Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptomic analyis of the response of porcine PBMC to stimulation with concanavalin A

ABSTRACT: One of the top priorities of the swine industry worldwide is to decrease the susceptibility of its animals to infectious diseases. There is therefore now an increased focus on the production of more M-bM-^@M-^\robustM-bM-^@M-^] animals, which have lower susceptibility to a broad range of infectious diseases. One general measure of immune competence is the proliferative response of peripheral blood mononuclear cells (PBMC) to mitogenic stimulation. Concanavalin A (ConA) is a mitogen that predominantly stimulates T lymphocytes within the PBMC fraction of blood. T cells play a crucial role in both cell-mediated and humoral immunity. Variation between individuals in the magnitude of the proliferative response to ConA has been estimated to have moderate heritability, and quantitative trait loci (QTL) for this trait have been identified. However, detailed knowledge of the genomic pathways which coordinate this process in swine PBMC is currently lacking. The recent development of a high density oligonucleotide microarray for the pig research community means that it is now possible to study simultaneously the expression of a large proportion of genes from the pig genome. The objective of this research project was to profile gene expression in ConA-stimulated PBMC to identify the gene networks that control the cellular response to this mitogen. Peripheral Blood Mononuclear Cells (PBMC) from 4 individual biological replicates were used for this experiment. Separate cell aliquots from these animals were grown in culture with the mitogen concanavalin A for 4 different times: 0, 3, 20, and 68 hours. For the hybridization experiment, 3 separate direct comparisons of gene expression were made (0 v 3 hr, 0 v 20 hr and 0 vs 68 hr). Time-point samples from the same individual were always compared on the same array. Each comparison consisted of 8 hybridizations (4 biological replicates; 2 dye-swap technical replicates), making a total of 24 hybridizations.

ORGANISM(S): Sus scrofa

SUBMITTER: Jamie Wilkinson

PROVIDER: E-GEOD-33760 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:This study tested the hypothesis that transcription of immediate early genes is inhibited in T cells activated in microgravity (mg). Immunosuppression during spaceflight is a major barrier to safe long-term human space habitation and travel. The goals of these experiments were to prove that mg was the cause of impaired T cell activation during spaceflight as well as understand the mechanisms controlling early T cell activation. T cells from 4 human donors were stimulated with concanavalin A (ConA) and anti-CD28 onboard the International Space Station (ISS). An onboard centrifuge was used to generate a 1g simultaneous control to isolate the effects of mg from other variables of spaceflight. Microarray expression analysis after 1.5 hours of activation demonstrated that mg- and 1g-activated T cells had distinct patterns of global gene expression and identified 47 genes that were significantly differentially down-regulated in mg. Importantly, several key immediate early genes were inhibited in mg. T cells were isolated from human volunteers. T cells from each donor were kept separate and loaded into individual chambers in separate cassettes for the following treatments: mg non-activated, mg activated, and 1g activated. Therefore, samples represent biological triplicates. Experimental units were launched into space and placed into the KUBIK facility onboard the International Space Station. The 1g units were placed in the central centrifuge positions and centrifuged with an applied 1g force. The mg units were place in the static positions for continued mg exposure. After 30 minutes of pre-incubation, mg non-activated units were fixed by addition of RNALater (QIAGEN, Valencia, CA), removed from the incubator, and stored in 4M-BM-0C. The mg and 1g activated units were injected with final concentration 10mg/ml Con A and 4mg/ml anti-CD28. These cassettes were replaced into KUBIK on either the centrifuge or static positions and activated for 1.5 hours. Activation was stopped with the addition of RNALater and the units were then moved to 4M-BM-0C storage. All units were returned to Earth for analysis.

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Transcriptomic analyis of the response of porcine PBMC to stimulation with concanavalin A

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