Spaceflight poses risks to the central nervous system (CNS), and understanding neurological responses is important for future missions. We report CNS changes in Drosophila aboard the International Space Station in response to microgravity (SFμg) and artificially simulated Earth-gravity (SF1g) via inflight centrifugation as a countermeasure. While inflight behavioral analyses of SFμg exhibit increased activity, postflight analysis displays significant climbing defects, highlighting the sensitivity of behavior to altered gravity. Multi-omics analysis shows alterations in metabolic, oxidative stress, and synaptic transmission pathways in both SFμg and SF1g; however, neurological changes immediately postflight, including neuronal loss, glial cell count alterations, oxidative damage, and apoptosis, are seen only in SFμg. Additionally, progressive neuronal loss and a glial phenotype in SF1g and SFμg brains, with pronounced phenotypes in SFμg, are seen upon acclimation to Earth conditions. Overall, our results indicate that artificial gravity partially protects the CNS from the adverse effects of spaceflight.

With technological advancements, human's desire to explore space is growing and more people are staying longer at the international space station (ISS). The impact of microgravity on stem cells (SC) is not fully understood. We explored the impact of microgravity on gene expression profile of cultured mesenchymal stem/stromal cells (MSCs) at the ISS. We also evaluated how the new knowledge gained sheds light on our understanding of human physiology on Earth. Primary cultures of MSCs were expanded at the ISS for 1 or 2 weeks and mRNA was isolated from samples of the cultured cells. Gene expression profiles were determined and compared with samples from real-time ground control cultures. Differential gene expression, gene set enrichment analysis and determination of key genes were performed that revealed for the first time the existence of potential 'master regulators' coordinating a systemic response to microgravity. Cyclin D1 (CCND1), a protein-coding gene that regulates cell cycle progression and CDK kinases, was identified as the most connected regulator at week 1. Further analysis showed the impacted genes from cultured MSCs significantly correlated with known gene pathways associated with cell division, chromosomal segregation and nuclear division, extracellular matrix structure and organization, muscle apoptosis and differentiation. This study exemplifies the utility of space research to advance our understanding of human physiology both on Earth and in space. To investigate the effects of microgravity on MSC growth and understand the differences in gene expression profiles between microgravity and ground control environments, two groups of MSC were sent to the ISS. One group was cultured for one week, while the other was cultured for two weeks, with corresponding control groups processed similarly on Earth. The cells were then preserved and transferred back to the laboratory. Further Gene expression profiles were compared between samples to identify differentially expressed genes.

The objective of the Rodent Research-9 (RR-9) mission was to use mice to understand the molecular basis of phenomena that affect astronauts during long-duration spaceflight particularly visual impairment and joint tissue degradation. To this end a flight group (FLT) of 10-week-old male C57BL/6J mice was launched from Kennedy Space Center (KSC) on 8/14/2017 and housed in Rodent Habitats on the ISS for 33 days before being returned alive to Earth. After splashdown in the Pacific Ocean the animals were transported to Loma Linda University (LLU) for testing euthanasia and dissection on 9/18/2018. A Basal Control (BSL) was housed in standard cages at Kennedy Space Center (KSC) and euthanized one day after launch of the FLT animals (8/15/2017). Ground Control (GC) and Vivarium Control (VIV) studies were planned to commence at KSC approximately one-week after the conclusion of the flight experiments. However all the GC and VIV mouse studies at KSC had to be cancelled due to Hurricane Irma and potential adverse effects on the animal housing facility. The GC and VIV studies were therefore rescheduled and begun in May 2018. The GC was euthanized and dissected 6/18/2018 - 6/20/2018 while the VIV was euthanized and dissected 6/22/2018 - 6/23/2018. Because this resulted in a different cohort of mice being used for the GC and VIV controls as compared to the flight (FLT) and basal (BSL) groups two cohort controls were included in the study. The first Cohort Control 1 (CC_C1) was from the same cohort as the FLT and BSL animals and was sacrificed and dissected 4 days after the FLT group (9/22/2017). The second Cohort Control 2 (CC_C2) was from the same cohort as the GC and VIV animals and was sacrificed and dissected 2-8 days after the GC and VIV groups (6/24/2018 - 6/26/2018). The CC_C1 and CC_C2 groups were housed in standard cages and fed standard chow in contrast to all other groups which received Rodent Foodbars. To clarify the connections between treatment groups and animal cohorts the following group abbreviations are used in the sample metadata: Flight (FLT_C1); Basal (BSL_C1); Ground Control (GC_C2); Vivarium Control (VIV_C2) Cohort Control 1 (CC_C1); Cohort Control 2 (CC_C2). Fecal pellets were isolated directly from mice during dissection and preserved by flash freezing in liquid nitrogen before stored at -80 C. DNA was then extracted shotgun metagenomic libraries generated and libraries sequenced (target 10 M clusters at PE 250 bp). Metagenomic data was generated from the following groups: Basal Control (n=5) Ground Control (n=5) Vivarium Control (n=5) Cohort Control 1 (n=5) Cohort Control 2 (n=5) Flight (n=5).

