Background: The radiation bystander response is an important component of the overall response of cells to radiation and critical to understanding health risks of radiation exposure to humans. The mechanism of radiation response includes inter-cellular signaling and intra-cellular communication by which the bystander signal is propagated. Methods: We measured the bystander response to 1Gy a-particle radiation in Mrad9-/- mouse stem cells and H1299shRAD9 cells using chromosomal aberration and micronucleus formation as DNA damage endpoints. In the H1299 model we used whole genome microarray analyses to profile the transcriptome of irradiated and bystander cells. Results: We investigated the role of RAD9 in the bystander response and showed that depletion or mutation of RAD9 had an effect of increasing chromosomal structural damage as well as micronucleus formation in bystander cells. The enhancement of the damage effect correlated strongly with a transcriptomic response in critical pathways. RAD9 depletion affected many pathways in the cell including the UV-MAPK pathway involving p38MAPK members STAT1 and PARP1 at the mRNA levels. There was an overall reduction of RNA biogenesis of gene members of this pathway suggesting that perhaps these signaling pathways do not function optimally after RAD9 depletion. Using network analysis we found there may be differential activation of transcriptional regulators between the irradiated and bystander cells involving the SP1 and NUPR1 transcription factors. Network analysis also suggested that HIF1a (Hypoxia induced factor 1a) activation could be a negative predictor of the bystander effect and perhaps that local hypoxic stress observed by cells that are directly exposed to radiation may predict whether or not they will elicit a bystander response. Gene expression in H1299 cells was measured at 4 hours after exposure to 1 Gy a-particles. There were two groups based on RAD9 status RAD9 normal and RAD9 depleted by siRNA. In each of these groups sham irradiated direct irradiated cells for positive bystanders positive bystanders direct irradiated cells for negative bystanders and negative bystanders; were identified based on micronucleus responses. Five biological replicates were analyzed for each experimental group.

The introduction of generally recognized as safe (GRAS) probiotic microbes into the spaceflight food system has the potential for use as a safe non-invasive daily countermeasure to crew microbiome and immune dysregulation. However the microgravity effects on the stress tolerances and genetic expression of probiotic bacteria must be determined to confirm translation of strain benefits and to identify potential for optimization of growth survival and strain selection for spaceflight. The work presented here demonstrates the translation of characteristics of a GRAS probiotic bacteria to a microgravity analog environment. Lactobacillus acidophilus ATCC 4356 was grown in the low shear modeled microgravity (LSMMG) orientation and the control orientation in the rotating wall vessel (RWV) to determine the effect of LSMMG on the growth survival through stress challenge and gene expression of the strain. No differences were observed between the LSMMG and control grown L. acidophilus suggesting that the strain will behave similarly in spaceflight and may be expected to confer Earth-based benefits.

Using a novel flood source construct we performed a direct comparison of genome-wide gene expression regulations resulting from exposure of primary human prostate fibroblast cultures to acute (10cGy and 200cGy) and longer-term chronic (1.0-2.45cGy cumulative over 24hr) exposures.

In the present study we analyzed miRNA and mRNA expression profiles in human peripheral blood lymphocytes (PBLs) incubated in microgravity condition simulated by a ground-based Rotating Wall Vessel (RWV) bioreactor. Our results show that 42 miRNAs were differentially expressed in MMG-incubated PBLs compared with 1g-incubated ones. Among these miR-9-5p miR-9-3p miR-155-5p miR-150-3p and miR-378-3p were the most dysregulated. To improve the detection of functional miRNA-mRNA pairs we performed gene expression profiles on the same samples assayed for miRNA profiling and we integrated miRNA and mRNA expression data. The functional classification of miRNA-correlated genes evidenced significant enrichments in the biological processes of immune/inflammatory response signal transduction regulation of response to stress regulation of programmed cell death and regulation of cell proliferation. We identified the correlation between miR-9-3p miR-155-5p miR-150-3p and miR-378-3p expression with that of genes involved in immune/inflammatory response (eg. IFNG and IL17F) apoptosis (eg. PDCD4 and PTEN) and cell proliferation (eg. NKX3-1 and GADD45A). Experimental assays of cell viability and apoptosis induction validated the results obtained by bioinformatics analyses demonstrating that in human PBLs the exposure to reduced gravitational force increases the frequency of apoptosis and decreases cell proliferation. microRNA expression profiling were carried out on total RNA extracted from PBLs of twelve healthy donors at the end of 24h-incubation time in MMG and in 1g conditions. Analyses were performed by using the Human miRNA Microarray kit (V2) (Agilent Technologies) that allows the detection of 723 known human (miRBase v.10.1) and 76 human viral miRNAs. By comparing the expression profile of MMG-incubated vs. 1g-incubated PBLs of the same donor we found 42 differentially expressed miRNAs 25 up-regulated and 17 down-regulated.

