We differentiated mouse bone marrow cells in the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14 days during the flight of space shuttle Space Transportation System (STS)-126. We tested the hypothesis that the receptor expression for M-CSF c-Fms was reduced. We used flow cytometry to assess molecules on cells that were preserved during flight to define the differentiation state of the developing bone marrow macrophages; including CD11b CD31 CD44 Ly6C Ly6G F4/80 Mac2 c-Fos as well as c-Fms. In addition RNA was preserved during the flight and was used to perform a gene microarray. We found that there were significant differences in the number of macrophages that developed in space compared to controls maintained on Earth. We found that there were significant changes in the distribution of cells that expressed CD11b CD31 F4/80 Mac2 Ly6C and c-Fos. However there were no changes in c-Fms expression and no consistent pattern of advanced or retarded differentiation during space flight. We also found a pattern of transcript levels that would be consistent with a relatively normal differentiation outcome but increased proliferation by the bone marrow macrophages that were assayed after 14 days of space flight. There also was a surprising pattern of space flight influence on genes of the coagulation pathway. These data confirm that a space flight can have an impact on the in vitro development of macrophages from mouse bone marrow cells.

We differentiated mouse bone marrow cells in the presence of recombinant macrophage colony stimulating (rM-CSF) factor for 14 days during the flight of space shuttle Space Transportation System (STS)-126. We tested the hypothesis that the receptor expression for M-CSF, c-Fms was reduced. We used flow cytometry to assess molecules on cells that were preserved during flight to define the differentiation state of the developing bone marrow macrophages; including CD11b, CD31, CD44, Ly6C, Ly6G, F4/80, Mac2, c-Fos as well as c-Fms. In addition, RNA was preserved during the flight and was used to perform a gene microarray. We found that there were significant differences in the number of macrophages that developed in space compared to controls maintained on Earth. We found that there were significant changes in the distribution of cells that expressed CD11b, CD31, F4/80, Mac2, Ly6C and c-Fos. However, there were no changes in c-Fms expression and no consistent pattern of advanced or retarded differentiation during space flight. We also found a pattern of transcript levels that would be consistent with a relatively normal differentiation outcome but increased proliferation by the bone marrow macrophages that were assayed after 14 days of space flight. There also was a surprising pattern of space flight influence on genes of the coagulation pathway. These data confirm that a space flight can have an impact on the in vitro development of macrophages from mouse bone marrow cells.

Microarray Analysis of Space-flown Murine Thymus Tissue Reveals Changes in Gene Expression Regulating Stress and Glucocorticoid Receptors. We used microarrays to detail the gene expression of space-flown thymic tissue and identified distinct classes of up-regulated genes during this process. We report here microarray gene expression analysis in young adult C57BL/6NTac mice at 8 weeks of age after exposure to spaceflight aboard the space shuttle (STS-118) for a period of 13 days. Upon conclusion of the mission thymus lobes were extracted from space flown mice (FLT) as well as age- and sex-matched ground control mice similarly housed in animal enclosure modules (AEM). mRNA was extracted and an automated array analysis for gene expression was performed. Examination of the microarray data revealed 970 individual probes that had a 1.5 fold or greater change. When these data were averaged (n=4) we identified 12 genes that were significantly up- or down-regulated by at least 1.5 fold after spaceflight (p < 0.05). Together these data demonstrate that spaceflight induces significant changes in the thymic mRNA expression of genes that regulate stress glucocorticoid receptor metabolism and T cell signaling activity. These data explain in part the reported systemic compromise of the immune system after exposure to the microgravity of space.

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.

Gene expression levels were determined in 3rd instar and adult Drosophila melanogaster reared during spaceflight to elucidate the genetic and molecular mechanisms underpinning the effects of microgravity on the immune system. The goal was to validate the Drosophila model for understanding alterations of innate immune responses in humans due to spaceflight. Five containers of flies with ten female and five male fruit flies in each container were housed and bred on the space shuttle (average orbit altitude of 330.35 km) for 12 days and 18.5 hours with a new generation reared in microgravity. RNA was extracted on the day of shuttle landing from whole body animals (3rd instar larvae and adults) hybridized to Drosophila 2.0 Affymetrix genome arrays and the expression level of all genes was normalized against the gene expression level from the corresponding developmental stage animals raised on ground. Spaceflight altered the expression of larval genes involved in the maturation of plasmatocytes (macrophages) and their phagocytic response as well as the level of constitutive expression of pattern recognition receptors and opsonins that specifically recognize bacteria and of lysozymes antimicrobial peptide pathway and immune stress genes hallmarks of humoral immunity. Larval microarrays (FL 6 samples) are based on RNA extracted from 6 independent sets of 50 mid 3rd instar larvae reared in microgravity and collected on the day of landing after 12 days and 18.5 hours on the space shuttle and the same number of control larvae raised on ground (GL 6 samples). Adults microarrays (F1 3 samples) are based on RNA from 3 sets of 20 adult females each that emerged during spaceflight and within 4 hours of landing and the same number of adult females from the corresponding ground control containers (G1 3 samples).

