Spaceflight results in a number of adaptations to skeletal muscle, including atrophy and shifts towards faster muscle fiber types. To identify changes in gene expression that may underlie these adaptations, microarray expression analysis was performed on gastrocnemius from mice flown on the STS-108 shuttle flight (11 days, 19 hours) versus mice maintained on earth for the same period. Additionally, to identify changes that were due to unloading and reloading, microarray analyses were conducted on calf muscle from ground-based mice subjected to hindlimb suspension (12 days) and mice subjected to hindlimb suspension plus a brief period of reloading (3.5 hours) to simulate the time between landing and sacrifice of the spaceflight mice.

Physical forces greatly influence the growth and function of an organism. Altered gravity can perturb normal development and induce corresponding changes in gene expression. Understanding this relationship between the physical and biological realms is important for NASA's space travel goals. We use combined RNA-Seq and qRT-PCR to profile changes in early Drosophila melanogaster pupae exposed to chronic hypergravity (3 g, three times Earth's gravity) to highlight gravity-dependent pathways and gene products. Robust transcriptional response was evident among the pupae developed in a hypergravity environment compared to control. 1,513 genes showed significantly (p less than 0.05) altered gene expression in the 3 g samples. These findings were supported with qRT-PCR data. Major biological processes affected include ion transport, redox homeostasis, immune and humoral stress response, proteolysis, and cuticle development.

Omics analyses of RNA and protein isolated from heads of microgravity reared adult Drosophila.

Exposure to weightlessness in microgravity and elevated space radiation are associated with rapid bone loss in mammals, but questions remain about their mechanisms of action and relative importance. In this study, we tested the hypothesis that bone loss during spaceflight in Low Earth Orbit is primarily associated with site-specific microgravity unloading of weight-bearing sites in the skeleton. Microcomputed tomography and histological analyses of bones from mice space flown on ISS for 37 days in the NASA Rodent Research-1 experiment show significant site-specific cancellous and cortical bone loss occurring in the femur, but not in L2 vertebrae. The lack of bone degenerative effects in the spine in combination with same-animal paired losses in the femur suggests that space radiation levels in Low Earth Orbit or other systemic stresses are not likely to significantly contribute to the observed bone loss. Remarkably, spaceflight is also associated with accelerated progression of femoral head endochondral ossification. This suggests the microgravity environment promotes premature progression of secondary ossification during late stages of skeletal maturation at 21 weeks. Furthermore, mice housed in the NASA ISS Rodent Habitat during 1g ground controls maintained or gained bone relative to mice housed in standard vivarium cages that showed significant bone mass declines. These findings suggest that housing in the Rodent Habitat with greater topological enrichment from 3D wire-mesh surfaces may promote increased mechanical loading of weight-bearing bones and maintenance of bone mass. In summary, our results indicate that in female mice approaching skeletal maturity, mechanical unloading of weight-bearing sites is the major cause of bone loss in microgravity, while sites loaded predominantly by muscle activity, such as the spine, appear unaffected. Additionally, we identified early-onset of femoral head epiphyseal plate secondary ossification as a novel spaceflight skeletal unloading effect that may lead to premature long bone growth arrest in microgravity. This study derives results from femur and vertebrae using the micro computed tomography assay.

Transcriptional crosstalk between mammary gland liver and adipose tissue Experiment Overall Design: Pregnant and Lactating rats exposed to 3 gravity conditions

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, but 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 advancement 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, we propose to test the hypotheses that miRNA profiles will be altered in the space environment, and that cellular responses to DNA damage in space are different from those on the ground.

Microphysiological systems provide the opportunity to model accelerated changes at the human tissue level in the extreme space environment. Spaceflight-induced muscle atrophy experienced by astronauts shares similar physiological changes to muscle wasting in older adults, known as sarcopenia. These shared attributes provide a rationale for investigating molecular changes in muscle cells exposed to spaceflight that may mimic the underlying pathophysiology of sarcopenia. We report the results from three-dimensional myobundles derived from muscle biopsies from young and older adults, integrated into an autonomous CubeLab™, and flown to the International Space Station (ISS) aboard SpaceX CRS-21 as part of the NIH/NASA funded Tissue Chips in Space program. Global transcriptomic RNA-Seq analyses comparing the myobundles in space and on the ground revealed downregulation of shared transcripts related to myoblast proliferation and muscle differentiation. The analyses also revealed downregulated differentially expressed gene pathways related to muscle metabolism unique to myobundles derived from the older cohort exposed to the space environment compared to ground controls. Gene classes related to inflammatory pathways were downregulated in flight samples cultured from the younger cohort compared to ground controls. Our muscle tissue chip platform provides an approach to studying the cell autonomous effects of spaceflight on muscle cell biology that may not be appreciated on the whole organ or organism level and sets the stage for continued data collection from muscle tissue chip experimentation in microgravity. We also report on the challenges and opportunities for conducting autonomous tissue-on-chip CubeLab™ payloads on the ISS.

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.

Research conducted on the International Space Station (ISS) in low-Earth orbit (LEO) has shown the effects of microgravity on multiple organs. To investigate the effects of microgravity on the central nervous system, we developed a unique organoid strategy for modeling specific regions of the brain that are affected by neurodegenerative diseases. We generated 3-dimensional human neural organoids from induced pluripotent stem cells (iPSCs) derived from individuals affected by primary progressive multiple sclerosis (PPMS) or Parkinson's disease (PD) and non-symptomatic controls, by differentiating them toward cortical and dopaminergic fates, respectively, and combined them with isogenic microglia. The organoids were cultured for a month using a novel sealed cryovial culture method on the International Space Station (ISS) and a parallel set that remained on Earth. Live samples were returned to Earth for analysis by RNA expression and histology and were attached to culture dishes to enable neurite outgrowth. Our results show that both cortical and dopaminergic organoids cultured in LEO had lower levels of genes associated with cell proliferation and higher levels of maturation-associated genes, suggesting that the cells matured more quickly in LEO. This study is continuing with several more missions in order to understand the mechanisms underlying accelerated maturation and to investigate other neurological diseases. Our goal is to make use of the opportunity to study neural cells in LEO to better understand and treat neurodegenerative disease on Earth and to help ameliorate potentially adverse neurological effects of space travel. This study hosts data from cortical organoids. Data for the dopaminergic organoids is available under OSD-871.

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. Overall design: 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)