The microgravity environment aboard the International Space Station (ISS) provides a unique stressor that can help understand underlying cellular and molecular drivers of pathological changes observed in astronauts with the ultimate goals of developing strategies to enable long- term spaceflight and better treatment of diseases on Earth. We used this unique environment to evaluate the effects of microgravity on kidney proximal tubule epithelial cell (PTEC) response to serum exposure and vitaminD biotransformation capacity. To test if microgravity alters the pathologic response of the proximal tubule to serum exposure, we treated PTECs cultured in a microphysiological system (PT-MPS) with human serum and measured biomarkers of toxicity and inflammation (KIM-1 and IL-6) and conducted global transcriptomics via RNAseq on cells undergoing flight (microgravity) and respective controls(ground). Given the profound bone loss observed in microgravity and PTECs produce the active form of vitamin D, we treated 3D cultured PTECs with 25(OH)D 3 (vitamin D) and monitored vitamin D metabolite formation, conducted global transcriptomics via RNAseq, and evaluated transcript expression of CYP27B1, CYP24A1, or CYP3A5 in PTECs undergoing flight (microgravity) and respective ground controls. We demonstrated that microgravity neither altered PTEC metabolism of vitamin D nor did it induce a unique response of PTECs to human serum, suggesting that these fundamental biochemical pathways in the kidney proximal tubule are not significantly altered by short-term exposure to microgravity. Given the prospect of extended spaceflight, more study is needed to determine if these responses are consistent with extended (greater than 6 months) exposure to microgravity.

We report the findings of an animal experiment onboard STS-135 investigating the molecular aspects of the impact of spaceflight on retinal biology by performing differential gene expression profiling between mice flown onboard STS-135 and their ground control counterparts.

While it has been shown that astronauts suffer immune disorders after spaceflight, the underlying causes are still poorly understood and there are many variables to consider when investigating the immune system in a complex environment. Additionally, there is growing evidence that suggests that not only is the immune system being altered, but the pathogens that infect the host are significantly influenced by spaceflight and ground-based spaceflight conditions. In this study, we demonstrate that Serratia marcescens (strain Db11) was significantly more lethal to Drosophila melanogaster after growth on the International Space Station than ground-based controls, but that the host immune system is not significantly altered amongst known immune genes. High-throughput sequencing of wild-type (w1118) adult hosts infected with either space or ground-reared S. marcescens revealed few changes in gene expression, with 11 genes significantly differentially expressed (q-values less than 0.05) and only one gene related to the immune system. This data supports the main findings of the paper, which state that both spaceflight and low-shear modeled microgravity conditions increase the virulence of this pathogen, independent of the host immune system. This data, which shows that there are no significant immune-related changes to the host when infected with space-grown sample compared to ground-grown sample, provides further evidence that there are likely phenotypic changes to the pathogen itself that is causing increased virulence in spaceflight and in low-shear modeled microgravity. RNA was extracted in triplicate from 2 pooled adult (2-3 day old female) Drosophila melanogaster (w1118) per treatment, with 4 total treatment groups (no injection control, sham injection with PBS, ground bacteria-injected, and space bacteria-injected) with poly(A)+ RNA libraries. Samples were multiplexed and sequenced 100bp paired-end ready were sequenced on one lane of the Illumina HiSeq-4000.