In the present study we analyzed miRNA and mRNA expression profiles in human peripheral blood lymphocytes (PBLs) incubated in microgravity condition simulated by a ground-based Rotating Wall Vessel (RWV) bioreactor. Our results show that 42 miRNAs were differentially expressed in MMG-incubated PBLs compared with 1g-incubated ones. Among these miR-9-5p miR-9-3p miR-155-5p miR-150-3p and miR-378-3p were the most dysregulated. To improve the detection of functional miRNA-mRNA pairs we performed gene expression profiles on the same samples assayed for miRNA profiling and we integrated miRNA and mRNA expression data. The functional classification of miRNA-correlated genes evidenced significant enrichments in the biological processes of immune/inflammatory response signal transduction regulation of response to stress regulation of programmed cell death and regulation of cell proliferation. We identified the correlation between miR-9-3p miR-155-5p miR-150-3p and miR-378-3p expression with that of genes involved in immune/inflammatory response (eg. IFNG and IL17F) apoptosis (eg. PDCD4 and PTEN) and cell proliferation (eg. NKX3-1 and GADD45A). Experimental assays of cell viability and apoptosis induction validated the results obtained by bioinformatics analyses demonstrating that in human PBLs the exposure to reduced gravitational force increases the frequency of apoptosis and decreases cell proliferation. Gene expression profiling was carried out in MMG-incubated PBLs vs. 1g-incubated PBLs on total RNA extracted from the same PBL samples assayed for miRNA profiling. We used the Whole Human Genome Oligo Microarray (Agilent) consisting of ~41.000 (60-mer) oligonucleotide probes which span conserved exons across the transcripts of the targeted full-length genes.

Worms spun at higher than normal g values for 4 days on 20G centrifuge vs. worms kept at 1 X G. Both are mixed stage cultures. A stimulus or stress experiment design type is where that tests response of an organism(s) to stress/stimulus. e.g. osmotic stress behavioral treatment Physical Characteristics: worms kept at increased g values for 4 days (x g value) stimulus_or_stress_design.

Plants exhibit a robust transcriptional response to gamma radiation which includes the induction of transcripts required for homologous recombination and the suppression of transcripts that promote cell cycle progression. Various DNA damaging agents induce different spectra of DNA damage as well as collateral damage to other cellular components and therefore are not expected to provoke identical responses by the cell. Here we study the effects of two different types of ionizing radiation (IR) treatment HZE (1 GeV Fe26+ high mass high charge and high energy relativistic particles) and gamma photons on the transcriptome of Arabidopsis thaliana seedlings. Both types of IR induce small clusters of radicals that can result in the formation of double strand breaks (DSBs) but HZE also produces linear arrays of extremely clustered damage. We performed these experiments across a range of time points (1.5-24 h after irradiation) in both wild-type plants and in mutants defective in the DSB-sensing protein kinase ATM. The two types of IR exhibit a shared double strand break-repair-related damage response although they differ slightly in the timing degree and ATM-dependence of the response. The ATM-dependent DNA metabolism-related transcripts of the DSB response were also induced by other DNA damaging agents but were not induced by conventional stresses. Both Gamma and HZE irradiation induced at 24 h post-irradiation ATM-dependent transcripts associated with a variety of conventional stresses; these were overrepresented for pathogen response rather than DNA metabolism. In contrast only HZE-irradiated plants at 1.5 h after irradiation exhibited an additional and very extensive transcriptional response shared with plants experiencing extended night. This response was not apparent in gamma-irradiated plants. We treated 5-day-old WT and atm-1 seedlings of Arabidopsis thaliana with 100 Gy of Gamma radiation (over a span of 15 minutes) or 30 Gy of HZE (over a span of approximately 12 minutes). Gamma irradiations were completed at 8:40 am while HZE irradiations were conducted in two runs (due to space limitations) which were completed at 1:09 and 1:28pm respectively. Gamma treated seedlings were sampled at 10:10 am 11:40 am 2:55 pm 8:40 pm and 8:40 am. HZE treated seedlings were sampled at 2:39 pm 4:09 pm 7:24 pm 1:09 am and 1:09 pm. Un-irradiated WT and atm-1 control seedlings were sampled at 10:45 am on Day #1 and 9:15 am on Day #2. There are a total of 22 experimental or control conditions with two replicates per condition yielding 44 samples overall.

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.

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.

One of the most likely risks astronauts on long duration space missions face is exposure to ionizing radiation associated with highly energetic and charged heavy (HZE) particles. Since access to medical expertise on such a mission is limited at best early diagnosis and mitigation of such exposure is critical. In order to accurately determine the dosage within 1 hour post-exposure dose-dependent biomarkers are needed. Therefore we performed a dose-course transcriptional analysis for radiation exposure at 0 0.3 1.5 and 3.0 Gy with corresponding time point at 1 hour (hr) post-exposure using Affymetrix GeneChip Human Gene 1.0 ST v1 Array chips. The analysis of our data suggests a set of sensitive genetic biomarkers specific to each radiation level as well as generic radiation response biomarkers. Upregulated biomarkers can then be used within lab-on-a-chip (LOC) systems to detect exposure to ionizing radiation. A total of sixteen human samples representing radiation exposure at levels 0 Gy 0.3 Gy 1.5 Gy and 3.0 Gy at time point 1 hour (hr) post-exposure were constructed. Blood samples were extracted from four human volunteers and were irradiated. Leukocytes were extracted and gene expression was measured. Samples for all four volunteers were measured at 1 hr for all four dose levels resulting in four replicates at each dose level. Thus a total of 4 samples at each of the four radiation levels were sampled yielding the total of 16 samples.