functional validation assays using luciferase reporter constructs, confirmed miRNA-mRNA interactions derived from target prediction analyses. By integrating the transcriptome and microRNome, evidence shows that modeled microgravity can affect the DNA-damage response to IR in human PBL. The gene expression signature was defined in PBL irradiated with gamma-rays (2.0 Gy), and incubated in modeled microgravity (MMG) and parallel ground conditions (1g) for 24h. Five independent experiments were performed for each donor to address which mRNAs were regulated on IR stress. The level of each transcript was represented as Log2.
This work was supported by grants from the Italian Space Agency (ASI, XMAB-from Molecules to Man, 1/014/06/0) to L.C., from the Associazione Italiana per la Ricerca sul Cancro-AIRC (2008/6014) to G.L. and from the University of Padova (CPDA061783) to M.M.
Experimentation on the International Space Station (ISS) 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. CARA Seed Lines and Planting: Three seed lines Wild-Type Wassilewskija (Ws), Columbia-0 (Col-0) and Col-0 PhyD (phyD) Mutants were tested for viability, sterility and ability to maintain dormancy before the launch. Tested batches of seeds were planted on phytagel plates as one genotype per plate for gene expression analysis in replicates of three. One set was planted for the flight and one for ground control. The plates were wrapped such that every surface of the plate was covered by two layers of Duvetyn Black-Out cloth (Seattle Fabrics) (Sng et al, 2014). The plates were stored 4° C until launch, and was then launched in a cold-stow bag to maintain the plates at 4° C until integration and activation on the ISS. On-orbit operations and harvest: The dormant plates were activated on station by removing the Black-Out cloth wrapping 12 days after launch. The plates were then placed on a fabric that was mounted in the US Laboratory module on the wall adjoining the MELFI freezer and secured using Velcro. The plants were allowed to grow on orbit for 11 days; some in the ambient light of ISS and some in the dark. The dark-grown plates were first activated by exposing the seeds to light for 4 hours, and then re-wrapped in Black-Out cloth for the duration of the growth period. A corresponding set of seedlings were grown as ground control in KSC. At 11 days, seedlings were photographed, harvested into KFT containing RNAlater solutions and returned for post-flight analysis. All plates were harvested into KFTs with their counterpart (e.g. Light 1 was harvested with Dark 1). Once the plants were placed in the KFTs, the KFT was actuated with RNAlater to preserve the sample. At 24 hours post-harvest, KFTs were then transferred to MELFI, the -32°C freezer. Following SpaceX-3 splashdown in the Pacific Ocean, the KFTs transferred to the Cold Stowage charter plane at the Long Beach Airport, placed into an insulated shipper with dry ice, and flown to Johnson Space Center. The KFTs were then transferred via FedEx ground to the Kennedy Space Center. The KFTs were removed from dry ice and transferred to a -80°C freezer on May 22, 2014. The PIs retrieved the plant samples from the KFTs on May 29, 2014 and transferred the samples back to the University of Florida. The harvested material was used to compare the transcriptomes of each genotype. The patterns of gene expression were compared among genotypes within a treatment (spaceflight or unit gravity) and between treatments (spaceflight versus ground control) for each genotype.
RNA-seq libraries were prepared using the Illumina TruSeq RNA kit, and the TrueSeq method was employed for mRNA enrichment. The libraries were quantified and samples were multiplexed in each lane of the flow cell. Cluster generation was performed and then sequenced on the Illumina HiSeq1000 system. Reads were mapped on the Human Genome Reference and a normalized expression table was generated. Among differentially expressed genes, 53 were up-regulated and 75 were down-regulated. Data demonstrate increased expression of genes associated with growth, development, and pro-survival in cardiac progenitors cultured under simulated microgravity compared with those cultured under standard gravity. RNA-sequencing analysis was performed to compare global gene expression profiles of cells at differentiation day 8 under simulated microgravity vs. standard gravity.
This study was supported in part by grants GA-2014-126 from the Center for the Advancement of Science in Space (CASIS) and R21HL118454 from the NIH.
Ionizing radiation (IR) can be extremely harmful for human cells since an improper DNA-damage response (DDR) to IR can contribute to carcinogenesis initiation. Perturbations in the DDR pathway can originate from alteration in the functionality of the microRNA-mediated gene regulation, being microRNAs (miRNAs) small noncoding RNA that act as post-transcriptional regulators of gene expression. In this study, insight was gained into the role of miRNAs in the regulation of DDR to IR under microgravity, a condition of weightlessness experienced by astronauts during space missions, which could have a synergistic action on cells, increasing the risk of radiation exposure. The miRNA expression profile of human peripheral blood lymphocytes (PBL) was analyzed, and incubated for 4 and 24 h in normal gravity (1 g), in modeled microgravity (MMG) during the repair time after irradiation with 0.2 and 2Gy of gamma-rays. Results show that MMG alters miRNA expression signature of irradiated PBL by decreasing the number of radio-responsive miRNAs. Moreover, let-7i*, miR-7, miR-7-1*, miR-27a, miR-144, miR-200a, miR-598, and miR-650 are deregulated by the combined action of radiation and MMG. Integrated analyses of miRNA and mRNA expression profiles, carried out on PBL of the same donors, identified significant miRNA-mRNA anti-correlations of the DDR pathway. Gene ontology analysis reports that the biological category of response to DNA damage is enriched when PBL are incubated in 1 g but not in MMG. Moreover, some anti-correlated genes of p53-pathway show a different expression level between 1 g and MMG. Functional validation assays using luciferase reporter constructs, confirmed miRNA-mRNA interactions derived from target prediction analyses. By integrating the transcriptome and microRNome, evidence shows that modeled microgravity can affect the DNA-damage response to IR in human PBL.
This work was supported by grants from the Italian Space Agency (ASI, XMAB-from Molecules to Man, 1/014/06/0) to L.C., from the Associazione Italiana per la Ricerca sul Cancro-AIRC (2008/6014) to G.L. and from the University of Padova (CPDA061783) to M.M.
To reveal outcomes of microgravity on molecular processes within the cellular environment we have employed a mass-spectrometry-based proteomics approach. Proteomics analysis based on mass spectrometry allows for the relative quantitation of a large number of proteins concurrently, and in a relatively unbiased manner. Mass-spectrometry-based proteomics can be rendered even more informative by addition of a labeling component to understand the dynamics of the changing protein content. In this study, we utilized a combination of proteomics techniques, namely label-free quantification and dynamic stable-isotope labeling by amino acids in cell culture (Dynamic SILAC) to characterize the microgravity stress response in primary cardiomyocytes.
Funding Source: This work was supported by the National Aeronautics and Space Association grant NNX12AK76G to Dawn E. Bowles.
DLD-1 and MOLT-4 human cancer cell lines were cultured in a rotating cell culture system to simulate microgravity. The mRNA expression profile was observed in comparison to static controls. Cells were grown in 10mL rotating vessels in an RCCS and in 60mm Petri dishes (test, control, respectively). Two replicates of test (microgravity) and control (static), each from DLD-1 and MOLT-4, were analyzed by microarray. Simulated microgravity affected the solid tumor cell line DLD-1 markedly, which showed a higher percentage of dysregulated genes compared to the hematological tumor cell line, MOLT-4. Microgravity affects the cell cycle of DLD-1 cells and disturbs expression of cell cycle regulatory gene networks. Multiple microRNA host genes were dysregulated and significantly, mir-22, tumor suppressor microRNA, is highly upregulated in DLD-1.