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.

Space radiations and microgravity both could cause DNA damage in cells but the effects of microgravity on DNA damage response to space radiations are still controversial. A mRNA microarray and microRNA microarray in dauer larvae of Caenorhabditis elegans (C. elegans) that endured spaceflight environment and space radiations environment during 16.5-day Shenzhou-8 space mission were performed. In our study wild type and ced-1 mutant strains of C.elegans endured three conditions during shenzhou-8 spaceflight mission including spaceflight static condition(ss) spaceflight 1-g centrifugal condition(sc) and ground control condition(gc). Limited to the quantity of worm samples we performed technical-repeat test but not sample-repeat test.Accordingly xef xbc x8csix miRNA microarrays were performed.

Space radiations and microgravity both could cause DNA damage in cells but the effects of microgravity on DNA damage response to space radiations are still controversial. A mRNA microarray and microRNA microarray in dauer larvae of Caenorhabditis elegans (C. elegans) that endured spaceflight environment and space radiations environment during 16.5-day Shenzhou-8 space mission were performed. In our study wild type and dys-1 mutant strains of C.elegans endured four conditions during shenzhou-8 spaceflight mission including spaceflight static condition(ss) spaceflight 1-g centrifugal condition(sc) ground control condition(gc) and no-transport control. Limited to the quantity of worm samples we performed technical-repeat test but not sample-repeat test. Accordingly eight miRNA microarrays were performed.

Space radiations and microgravity both could cause DNA damage in cells but the effects of microgravity on DNA damage response to space radiations are still controversial. A mRNA microarray and microRNA microarray in dauer larvae of Caenorhabditis elegans (C. elegans) that endured space xef xac x82ight environment and space radiations environment during 16.5-day Shenzhou-8 space mission were performed. In our study wild type and dys-1 mutant strains of C.elegans endured four conditions during shenzhou-8 spaceflight mission including spaceflight static condition(ss) spaceflight 1-g centrifugal condition(sc) ground control condition(gc) and no-transport control. Limited to the quantity of worm samples we performed technical-repeat test but not sample-repeat test. Accordingly eight miRNA microarrays were performed.

Microgravity alters the immune response to in vitro LPS assault engineered in spaceflight: A multi-omics study Microgravity can facilitate creation of a potent environment for opportunistic infection by augmenting virulence and suppressing the host defense. Presumably extraterrestrial infection may trigger potentially novel bionetworks different from the terrestrial equivalent which could only be probed by investigating the host-pathogen relationship with minimum terrestrial bias. Towards this objective we strategically engineered a cell culture module equipped with a feedback controlled semi-automated platform to expose human endothelial cells to lipopolysaccharide (LPS). The assay was carried out in the STS-135 space shuttle and a concurrent ground study constituted the baseline. Transcriptomic investigation revealed an immune blunting in microgravity; Lbp MyD88 and MD-2 failed to encode proteins responsible for early LPS uptake. Longer exposure results implied that there was a delayed response potentially ineffectual in preventing pathogens from opportunistically modulating the infection network. Lack of recruitment of growth factors and a debilitated apoptosome supported this potential explanation. Certain cytokines such as IL-6 and IL-8 surged in response to LPS insult in microgravity. Contrasting expressions of B2M TIMP-1 and VEGRs suggested impaired pro-survival adaptation and healing mechanisms. The susceptibility of oxidative stress and immune regulation to microgravity compelled further investigation of the respective microRNA modulators such as miR-200a and miR-146b. These miRNAs were expressed differently in response to LPS assaults in different gravitational limits. In conclusion despite a serious drawback attributed to the small sample size we delineated some of the important aspects of the extraterrestrial etiology; more comprehensive follow up studies are warranted. Present study though compromised by the small sample size was able to shade lights on several aspects of immunological responses to the endotoxic assault mediated by uG. Implementing the host-pathogen interactions in the spaceflight and subsequently lysing the cells onboard presented the critical distinguishing features of the present study from the past reports. We identified the CCM of Tissue Genesis Inc. HI as the suitable hardware system to carry out the experiment in the spaceflight. CCM is an automated feedback controlled module that can concurrently support 24 bioreactors following protocols exclusively programmed for individual bioreactor. For this experiment we use samples EA41 EA 47 EA45 and EA155 that were exposed to LPS for 4 hours. Samples EA123 EA165 EA127 EA126 were exposed to LPS for 8Hrs. Samples EA33 EA 125 EA79 and EA 39 were controls in this experiment.