Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Inflammatory response of trout head kidney to in vivo challenge with IHN virus and LPS


ABSTRACT: The response of the trout, O.mykiss, head kidney to bacterial lipopolysaccharide (LPS) or active and attenuated infectious hematopoietic necrosis virus (IHNV and attINHV respectively) intraperoniteal challenge, 24 and 72 hours post-injection, was investigated using a salmonid-specific cDNA microarray. The head kidney response to i.p. LPS-induced inflammation in the first instance displays an initial stress reaction involving suppression of major cellular processes, including immune function, followed by a proliferative hematopoietic-type/biogenesis response 3 days after administration. The viral response at the early stage of infection highlights a suppression of hematopoietic and protein biosynthetic function and a stimulation of immune response. In fish infected with IHNV a loss of cellular function including signal transduction, cell cycle and transcriptional activity 72 hours after infection reflects the tissue-specific pathology of IHNV infection. attIHNV treatment on the other hand shows a similar pattern to native IHNV infection at 24 hours however at 72hours a divergence from the viral response is seen and replace with a recovery response more similar to that observed for LPS is observed. Rainbow trout, Oncorhynchus mykiss were intraperitoneally injected with saline (control) and LPS (6mg/Kg), or with 100 μl of a 106 pfu/ml dilution of IHNV or attIHNV or culture medium (negative control). At defined time points, 24 and 72 hours post i.p. injection, animals (total n=6), control and experimental, were selected from each tank (n=2) and sacrificed. Head kidneys (HK) were immediately dissected out, pooled and processed for total RNA purification using Tri Reagent (Molecular Research Center, Cincinnati, OH, USA) according to the manufacturer’s protocol.The design of the microarray includes 1380 genes printed in six replicates each. Random clones from common and subtracted cDNA libraries (976) were compared with the known vertebrate proteins using blastx and 686 genes were identified; the functional annotations were transferred from the putative homologs. These clones were supplemented with 297 genes selected by the categories of Gene Ontology. Overall, each microarray was enriched in a number of functional classes, such as stress and defense response (145 and 105 genes, respectively), cell cycle (62 genes), signal transduction (114 genes), chaperone activity (41 genes), and apoptosis (79 genes). Total RNA obtained from head kidney tissue was verified for quantity and integrity by denaturing electrophoresis and labeling with Cy3- and Cy5-dCTP (Amersham Pharmacia) was completed using SuperScript III reverse transcriptase (Invitrogen) and oligo(dT) primer, and cDNA was purified with Microcon YM30 (Millipore). We used a dye swap experimental design and each sample was hybridized to two microarrays. For the first slide, test and control cDNA were labeled with Cy5 and Cy3 respectively, and for the second array dye assignment was reversed. All head kidney samples were analyses using the dye swap protocol. Scanning was performed with ScanArray 5000 and images were processed with QuantArray (GSI Luminomics. After subtraction of mean background, locally weighted non-linear regression (Lowess) normalization was performed separately for each slide. To assess differential expression of genes, the normalized log intensity ratios were analyzed with Student’s t-test (p<0.01). The Bayesian modification to the false discovery rate (FDR) was used to correct for multiple comparison tests, estimating the q-value for the set of differentially expressed genes

ORGANISM(S): Oncorhynchus mykiss

SUBMITTER: Aleksei Krasnov 

PROVIDER: E-GEOD-10272 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Comparative analysis of the acute response of the trout, O. mykiss, head kidney to in vivo challenge with virulent and attenuated infectious hematopoietic necrosis virus and LPS-induced inflammation.

MacKenzie Simon S   Balasch Joan C JC   Novoa Beatriz B   Ribas Laia L   Roher Nerea N   Krasnov Aleksei A   Figueras Antonio A  

BMC genomics 20080326


<h4>Background</h4>The response of the trout, O. mykiss, head kidney to bacterial lipopolysaccharide (LPS) or active and attenuated infectious hematopoietic necrosis virus (IHNV and attINHV respectively) intraperitoneal challenge, 24 and 72 hours post-injection, was investigated using a salmonid-specific cDNA microarray.<h4>Results</h4>The head kidney response to i.p. LPS-induced inflammation in the first instance displays an initial stress reaction involving suppression of major cellular proces  ...[more]

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