This work was supported by Defense Research Development Organization (DLS/81/48222/LSRB-189/ID/2009 and DLS/81/48222/LSRB- 273/SH&DD/2013) to RSV.
Proton irradiation is touted for its improved tumor targeting due to the physical advantages of ion beams for radiotherapy. Recent studies from our laboratory have shown that, in addition to targeting advantages, proton irradiation can inhibit angiogenic and immune factors and thereby modulate tumor progression. High-energy protons also constitute a principal component of the galactic cosmic rays to which astronauts are exposed. Increased understanding of the biological effects of proton exposure would thus contribute to both improved cancer therapy and carcinogenesis risk assessment for space travel. In addition, age plays a major role in tumor incidence and is a critical consideration for estimating cancer risk. We investigated the effects of host age and proton exposure on tumor progression. Tumor lag time and growth dynamics were tracked following injection of murine Lewis lung carcinoma (LLC) cells into young (68 day) versus old (736 day) mice with or without coincident irradiation. Tumor progression was suppressed in old compared to young mice. Differences in progression were further modulated by proton irradiation (1GeV), with increased inhibition evident in old mice. Through global transcriptome analysis, TGFB1 and TGFB2 were determined to be key players that contributed to the tumor dynamics observed. These findings point to older hosts providing decreased systemic tumor support, which can be further inhibited by proton irradiation.
This material is based upon work supported by the National Aeronautics and Space Administration under NSCOR grants NNJ06HA28G and NNX11AK26G issued through the Human Research Program and by award number U54CA149233 from the National Cancer Institute to LH.
RAD51, a multifunctional protein, plays a central role in DNA replication and homologous recombination repair, and is known to be involved in cancer development. We identified a novel role for RAD51 in innate immune response signaling. Defects in RAD51 lead to the accumulation of self-DNA in the cytoplasm, triggering a STING-mediated innate immune response after replication stress and DNA damage. Our data suggest that in addition to playing roles in homologous recombination-mediated DNA double-strand break repair and replication fork processing, RAD51 is also implicated in the suppression of innate immunity. Thus, our study reveals a previously uncharacterized role of RAD51 in initiating immune signaling, placing it at the hub of new interconnections between DNA replication, DNA repair, and immunity. Overall design of the gene expression analysis of WT and sh RAD51 HT1080 cells was carried out before and after 1 Gy of high LET Fe particle radiation.
National Aeronautics and Space Administration [NNX13AD57G and NNX15AE06G]; Cancer Prevention and Research Institute of Texas [RP160520]; National Institute of Health [R01AG053341] to A.A
The whole-genome sequences of eight fungal strains that were selected for exposure to microgravity at the International Space Station are presented here. These baseline sequences will help to understand the observed production of novel bioactive compounds.
Part of the research described in this publication was carried out at the Jet Propulsion Laboratory (JPL), California Institute of Technology, under a contract with NASA. This research was supported by the JPL Advanced Concept Development fund awarded to K. Venkateswaran, which funded a student fellowship to A. Blachowicz. Part of the research described in this publication was carried out at Lawrence Berkeley National Laboratory (T. Torok) and the University of Southern California (C.Wang), which funded a student fellowship to J.Romsdahl.
The general objective of this study was to determine modulation of gene expression by environmental factors, with a special emphasis on bone formation. For this reason, the specific period of treatment was chosen between 5-6 days post-fertilization (dpf), when bone formation and calcification are taking place. Zebrafish larvae were placed at 5 dpf into a large diameter centrifuge and brought to a gravitational force of 3g or 5g for 24 hours. This treatment causes a clear increase of bone formation, as illustrated by cranial skeleton staining of the bone matrix by Alizarin Red, by morphometric analysis of the resulting images, and by gene expression studies of selected genes. Thus, a whole genome micro-array experiment was conducted to identify genes that may be involved in the observed effect on bone formation.
This work was supported by the Fonds de la Recherche Fondamentale Collective; 2.4555.99/ 2.4542.00/2.4561.10, the SSTC; PAI: P5/35, the University of Liege; GAME project, the European Space Agency projects AO-99-LSS-003 and AO-99-LSS-006, the Belgian Space Agency Prodex projects FISH-GSIM and FISH-SIM.
Microgravity, or an altered gravity environment from the static 1g, has been shown to influence global gene expression patterns and protein levels in cultured cells or animals. However, it is unclear how these changes in gene and protein expressions are related to each other or are related to other factors regulating such changes. Recent advancements in the field of molecular biology revealed that a different class of RNA, the small non-coding microRNA (miRNA), can have a broad effect on gene expression networks by mainly inhibiting the translational process. In this experiment conducted on the International Space Station, the hypotheses to be tested are: miRNA profiles will be altered in the space environment, and cellular responses to DNA damage in space are different from those on the ground.
This work was supported by the NASA Fundamental Space Biology Program.
Mice were flown onboard STS-135 and returned to Earth for analysis. Livers were collected within 3-4 hours of landing and snap frozen in liquid nitrogen. Samples were shipped to Metabolon, Inc. for analysis.
This study was supported by the NASA Cooperative Agreement NNX10AJ31G 'Cooperative Research in Proton Space Radiation' and NNX10AE39G, the Loma Linda University Medical Center Department of Radiation Medicine, and the University of Colorado Anschutz Medical Center Department of Anesthesiology. Liver samples were obtained through the NASA Biospecimen Sharing Program.
To explore possible mechanisms, changes in gene expression profiles implicated in oxidative stress and in ECM remodeling in mouse skin were examined after space flight. The metabolic effects of space flight in skin tissues were also characterized.
This study was supported by NASA grant NNX10AJ31G and the LLUMC Dept. of Radiation Medicine.
Spaceflight imposes numerous adaptive challenges for terrestrial life. The reduction in gravity or microgravity, represents a novel environment that can disrupt homeostasis of many physiological processes. Additionally, it is becoming increasingly clear that an organism’s microbiome is critical for host health and examining its resiliency in microgravity represents a new frontier for space biology research. In this study, we examine the impact of microgravity on the interactions between the squid Euprymna scolopes and its beneficial symbiont Vibrio fischeri, which form a highly specific binary mutualism. First, animals inoculated with V. fischeri aboard the space shuttle showed effective colonization of the host light organ, the site of the symbiosis, during space flight. Second, RNA-Seq analysis of squid exposed to modeled microgravity conditions exhibited extensive differential gene expression in the presence and absence of the symbiotic partner. Transcriptomic analyses revealed in the absence of the symbiont during modeled microgravity there was an enrichment of genes and pathways associated with the innate immune and oxidative stress response. The results suggest that V. fischeri may help modulate the host stress responses under modeled microgravity. This study provides a window into the adaptive responses that the host animal and its symbiont use during modeled microgravity.
This study was supported by NASA Space Biology program grant NNX13AM44G.
