Unknown,Transcriptomics,Genomics,Proteomics

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Effect of blood sample storage on the study of gene expression in response to ionizing radiation exposure

ABSTRACT: For around ten years, microarrays have been suggested for the diagnosis of ionizing radiation exposure. We assessed for the first time the relevance of gene expression profiling in a real accidental case. This study was performed on peripheral blood mononuclear cells of 41 potential victims. The different strategies of analysis highlighted a huge effect of the blood sample handling on the gene expression profiles. This effect was so high that it could mask specific modulations as a potential effect of ionizing radiation exposure. Thus, we assessed a new way of blood sampling adapted to gene expression analysis: PAXgene. With this method, more than 70% of the modulations of gene expression induced 3 hours after an ex vivo exposure to 0.5 Gy were preserved even in a 24-hour delayed analysis (as for transportation of blood sample from the accident location to the laboratory). We validated a new methodology in order to propose a new strategy of blood sampling and handling for gene profiling. This system could be used in case of accidental overexposure to study whether gene expression is a relevant biomarker of ionizing radiation exposure. Radiation induced gene expression in human blood was measured at 3 hours after exposure to doses of 0 and 0.5 grays. Following the incubation of 3 hours at 37Â°C, the RNA extractions were performed either immediately or 24h later (as for transportation of blood sample at room temperature). Two different blood preservation methods were compared: classical anticoagulant and PAXgene Blood RNA System. Venous blood samples of 6 donors were used (3 for anticoagulant study, 3 for Paxgene study). Each sample was hybridized twice.

ORGANISM(S): Homo sapiens

SUBMITTER: Gaetan GRUEL

PROVIDER: E-GEOD-21240 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:The Biomarkers of Exposure to ARsenic (BEAR) pregnancy cohort in Gómez Palacio, Mexico was recently established to better understand the impacts of prenatal exposure to inorganic arsenic (iAs). In this study, we examined a subset (n = 40) of newborn cord blood samples for microRNA (miRNA) expression changes associated with in utero arsenic exposure. Levels of iAs in maternal drinking water (DW-iAs) and maternal urine were assessed. Levels of DW-iAs ranged from below detectable values to 236 µg/L (mean = 51.7 µg/L). Total arsenic in maternal urine (U-tAs) was defined as the sum of iAs and its monomethylated and dimethylated metabolites (MMAs and DMAs, respectively) and ranged from 6.2 to 319.7 µg/L (mean = 64.5 µg/L). Genome-wide miRNA expression analysis of cord blood revealed 12 miRNAs with increasing expression associated with U-tAs. Transcriptional targets of the miRNAs were computationally predicted and subsequently assessed using transcriptional profiling. Pathway analysis demonstrated that the U-tAs-associated miRNAs are involved in signaling pathways related to known health outcomes of iAs exposure including cancer and diabetes mellitus. Immune response-related mRNAs were also identified with decreased expression levels associated with U-tAs, and predicted to be mediated in part by the arsenic-responsive miRNAs. Results of this study highlight miRNAs as novel responders to prenatal arsenic exposure that may contribute to associated immune response perturbations. We assessed the impact of prenatal exposure to arsenic on genome-wide miRNA expression profiles and their potential influence on gene expression patterns in the Biomarkers of Exposure to ARsenic (BEAR) prospective pregnancy cohort. This cohort includes residents from Gómez Palacio, located in the state of Durango in the Lagunera region of Northern Mexico. A total of 200 pregnant women residing in Gómez Palacio, State of Durango, Mexico, were recruited at the General Hospital of Gómez Palacio to participate in the BEAR prospective pregnancy cohort. The present study focuses on miRNA expression profiles and utilizes 40 samples obtained from mother-newborn pairs selected from the larger cohort (n=200). The subcohort was selected to include subjects exposed to varying levels of arsenic as determined by both total arsenic in maternal urine (U-tAs) and inorganic arsenic in drinking water (DW-iAs). Cord blood samples were collected from the newborns immediately after infant delivery. Blood samples were collected using PreAnalytix PaxGene RNA tubes and extracted using the PAXgene RNA Kit, per standard protocol (Qiagen, Valencia, CA). Isolated RNA used for microarray analysis were amplified and labeled using the NuGEN Ovation Pico WTA System V2 and Encore Biotin Module, respectively (NuGEN, San Carlos, CA). RNA isolated from 40 cord blood samples were labeled and hybridized to the Agilent Human miRNA Microarray, based off miRBase v16.0.

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Effect of blood sample storage on the study of gene expression in response to ionizing radiation exposure

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