Transcriptional profiling of mammary tissue irradiated at 10 weeks of age with either 100 cGy sparsely ionizing gamma-rays or 10 cGy or 30 cGy densely ionizing radiation (350 MeV/amu Si). Mammary tissue was collected 1 weeks 4 weeks and 12 weeks post-irradiation. Four radiation treatment groups: sham 100 cGy sparsely ionizing gamma-rays 10 cGy or 30 cGy densely ionizing radiation (350 MeV/amu Si). Three time points post-irradiation (1 4 and 12 weeks). Three or four replicates per time point.

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.

Drosophila samples were exposed to the levitation magnet inside a 25mm diameter tubes with 3 ml of yeast-based Drosophila food in the bottom and a chamber of only 5 mm of height over the food. This small region is needed in order to guarantee that all the flies were located in the effective g area so a maximum of 35 to 40 imagos or pupa can be exposed to each condition per experiment. All experiments were carried out with a parallel 1g external control in a temperature regulated incubator outside the magnet. Three experiments of different duration were performed inside the magnet system to analyze the effect of strong magnetic fields and magnetic levitation during different stages of the Drosophila development

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.

Transcriptional profiling of mammary tissue irradiated at 10 weeks of age with either 100 cGy sparsely ionizing gamma-rays or 10 cGy or 30 cGy densely ionizing radiation (350 MeV/amu Si). Mammary tissue was collected 1 weeks 4 weeks and 12 weeks post-irradiation. Four radiation treatment groups: sham 100 cGy sparsely ionizing gamma-rays 10 cGy or 30 cGy densely ionizing radiation (350 MeV/amu Si). Three time points post-irradiation (1 4 and 12 weeks). Three or four replicates per time point.

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. 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.

Drosophila samples were exposed to the levitation magnet inside a 25mm diameter tubes with 3 ml of yeast-based Drosophila food in the bottom and a chamber of only 5 mm of height over the food. This small region is needed in order to guarantee that all the flies were located in the effective g area so a maximum of 35 to 40 imagos or pupa can be exposed to each condition per experiment. All experiments were carried out with a parallel 1g external control in a temperature regulated incubator outside the magnet. Three experiments of different duration were performed inside the magnet system to analyze the effect of strong magnetic fields and magnetic levitation during different stages of the Drosophila development

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) proton (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. Transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy 1 GeV proton (1H) and 15 cGy 1 GeV/nucleon (n) proton (56Fe) particles 1 3 7 14 and 28 days after exposure.

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.

This is a genome-wide approach to identifying genes persistently induced in the mouse mammary gland by acute whole body low dose ionizing radiation (10cGy) 1 and 4 weeks after exposure. Gene expression that is modified under these parameters were compared between Tgfb1 wild type and heterozygote littermates in order to determine which genes induced or repressed by radiation were mediated via Tgfb1 status. Differential gene expression was analyzed in Tgfb1 heterozygote and wild type littermate 4th mammary glands after whole body exposure to an acute dose of 10cGy ionizing radiation. Estrus cycle was normalized in all mice two days prior to irradiation by injection with an estrogen and progesterone mixture. It is widely believed that the carcinogenic action of ionizing radiation is due to targeted DNA damage and resulting mutations but there is also substantial evidence that non-targeted radiation effects alter epithelial phenotype and the stromal microenvironment. Activation of transforming growth factor beta 1 (TGFbeta) is a non-targeted radiation effect that mediates cell fate decisions following DNA damage and regulates microenvironment composition; it could either suppress or promote cancer. Gene expression profiling shown herein demonstrates that low dose radiation (10 cGy) elicits persistent changes in Tgfb1 wild type and heterozygote murine mammary gland that are highly modulated by TGFbeta. We asked if such non-targeted radiation effects contribute to carcinogenesis by using a novel radiation chimera model. Unirradiated Trp53 null mammary epithelium was transplanted to the mammary stroma of mice previously exposed to a single low (10 -100 cGy) radiation dose. By 300 days 100% of transplants in irradiated hosts at either 10 or 100 cGy had developed Trp53 null breast carcinomas compared to 54% in unirradiated hosts. Tumor growth rate was also increased by high but not low dose host irradiation. In contrast irradiation of Tgfb1 heterozygote mice prior to transplantation failed to decrease tumor latency or increase growth rate at any dose. Host irradiation significantly reduced the latency of invasive ductal carcinoma compared to spindle cell carcinoma as well as those tumors negative for smooth muscle actin in wild type but not Tgfb1 heterozygote mice. However irradiation of either host genotype significantly increased the frequency of estrogen receptor negative tumors. These data demonstrate two concepts critical to understanding radiation risks. First non-targeted radiation effects can significantly promote the frequency and alter the features of epithelial cancer. Second radiation-induced TGFbeta activity is a key mechanism of tumor promotion. Keywords: Differential gene expression after low dose irradiation Two genotypes: TGBbeta1 heterozygote and wildtype mouse mammary glands. Two time points post-10cGy-irradiation per genotype (1 week 4 weeks); control time point was 1 week post-sham-irradiation. Two or three replicates per time point.