Rodent Research-1 (RR1) NASA Validation Flight:
Mouse tibialis anterior muscle transcriptomic, proteomic, and epigenomic data
Mouse soleus muscle transcriptomic and epigenomic data
Mouse quadriceps muscle transcriptomic, proteomic, and epigenomic data
Mouse kidney transcriptomic, proteomic, and epigenomic data
Mouse gastrocnemius muscle transcriptomic, proteomic, and epigenomic data
Mouse eye transcriptomic and epigenomic data
Mouse extensor digitorum longus muscle transcriptomic and epigenomic data
Mouse adrenal gland transcriptomic, proteomic, and epigenomic data
Mouse liver transcriptomic, proteomic, and epigenomic data
The samples were generated from protein and RNA extracts of Arabidopsis seedlings grown in mock BRIC hardware and were grown simultaneously with the ground control samples from the BRIC-20 study. “n2” samples were preserved by direct submersion in liquid nitrogen, whereas the “rl” samples were submerged in RNAlater for 12 hours prior to freezing at -80C. The protein and RNA extracts were analyzed in the same manner as the space flight and ground control samples.
The effect of microgravity on gene expression in C.elegans was comprehensively analyzed by DNA microarray (Agilent). Hypergravity and clinorotation experiments were performed as reference against the flight experiment. Mix staged C. elegans N2 was exposed microgravity for 10 days.
ICE-FIRST was a collaborative effort among four nations: France, United States, Japan, and Canada.
Prolonged skeletal unloading through bedrest results in bone loss similar to that observed in elderly osteoporotic patients, but with an accelerated timeframe. This rapid effect on weight-bearing bones is also observed in astronauts who lose up to 2% of their bone mass per month spent in space. Despite important implications for space-flight travelers and bedridden patients on Earth, the exact mechanisms involved in disuse osteoporosis have not been elucidated. Parathyroid hormone-related protein (PTHrP) regulates many physiological processes including skeletal development, and has been proposed as a gravisensor. To investigate the role of PTHrP in microgravity-induced bone loss, trabecular osteoblasts (TOs) from Pthrp+/+ and -/- mice were exposed to simulated microgravity for 6 days. Viability of TOs decreased in inverse proportion to PTHrP expression levels. Microarray analysis of Pthrp+/+ TOs after 6 days at 0g revealed expression changes in genes encoding prolactins, apoptosis and survival molecules, bone metabolism and extra-cellular matrix composition proteins, chemokines, IGF family and Wnt-related signaling molecules. Importantly, 88% of 0g-induced expression changes in Pthrp+/+ cells overlap those observed in Pthrp-/- cells in normal gravity. Pulsatile treatment with PTHrP1-36 peptide during microgravity exposure reversed a large proportion of 0g-induced changes in Pthrp+/+ TOs. Results confirm PTHrP efficacy as an anabolic agent to prevent microgravity-induced cell death in TOs. Total RNA samples extracted from Pthrp+/+and -/- trabecular osteoblasts (TOs) exposed for 6 days to simulated 0g in Synthecon rotating cell, or left 6 days in culture at 1g. Cells had either been treated with a pulsatile treatment (2 h/day) of PTHrP1-36 peptide (10-8M) or received a change in growth medium.
This work was completed through the Canadian Space Agency (CSA) E/Osteo program contract # 9F-007-05-1657 and a grant to Dr. David Goltzman from the Canadian Institutes of Health Research (FRN-86703).
The effect of microgravity on C. elegans gene expression was analyzed by the whole genome microarray. The worms were cultivated under microgravity for 4 days in the Japanese module of the International Space Station. C. elegans N2 were exposed to microgravity for 4 days. The worms synchronously were cultivated from L1 larvae to adult. There were two control groups, onboard 1G and ground 1G control.
This experiment was supported by the Cell Biology Experiment Project conducted by the Institute of Space and Astronautical Science in JAXA, and was funded in part by JSPS KAKENHI Grant Numbers 26506029, 15H05937, the Medical Research Council UK (G0801271), and National Institutes of Health (NIH NIAMS ARO54342).
The water flea Daphnia is an interesting candidate for bioregenerative life support systems (BLSS). These animals are particularly promising because of their central role in the limnic food web and its mode of reproduction. However, the response of Daphnia to altered gravity conditions has to be investigated, especially on the molecular level, to evaluate the suitability of Daphnia for BLSS in space. In this study, a proteomic approach was used to identify key proteins and pathways involved in the response of Daphnia to simulated microgravity generated by a 2D-clinostat. Five biological replicates were analyzed using 2D-DIGE proteomic analysis. Identified were 109 protein spots differing in intensity (p < 0.05). Substantial fractions of these proteins are involved in actin microfilament organization, indicating the disruption of cytoskeletal structures during clinorotation. Furthermore, proteins involved in protein folding were identified, suggesting altered gravity induced break-down of protein structures in general. In addition, simulated microgravity increased the abundance of energy metabolism related proteins, indicating an enhanced energy demand of Daphnia. Most of the Daphnia protein sequences are well conserved throughout taxa, indicating that the response to altered gravity conditions in Daphnia follows a general concept.
Financial support was received from BMWi and DLR (Grant No. 50WB1029), as well as the ESA-GBF-program (Contract No. 4000103571).
Microgravity exposure as well as chronic muscle disuse are two of the main causes of physiological adaptive skeletal muscle atrophy in humans and murine animals. The aim of this study was to investigate, at both morphological and global gene expression levels, skeletal muscle adaptation to microgravity in mouse soleus and extensor digitorum longus (EDL). Adult male mice C57BL/N6 were flown aboard the BION-M1 biosatellite for 30 days on orbit (BF) or housed in a replicate flight habitat on Earth (BG) as reference flight control. This study investigated for the first-time, gene expression adaptation to 30 days of microgravity exposure in mouse soleus and EDL, highlighting potential new targets for improvement of countermeasures able to ameliorate or even prevent microgravity-induced atrophy in future spaceflights. C57BL/N6 mice were randomly divided in 3 groups: Bion Flown (BF), mice flown aboard the Bion M1 biosatellite in microgravity environment for 30 days; Bion Ground (BG), mice housed in the same habitat of flown animals but exposed to earth gravity; and Flight Control (FC), mice housed in a standard animal facility.
This work was supported from grants of the Department of Economics and Technology of the German Government (BMWi) through the German AeroSpace Board, Deutsches Zentrum fur Luftund Raumfahrt (DLR), e.V. Bonn, Germany (grant # 50WB821 and 1121 to DB).
The purpose of this study was to understand gene expression responses in confluent human fibroblasts in microgravity conditions to bleomycin treatment. Confluent human fibroblasts AG01522 were flown to the International Space Station (ISS). After three day on ISS, the cells were treated with bleomycin at 1.0 μg/mL for 3 hours. The cells were washed and fixed with RNALater. Total RNA were isolated after the samples were returned to Earth and subjected to microarray analysis.
This work was supported by the NASA Fundamental Space Biology Program.