Bone loss and immune dysregulation are among the main adverse outcomes of spaceflight challenging astronaut xe2 x80 x99s health and safety. However consequences on B cell development and responses are still under-investigated. Up to now most studies addressing these questions were performed using an amphibian species. Consequently we used advanced proteomics analysis of femur bone and marrow of mice flown for one month on board the BION-M1 biosatellite to determine whether extreme conditions encountered during a real spaceflight affect B cell development in mice and to examine reversibility of the effects upon return to Earth. Our data revealed that adverse effects on B lymphopoiesis were more marked one week after landing and that this phenomenon was associated with a 41% reduction of B cells in the spleen. Thus the effects of spaceflight persisted during at least one week after landing. These reductions may contribute to explain increased susceptibility to infection even if we confirmed that animals were able to mount a humoral immune response.

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 and matching 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. Brain samples from three flight and three ground control animal groups were cut in half between hemispheres. One hemisphere of each brain was used for generating spatially resolved transcriptional profiling data. Hemispheres were cryosectioned so that 2 consecutive sections from the hippocampus of each brain was placed on Visium Gene Expression arrays. Samples were fixed stained with Hematoxylin and Eosin and imaged. Imaging was followed by tissue permeabilization to release mRNA molecules from cells for capture onto the array surface. Subsequently following the 10XGenomics Visium Gene Expression protocol Spatial Transcriptomics RNA-seq libraries were prepared and sequenced.

We report the findings of an animal experiment onboard STS-135 investigating the molecular aspects of the impact of spaceflight on retinal biology by performing differential gene expression profiling between mice flown onboard STS-135 and their ground control counterparts.

The JAXA MHU-2 mission had two objectives: 1) To increase understanding of effects of spaceflight on the gut environment (microbiota and metabolites) and immune system using multi-omics based analysis; 2) To evaluate whether fructo-oligosaccharides added to the diet as prebiotics improve the gut environment and immune function during spaceflight. Twelve 16-18 week old male C57BL/6J mice were singly housed in the JAXA Habitat Cage Units (HCUs) on the ISS for 30 days. Six flight mice were housed in microgravity while six were exposed to simulated 1g by centrifugation. These two flight groups were further divided in half so that three mice in each group received standard JAXA chow while the other three were fed chow supplemented with fructooligosaccharides (FOS). Mice were returned live and euthanized and dissected <1 day after splashdown. Ground controls (n=6) were asynchronous and housed in HCUs. Vivarium controls (n=6) were asynchronous and housed in standard habitats. Three ground control and three vivarium animals received standard chow while the other three each ground control and vivarium animals received FOS-supplemented chow. Ground and vivarium samples were dissected by a separate dissection team than flight samples. Dorsal skin was dissected 30 minutes after euthanasia and snap frozen in liquid nitrogen. Total RNA was extracted and sequenced at a target depth of 60 M clusters per sample (ribodepleted paired end 150). Study Factor Levels: 1)Spaceflight ug Std. Chow: 3; 2)Spaceflight ug FOS: 3; 3) Spaceflight Artificial 1g Std. Chow: 3; 4)Spaceflight Artificial 1g FOS: 3; 5)Ground 1g Std. Chow: 3; 6)Ground 1g FOS: 3; 7)Vivarium 1g Std. Chow: 3; 8)Vivarium 1g FOS: 3.