Astronauts have been previously shown to exhibit decreased salivary lysozyme and increased dental calculus and gingival inflammation in response to space flight, host factors that could contribute to oral diseases such as caries and periodontitis. However, the specific physiological response of caries-causing bacteria such as Streptococcus mutans to space flight and/or ground-based simulated microgravity has not been extensively investigated. In this study, High Aspect Ratio Vessel (HARV) S. mutans simulated microgravity and normal gravity cultures were assessed for changes in metabolite and transcriptome profiles, H2O2 resistance, and competence in sucrose-containing biofilm media.
Stationary phase S. mutans simulated microgravity cultures displayed increased killing by H2O2 compared to normal gravity control cultures, but competence was not affected. RNA-seq analysis revealed that expression of 153 genes was up-regulated 2-fold and 94 genes down-regulated 2-fold during simulated microgravity HARV growth. These included a number of genes located on extrachromosomal elements, as well as genes involved in carbohydrate metabolism, translation, and stress responses. Collectively, these results suggest that growth under microgravity analog conditions promotes changes in S. mutans gene expression and physiology that may translate to an altered cariogenic potential of this organism during space flight missions. Differential gene expression was compared between RNA from S. mutans grown in normal gravity HARVs (n=3 independent cultures) and RNA from S. mutans grown in simulated microgravity HARVs (n=3 independent cultures).
This work was funded by 2012 NASA Space Biology solicitation NNH12ZTT001N.
Exposure to radiation provokes cellular responses controlled in part by gene expression networks. MicroRNAs (miRNAs) are small non-coding RNAs which mostly regulate gene expression by degrading the messages or inhibiting translation. Here, we investigated changes in miRNA expression patterns after low (0.1 Gy) and high (2.0 Gy) doses of X-ray in human fibroblasts. At early (0.5 h) and late (6 and 24 h) time points, irradiation caused qualitative and quantitative differences in the down-regulation of miRNA levels, including miR-92b, 137, 660, and 656. A transient up-regulation of miRNAs was observed after 2 h post-irradiation following high doses of radiation, including miR-558 and 662. MicroRNA levels were inversely correlated with targets from mRNA and proteomic profiling after 2.0 Gy of radiation. MicroRNAs miR-579, 608, 548-3p, and 585 are noted for targeting genes involved in radioresponsive mechanisms, such as cell cycle checkpoint and apoptosis. We suggest here a model in which miRNAs may act as hub regulators of specific cellular responses, immediately down-regulated so as to stimulate DNA repair mechanisms, followed by up-regulation involved in suppressing apoptosis for cell survival. Taken together, miRNAs may mediate signaling pathways in sequential fashion in response to radiation, and may serve as biodosimetric markers of radiation exposure. The gene expression patterns in human fibroblasts after 2.0 Gy of low-LET radiation was determined at 2 and 24 hrs post-irradiation time in technical triplicates. Control non-irradiated samples were also prepared in triplicates.
This work was funded by National Aeronautics and Space Administration, NASA Johnson Space Center, Houston, Texas. Grant Number: NNJ06HF62G.
We examined molecular responses using transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy, 1 GeV proton (1H) and 15 cGy, 1 GeV/nucleon (n) iron (56Fe) particles, 1, 3, 7, 14 and 28 days after exposure. Unsupervised clustering analysis of gene expression segregated samples according to the radiation (IR) response, and time after exposure with 56Fe-IR showing the greatest level of gene modulation. 1H-IR exposures showed little differential transcript modulation. Network analysis categorized the major differentially expressed genes into cell cycle, oxidative responses and transcriptional regulation functional groups. Transcriptional networks identified key nodes regulating expression. Individual transcription factors were inferred to be active at 1, 3, 7, 14 and 28 days after exposure. Validation of the signal transduction network by protein analysis showed that particle IR clearly regulates a long-lived signaling mechanism for p38 MAPK signaling and NFATc4 activation. Electrophoresis mobility shift assays supported the role of additional key transcription factors GATA-4, STAT-3 and NF-kB as regulators of the response at specific time points. These data suggest that the molecular response to 56Fe-IR is unique and shows long-lasting gene expression in cardiomyocytes, up to 28 days after exposure. Additionally, proteins involved in signal transduction and transcriptional activation via DNA binding play a role in the response to high charge (Z) and energy (E) particles (HZE). Our study may have implications for NASA's efforts to develop heart disease risk estimates for astronauts’ safety via identification of specific HZE-IR molecular markers and for patients receiving conventional and particle radiotherapy.
This work was supported by NASA and American Heart Association to D. A. Goukassian. This work was also supported in part by the U.S. Department of Energy (DOE) with funding from the U.S. DOE Low Dose Radiation Research Program to M. A. Coleman. The AHA and National Heart, Lung, and Blood Institute supported X. Yan. This work was also supported by National Space Biomedical Research Institute (NSBRI) to M. Natarajan.
NASA's Rodent Research (RR) project is playing a critical role in advancing biomedical research on the physiological effects of space environments. In the current set of investigations, GeneLab performed epigenomic, transcriptomic, and proteomic assays on DNA, RNA, and protein extracted from archived RR–1 tissue samples. At the end of the RR–1 mission, mice samples were collected. A variety of tissue types were harvested and either snap-frozen or preserved with RNAlater, and then stored at least a year at -80°C. Tissues available for analyses were prioritized based on the likelihood of significant scientific value in the context of either astronaut effects or previously documented rodent tissue effects. 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, kidney and liver. In addition to providing other 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.
GeneLab is proud to announce that we have included 5–methyl–cytidine (m5C)–RNAseq analysis for some of the RR–1 tissues including liver, kidney, quadriceps, gastrocnemius, and tibialis anterior. m5C residues are present in both coding and non-coding RNAs as well a the more commonly known reversible epigenetic marking of DNA on 5–methyl cytosine known to regulate transcription.
This investigation was funded by the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division with additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
Bacterial behavior has been observed to change during spaceflight. Higher final cell counts, enhanced biofilm formation, increased virulence, and reduced susceptibility to antibiotics have been reported to occur for cells cultured in space. Most of these phenomena are theorized as being an indirect effect of an altered extracellular environment, where the carbon source uptake is inhibited and excreted acidic byproducts buildup around the cell due to the lack of gravity-driven transport forces. The gene expression results from this spaceflight mission entitled Antibiotic Effectiveness in Space–1 (AES–1) corroborate this hypothesis: the data indicate an overexpression of genes associated with starvation, the search for alternative energy sources, increased metabolism, enhanced acetate production, and other systematic responses to acidity - all of which can be associated with reduced extracellular mass transport.
The data analysis effort for this investigation was funded under Grant Agreement (GA-2014-146) as part of the International Space Station (ISS) National Laboratory through the Center for the Advancement of Science in Space, Inc. (CASIS) in accordance with NASA Cooperative Agreement No. NNH11CD70A (http://www.iss-casis.org/). Luis Zea's work in Germany was funded by the German Academic Exchange Service (DAAD) and the German Aerospace Center (DLR) for their support through the DAAD Research Fellowship for Doctoral Candidates and Young Scientists (https://www.daad.org/).
Certain mouse strains such as CBA, C3H and RFM, have high incidence of radiation-induced acute myeloid leukemia (AML). The data in this series was generated by using spleen DNA from Male CBA/CaJ mice irradiated at 8–14 weeks of age with either gamma-rays or heavy ion (1 GeV/nucleon 56Fe ions) particles. Spleen DNA with radiation-induced AML was compared with DNA from normal CBA mice. Array comparative genomic hybridization were used to show that PU.1 mutations are common in both low-LET and high-LET rAML. The manuscript describing the full data highlights the similarities in molecular characteristics of high-LET and low-LET rAML and confirm the presence of ongoing chromosomal and microsatellite instability in murine rAML.
This study was funded by an NSCOR grant from the National Aeronautics and Space Administration (NAG-1569 to M. Weil); the US Department of Energy Low Dose Radiation Research Program (DE-FG02-05ER63946 to M. Weil and R. Ulrich); and the National Aeronautics and Space Administration (NNX07AP85G to J. Bedford and Y. Peng)
In prospective human exploration of outer space, the need to maintain a species over several generations under changed gravity conditions may arise. This paper reports the analysis of the third generation of fruit fly Drosophila melanogaster obtained during the 44.5-day space flight (Foton–M4 satellite, 2014, Russia), followed by the fourth generation on Earth and the fifth generation under conditions of a 12-day space flight (2014, in the Russian Segment of the ISS).
The study was supported by the program of fundamental research SSC RF–IBMP RAS, program Cell and Molecular Biology of the RAS Presidium.
Ionizing radiations are categorized by linear energy transfer (LET). High–LET are primarily found in space and have been shown to have a higher relative biological effectiveness (RBE) than low–LET radiations for many processes. This data–set provides gene-expression in yeast exposed to high–LET radiations (fast neutron, heavy ion (C) and thermal neutron) and low–LET radiation (gamma ray). Oxidative stress response is seen for all four types of radiation, but high–LET radiations induce a stronger response in these pathways. Such data are critical to evaluate the unique response of living cells exposed to a radiation environment only found in space and evaluate its RBE.
This study was supported in part by the Budget for Nuclear Research of the Ministry of Education, Culture, Sports, Science and Technology, based on screening and counseling by the Atomic Energy Commission. This work was carried out in part under the Visiting Researcher's Program of the Research Reactor Institute of Kyoto University and Research Project with Heavy Ions at NIRS-HIMAC
Accumulating data suggest that the biological responses to high and low doses of radiation are qualitatively different, necessitating the direct study of low dose responses. Most such studies have utilized 2–dimensional culture systems, which may not fully represent responses in 3–dimensional tissues. This data-set provides changes in gene expression in EPI-200, a 3–dimensional tissue model that imitates the structure and function of human epidermis, at 4, 16 and 24 hours after exposure to high (2.5 Gy) and low (0.1 Gy) doses of low LET protons. Highlight of the published analysis on these data showed that low dose was associated with gradual recovery and tissue remodeling, with HNF4A being a novel transcription factor not previously associated with radiation response, being most prominent in the low dose response.
This work was supported by the Office of Science (BER), U.S. DOE, Grant No. DE-FG02-07ER46336, and by grant number P41 EB002033-13, Radiological Research Accelerator Facility (RARAF), from the National Institutes of Health/National Institute of Biomedical Imaging and Bioengineering (NIBIB).
Carbon–ion (C–ion) radiation is used to treat tumors and humans are heterogeneous in their response to the treatment. Astronauts are exposed to whole body, chronic cosmic radiation, where protons and heavy ions are an important component and genetic differences in humans may allow some astronauts to withstand radiation better than others. There are hundreds of inbred strains of mice and strain differences have been important to the elucidation of disease mechanisms since different strains respond differently to diseases - including those caused by radiation damage. In this study, the authors performed whole lung C–ion irradiation using three different strains of mice to examine whether strain-dependent differences in radiation effects occur in high–LET C–ion thoracic irradiation. Microarray analysis was used to identify the key genes that are differentially regulated in different mouse strains after C–ion irradiation and to determine the mechanism of strain-dependent pulmonary damage after high–LET C–ion irradiation. The authors identified candidate molecules that could be implicated in the between-strain variance to early hemorrhagic pneumonitis after C–ion irradiation.
This research was supported in part by the funds for Research Project with Heavy Ions at National Institute of Radiological Sciences-Heavy Ion Medical Accelerator in Chiba (NIRS-HIMAC), Japan.
DNA sequencing provides a wealth of information about biological systems and is a pillar of modern science. If deployed to space, DNA sequencing would improve our capacity to monitor astronaut health, understand the microbial populations that inhabit spacecraft, and detect extraterrestrial life. Until recently, DNA sequencers were large and challenging to run, making their operation in space prohibitive. Nanopore DNA sequencers detect changes in conductivity as DNA bases pass through pores with diameters of ~1 nm and can be dramatically smaller and simpler to operate. This study tests the operation of the Oxford Nanopore MinION DNA sequencer on the International Space Station (ISS) and is the first demonstration of DNA sequencing in space. Multiple samples containing mixtures of DNA from Enterobacteria phage lambda, Escherichia coli and Mus musculus were sequenced with MinION sequencers on the ISS and on the ground using matched conditions. The same DNA mixtures were also sequenced with more traditional technologies (PacBio RSII, Illumina MiSeq) to allow comparison.
This work was funded in part by the NASA International Space Station program office; the NASA Postdoctoral Program; Starr Cancer Consortium grant I9-A9-071; the Irma T. Hirschl and Monique Weill-Caulier Charitable Trusts; the Bert L and N Kuggie Vallee Foundation; the WorldQuant Foundation; the Pershing Square Sohn Cancer Research Alliance; NASA grants NNX14AH50G and 15Omni2-0063; National Institutes of Health grants R25EB020393, R01ES021006 and R21AI120977; Bill and Melinda Gates Foundation grant OPP1151054; Alfred P. Sloan Foundation grant G-2015-13964; the California Initiative to Advance Precision Medicine; and Abbott Laboratories, Inc.
The microbes that colonize the surfaces of the International Space Station hitch a ride there from earth on astronauts, supplies or hardware. Regular cleaning removes many microbes, and the ones that persist may be resistant to the cleaning regimens. Understanding how the constellation of microbes on the surfaces of the International Space Station changes over time is important to reduce potential health hazards to the crew, as well as to determine how to best preserve the structural integrity of the spaceship. This study sampled heavily used surfaces of the ISS Kibo module including the incubator, air intake and handrail over three missions.
This study was supported by JAXA.
Robust microbial growth occurs in a fairly narrow range of pressures, but many bacteria can survive exposures to low pressure. This experiment explored how Bacillus subtilis respond to a low pressure of 5 kPa, such as might be encountered in the upper atmosphere at an altitude of 20 km (Mars atmosphere <1 kPa). This study will help reveal the cellular responses that are affected by exposure of B. subtilis to low pressure.
This work was funded in part by grants from the NASA Astrobiology: Exobiology and Evolutionary Biology program (NNX08AO15G); the NASA Planetary Biology Internship (PBI) program, and the NASA Earth and Space Science Fellowship (NESSF) program (13-PLANET13F-0084)
Exposure to Galactic Cosmic Radiation (GCR) presents a significant health risk to astronauts on long-duration missions beyond low Earth orbit. The researchers studied the impact of a particular form of radiation called high-mass, high-charged (HZE) particles which come in many forms. In Space, these high-energy HZE particles originate from exploding stars and are propelled through space at high speeds. HZE particles can penetrate solid objects such as the protective shielding of space vehicles, and cause significant cellular damage. In this ground-based study, human bronchial epithelial cells were exposed to HZE particles composed of either iron or silicon isotopes of varying energies, as well as to lower energy gamma rays. Cellular survival was determined and transcriptional profiling by microarray analysis was carried out at four time-points within the 24 hours following radiation exposure. This study will also aid in the assessment of HZE radiation treatment as a cancer therapy.
This work was supported by NASA NSCOR NNJ05HD36G and NNX11AC54G; Lung Cancer Spore P50 CA70907; Cancer Center Support Grant P30 CA142543
Breast cancer is one of the most common cancers associated with radiation exposure. Cosmic radiation exposure during long duration spaceflight carries uncharacterized risks. This experiment sought to understand how radiation exposure to high-energy HZE particles, that might be experienced during spaceflight beyond low earth orbit, affects mammary tumor development compared to lower energy gamma-radiation exposure commonly encountered on Earth. The researchers implanted mammary tumor cells into mice that had previously undergone exposure to radiation varying in energy and dose including densely ionizing silicon particles, lower energy gamma-radiation, or control sham-irradiation. Thus the effect of the environment on tumor progression was studied. Tumors were analyzed for global gene expression using microarray analysis and characterized histologically for molecular markers indicative of tumor stage. The study indicated that more aggressive tumors formed in animals exposed to densely ionizing particles.
This competitively selected study was funded by the NASA Space Radiation Program NSCOR award to Mary Helen Barcellos-Hoff (NASA Taskbook Grant NNX09AM52G)
Children exposed to ionizing radiation have a substantially greater breast cancer risk than adults exposed to ionizing radiation. To distinguish between several models for how exposure as a child increases the risk, this study used a mouse model and compared breast tissue in animals exposed as juveniles to ionizing radiation, gamma irradiation, or a control sham irradiation. In addition to the expression microarrays available here, the researchers developed in silico models to distinguish between various hypotheses. The researchers conclude that irradiation during puberty transiently increases stem cell self-renewal, which increases susceptibility to developing breast cancer.
This competitively selected study was funded by the NASA Space Radiation Program Specialized Center for Research in Radiation Health Effects (NSCOR) award to Mary Helen Barcellos-Hoff (NASA Taskbook Grant NNX09AM52G) and by the DOE Low-Dose Radiation program
Ionizing radiation, such as astronauts would encounter on missions beyond low-Earth orbit, is a well-established carcinogen in rodent models and a risk factor associated with human cancer. This study was designed to tease out the effects of ionizing radiation on the cells that support and influence a tumor's growth and malignancy, versus the effects on the tumor itself. The researchers implanted non-irradiated tumor tissue into mice that had been previously irradiated with ionizing radiation, lower energy gamma-rays or sham irradiated and then followed tumor development. More aggressive tumors were formed in mice exposed to ionizing irradiation prior to tumor implantation. The datasets presented here represent the expression profiles of the tumors in each of the different host environments. The results have implications in understanding tumor etiology and assessing tumor risk in individuals who have been previously exposed to ionizing radiation.
This competitively selected study was funded by the NASA Space Radiation Program Specialized Center for Research in Radiation Health Effects (NSCOR) award to Mary Helen Barcellos-Hoff (NASA Taskbook Grant NNX09AM52G) and the Department of Energy, Office of Biological and Environmental Research program on Low-Dose Radiation. Additional funding was provided from the NCI Breast SPORE program P50-CA58223, by RO1-CA138255 and RO1-CA148761, and by the Breast Cancer Research Foundation.
Most breast cancers are carcinomas that arise from the mammary epithelium. Astronauts are exposed to higher levels of ionizing radiation (IR), which is the most firmly established environmental cause of human breast cancer in both women and men. IR is also used as a cancer treatment. This study investigated the combined effect on cultured human mammary epithelial cells of exposure to IR and to the cytokine transforming growth factor beta1 (TGFbeta). TGFbeta plays a very complicated role in breast cancer; it suppresses tumor growth during the initial stage of tumorigenesis, but can switch to a tumor promoter as the stage of the tumor progresses. Gene expression was studied by transcription profiling using microarray analysis of these cells, and shows that IR causes human mammary epithelial cells to undergo an epithelial to mesenchymal transition in the presence of TGFbeta. The epithelial–mesenchymal transition (EMT) is a process by which epithelial cells lose their characteristic shape and tight connections to one another, and gain the ability to move and invade tissues: properties of mesenchymal stem cells. Carcinoma cells undergoing an EMT may invade and metastasize and thereby generate the final, life-threatening manifestations of cancer progression.
This study was supported by a National Aeronautics and Space Administration Specialized Center for Research in Radiation Health Effects (NSCOR) award to Mary Helen Barcellos-Hoff (NASA Taskbook Grant NNX09AM52G) ; the Low Dose Radiation Program of the U.S. Department of Energy Office of Biological Effects Research; U.S. Department of Defense DAMD17-00-1-0224 (A.C. Erickson); and the Office of Health and Environmental Research, Health Effects Division, U.S. Department of Energy (contract 03-76SF00098).
Ground-based simulators of microgravity are valuable tools for preparing spaceflight experiments and cost-efficient platforms for gravitational research. The effects of clinorotation on gene expression during Zebrafish development were studied after one day of clinorotation of 5-day old free-swimming larvae. The morphological effects of clinorotation on bone and cartilage structure were studied after longer exposures of 5 days clinorotation.
This work was supported by the 'Fonds de la Recherche Fondamentale Collective', the University of Liege GAME project, the European Space Agency projects, the Belgian Space Agency Prodex projects, Netherlands Organisation for Scientific (NWO) Research Earth, and Life Sciences via the Netherlands Space Office (NSO) and ESA
Medaka or killifish are small fish that inhabit rice paddies and have the distinction of having been the first vertebrate to mate in orbit on the Space Shuttle Columbia in 1994. Medaka are model organisms to study vertebrate development and space adaptation. In a multi-investigator experiment, the response to space flight was studied in six-week-old male and female Japanese medaka that were maintained for two months in the Aquatic Habitat system on the ISS. The transcriptome of six tissues (brain, eye, ovary, testis, liver and intestine) was studied using RNA-seq analysis. Histological analysis was also conducted. Though histological analysis indicated the ovary was the only tissue that showed significant spaceflight effects, all tissues showed significant changes at the level of transcription.
This research was supported by a Grant-in-Aid for Scientific Research to Hiroshi Mitani from the Japanese Ministry of Education, Culture, Sports, Science. Additional funding was from Technology of Japan and Japan Aerospace eXploration Agency (JAXA)
RNA interference (RNAi) is a biological pathway by which RNA molecules turn down or shut off the expression of genes by directly binding to complementary RNAs and activating molecular machinery to destroy the RNA. The RNAi pathway was discovered in the nematode C. elegans, but is common in all animals including humans, and is a promising therapeutic approach for combating diseases and even viral infections on Earth. This experiment used nematodes to determine whether the RNAi pathway is functional in the spaceflight environment. Crucial functional parts of the RNAi machinery, microRNAs, as well as transcripts and protein levels of the molecular components were assayed. The authors concluded that "treatment with RNAi works as effectively in the space environment as on Earth within multiple tissues, suggesting RNAi may provide an effective tool for combating spaceflight-induced pathologies aboard future long-duration space missions. Furthermore, this is the first demonstration that RNAi can be utilized to block muscle protein degradation, both on Earth and in space" indicating promise for the use of RNAi as a therapeutic approach to combat muscle degradation in space.
This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan, the Japan Society for the Promotion of Science, and Ground-Based Research Announcement for Space Utilization promoted by the Japan Space Forum. TE was supported by the Medical Research Council UK. NJS was supported by the National Institutes of Health.
Astronaut intestinal health may be impacted by microgravity, radiation, and diet. The aim of this study was to characterize how high and low linear energy transfer (LET) radiation, microgravity, and elevated dietary iron affect colon microbiota and colon function. Microbiota and mucosal characterization in these models is a first step in understanding the impact of the space environment on intestinal health. Three independent experiments were conducted. Experiment #1 compared four groups of rats exposed to adequate or high levels of dietary iron with or without low LET gamma radiation. In experiment #2, female mice were subjected to high LET particle exposure while suspended so that they were only partially weight-bearing. In experiment #3, the intestinal microbiota was assayed in mice that had flown for thirteen days on the Space Shuttle Atlantis.
This study was funded by the NASA Human Research Program's Human Health Countermeasures Element; a National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Diseases grant, and a National Space Biomedical Research Institute grant.
In the future, NASA astronauts journeying into deep space may use small devices or sensors to monitor exposure to hazardous ionizing radiation. The first step in the development of such a device is to determine how the body responds to radiation in the form of biomarkers. In this study, the researchers exposed human blood to varying doses of radiation and then used microarray analysis to analyze gene expression at varying times to determine which genes could serve as biomarkers for radiation exposure.
Funding was provided by the National Aeronautics and Space Administration (NASA) contract NNX10AJ36G; the National institutes of Health (NIH) grants P20RR016481, 3P20RR016481-09S1, and P20GM103436.
A major difficulty with biological spaceflight experiments is to differentiate the response to microgravity from the other potential confounding environmental influences including such factors as suboptimal temperature or pressure. Here the authors used a suite of ground based facilities to alter gravity to tease out the transcriptional responses of fruit flies to a suboptimal environment versus the transcriptional responses to altered gravity.
This work was supported by grants from the Spanish Space Program in the “Plan Nacional de Investigacion Cientifica y Desarrollo Tecnologico” [ESP2006-13600-C02-01] and MICINN [AYA2009-07792-E]; the Dutch NWO-ALW-SRON grant [MG-057] and the UK Engineering and Physical Sciences Research Council (EPSRC), Basic Technology grants [GR/S83005/01 and EP/G037647/1]
NASA's Microbial Payload Tracking (Microbial Observatory-1) is an ongoing census of the microbial community on the International Space Station (ISS). Sampling of the surfaces and atmosphere of the ISS over time is performed by crew members. To evaluate the potential risk to fouling of clean air supplies or contamination of fluids and food, areas that crew members contact daily are targeted for sampling including the dining area, exercise equipment, lavatory, and cupola (the best view in the house). Identifying which microbes flourish in the spaceflight and microgravity environment is important from a crew health perspective given the published findings that pathogenic bacteria become more virulent in this environment. The analysis in these six new datasets was led by Dr. Kasthuri Venkateswaran at NASA's Jet Propulsion Laboratory. More information on the Microbial Observatory-1 series can be found here.
The ISS has its own environmental microbiome shaped by microgravity, radiation, and limited human presence. To determine the microbial diversity of the ISS, environmental samples were collected from several ISS surface locations from three flight opportunities. Microbe abundance was determined by 16S ribosomal RNA gene (bacteria and archaea) and ITS (fungi) sequencing. A larger goal of this study was to determine the cultivable, total, and viable microbial diversity from the collected ISS surface samples.
To further understand how the microbial diversity of the International Space Station changes over time, researchers performed sequencing studies on environmental samples taken from eight different locations on three consecutive sampling sessions. In the study, the main objective was to identify the pool of genes for each location during each sample time to understand the functional and metabolic diversity of microorganisms in the ISS. Identification of the genes was achieved by random DNA sequencing of the pooled samples and mapping to a protein database.
Tracking of antimicrobial resistance genes is crucial to understanding the risk to for infection and illness to crew working in the closed environment of the ISS. In this study, DNA extracted from each environmental sample was used to create amplicon libraries based on a customized panel of 500 antimicrobial resistance genes followed by next-generation sequencing.
BSL-2 organisms have moderate potential hazard to humans and the environment. The classification includes various microbes that cause mild disease in humans. In this study researchers isolated and characterized bacterial strains from the ISS that showed multiple drug resistance to antibiotics. Whole genome sequencing was performed for 21 strains and is provided in this investigation. Analysis of these strains could lead to further insight of the influence of microgravity on the pathogenicity and virulence of the microorganisms..
Crew-associated environmental samples were collected from the Kibo Japanese Experiment Module (JEM), US Segment Harmony Node 2, and Russian Segment Zvezda module of the International Space Station and cultured in the laboratory of Dr. Venkateswaran at the Jet Propulsion Laboratory to isolate individual bacterial species. 16S rRNA gene sequencing identified 11 Bacillus isolates belonging to a subgroup of the Bacillus genus known as the B. anthracis/B. cereus/B. thuringiensis group. Whole genome sequence of each of the 11 isolates is provided here.
The overall analysis places these strains defines a previously uncharacterized Bacillus species, now called Bacillus issensis.
As part of the ISS Microbial Study, researchers identified two Aspergillus fumigatus strains isolated from the HEPA filter and the surface of the cupola of the ISS. Initial whole genome sequence analysis identified the isolates as A. fumigatus. This fungus can cause opportunistic infection termed aspergillosis in individuals with a compromised immune system. A long journey in space may actually compromise the immune system and make astronauts more susceptible to diseases. Researchers also conducted pathogenicity tests using the zebrafish larval model and determined that ISSF-21 is more virulent than two clinical strains (Af293 and CEA10); virulence for strain IF1SW-F4 is still being tested.
In this study, researchers present the draft genome sequences of the two strains, obtained through whole-genome sequencing.
Thousands of yeast strains from the Yeast Deletion Collection were flown on Space Shuttle flight STS-135 to the ISS in an experiment to identify genes required for survival in microgravity. Each individual strain was a mutant of the species Saccharomyces cerevisiae (brewer's yeast) that carried a deletion in a single gene. The deletion was marked by a "barcode" DNA sequence to allow identification of the surviving strains after flight. The strains were pooled together to allow competition for growth over approximately 21 generations during the spaceflight experiment. Strains missing from the mix after flight had deletions in genes that were required for survival.
This investigation was funded by the NASA Space Biology Program Office grant NNX10AP01G.
Primary cultures from mouse bone marrow were induced to differentiate by the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14-days during spaceflight. Cells were fixed to preserve RNA during flight and returned to Earth for transcriptional profiling using microarray analysis. Complementary analyses included cell proliferation studies and flow cytometry to detect antigens specific to the macrophage lineage.
This investigation was funded by the NASA Space Biology Program Office grant NNX08BA91G and also supported by the American Heart Association grant 0950036G, NIH grants AI55052, AI052206, AI088070, RR16475 and RR17686, the Jerry C. Johnson Center for Basic Cancer Research and the Kansas Agriculture Experiment Station.
During germination, a calcium current is triggered in the fern spore by a gravity sensing mechanism. The calcium current is part of a signalling cascade that orients the growth of the fern. Prior studies have determined that the critical timepoint for the gravity signal in the germination process occurs ten hours after the light signal that causes the onset of germination. In this study, transcriptional analysis was performed at ten hours post-germination to identify genes potentially important for the gravity response.
This study was supported by NASA Space Biology grants NNX09AH45G, NNX09AB41A, and NNX11AF48A to D. Marshall Porterfield and NSF grant IOS-1027514 to Stanley J. Roux
Liver is the metabolic hub of the vertebrate organ system and is involved in detoxification, regulation of glycogen storage, protein synthesis, and digestion, among other functions. Previous spaceflight experiments have demonstrated many changes in liver gene expression and in the activity of liver enzymes. GeneLab engaged in a sample sharing mission with the Rodent Research-1 (RR-1) project (GLDS-48) and with NASA's ISS National Lab managed by CASIS (GLDS-47) to provide tissue processing and extensive omics analyses on liver tissue from mice flown in microgravity. The RR-1 mission comprised the maiden voyage and validation of NASA's Rodent Research Hardware System. The RNA, protein, and DNA methylation data sets released here complement previous omics analyses from rodents in spaceflight and can be part of longitudinal studies for future rodent missions.
For the NASA investigation, samples were provided to GeneLab by the Rodent Research-1 project. The investigation was funded by the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division, and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
For the National Lab investigation, samples were provided to GeneLab by Dr. Sam Cadena (Novartis Institutes for Biomedical Research) through the Rodent Research-1 project. This investigation was funded by the Center for Advancement of Science in Space (CASIS), the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
This study provides a ground to microgravity comparative gene expression data set of female C57BL/6J mice utilizing transcriptional microarray technology. This data release in conjunction with data from previous studies in spleen and thymus using mice flown on the same mission, will allow researchers to perform network analyses that will help to gain a better understanding of the precise mechanisms that result in changes and possible health consequences associated with spaceflight.
This study was supported by the NASA Cooperative Agreement NNX10AJ31G “Cooperative Research in Proton Space Radiation", the LLUMC (Loma Linda University Medical Center) Department of Radiation Medicine and the University of Colorado Anschutz Medical Center Department of Anesthesiology. Liver samples were obtained through the NASA Biospecimen Sharing Program.
Built environments like the ISS are known to have their own microbiomes. Next-generation sequencing methods are being used to explore the ISS microbial profile to enable the development of appropriate safety and maintenance practices. This study provides strong evidence that specific human skin-associated microorganisms constitute a significant population of the ISS microbiome, generating notable differences between the ISS microbiome and cleanrooms on Earth.
This research was funded by NASA Space Biology Grant no. 19-12829-27 under Task Order NNN13D111T award to K. Venkateswaran.
This study provides a ground-to-microgravity comparative gene expression analysis of Arabidopsis thaliana seedlings. The core project examines global gene expression by RNASeq and the composition of the soluble protein fraction. The GeneLab collaboration augments the core investigation with an additional membrane protein data set. The data will allow researchers to perform network analyses to add to the knowledge of physiological effects of spaceflight during seedling growth.
This competitively selected study was funded by the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division, NASA Taskbook Grant NNX13AM48G to Sarah Wyatt and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
The core study characterizes transcriptional patterns of Arabidopsis thaliana induced during germination and growth on the International Space Station. The GeneLab collaboration allows the comparative study of three additional ecotypes. This investigation will aid researchers in assessing the common and ecotype-specific effects of spaceflight on gene expression and will facilitate cross-study data comparisons with future experiments utilizing these strains. This data release includes 48 out of 56 sample expression files with the remaining 8 files to be released at a later date.
This competitively selected study was funded by the NASA Space Biology Program Office, Space Life and Physical Sciences Research and Applications Division, NASA Taskbook Grant No. NNX13AM50G to Simon Gilroy and additional funding from the International Space Station Research Integration Office to the Space Biology GeneLab Project.
This study investigates the effects of microgravity during spaceflight on bone loss due in part to decreased bone formation by unknown mechanisms. Because it is difficult to perform experiments in space, researchers used ground-based simulators such as the Rotating Wall Vessel (RWV) and the Random Positioning Machine (RPM) to study the microgravity environment. In this study, researchers exposed 2T3 preosteoblast cells to the RWV for 3 days and found that alkaline phosphatase activity, a marker of differentiation, was inhibited. In addition, they found 61 genes downregulated and 45 genes upregulated by more than twofold compared to static 1 g controls, as shown by microarray analysis. These mechanosensitive genes may provide novel insights into understanding the mechanisms regulating bone formation and potential targets for countermeasures against decreased bone formation during spaceflight and in pathologies associated with lack of bone formation.
Using an Earth-based microgravity simulation technique that utilizes a high gradient magnetic field to levitate a biological organism, researchers investigated the biological response to weightlessness in D. melanogaster. From these experiments, researchers observed a delay in the development of the fruit flies from embryo to adult. Microarray analysis indicated significant changes in the expression of immune-, stress-, and temperature-response genes.
This study investigates the effects of microgravity on Murine Bone Marrow Stromal Cells (BMSC) that were flown to the International Space Station. The researchers use Genechip technology to detect differences in cell proliferation and cell-cycle genes between flight and control samples. This study represents the first report on the behavior of the potentially osteogenic murine BMSC in a 3D culture system.
Spores of B. subtilis 168 were exposed to real space conditions and to simulated Martian conditions for 559 days in low Earth orbit mounted on the EXPOSE-E exposure platform outside the European Columbus module on the International Space Station. Upon return, spores were germinated, total RNA extracted and fluorescently labeled and used to probe a custom Bacillus subtilis microarray to identify genes preferentially activated or repressed relative to ground control spores. Using microarray technology, this study reveals a change in expression of stress-related regulons responding to DNA damage.
Total RNA was extracted from R. rubrum S1H grown after 10 days in space flight or after 10 days in simulated ionizing radiation or simulated microgravity. Each microarray slide contained 3 technical repeats.
Researchers investigated both transcriptomic and proteomic changes in R. rubrum S1H cultures after a 10-day flight on the International Space Station and compared results to corresponding ground controls. Ground simulation of space ionizing radiation and space gravity were performed under identical culture setup and growth conditions encountered during the actual space journey. Whole-genome oligonucleotide microarray was used to test the effects of space flight. This study is unique in combining the results from an actual space experiment with the corresponding space ionizing radiation and modeled microgravity ground simulations, which allows distinguishing the different factors acting in spaceflight conditions.