Unknown,Transcriptomics,Genomics,Proteomics

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Alterations in the Brain Transcriptome in Plasmodium Berghei ANKA Infected Mice

ABSTRACT: We have used cDNA microarrays to compare gene expression profiles in brains from normal mice to those infected with the Anka strain of Plasmodium berghei, a model of cerebral malaria. For each of three brains in each group, we computed ratios of all quantifiable genes with a composite reference sample and then computed ratios of gene expression in infected brains with untreated controls. Of the almost 12,000 unigenes adequateluy quantified in all arrays, about 3% were significantly downregulated (p<0.05, >50% fold change) and about 7% were upregulated. Upon inspection of the lists of regulated genes, we identified a high number encoding proteins of importance to normal brain function or associated with neuropathology. These results emphasize the important impact of malarial infection on gene expression in brain and provide tentative target biomarkers that might provide novel therapeutic targets for neurological sequelae of disease. We have used a previously published protocol (Iacobas et al., Physiol Genomics 2005) and a composite reference RNA sample (R) prepared in sufficient quantity for the entire experiment from ten adult mouse tissues (aorta, brain, heart, kidney, liver, lung, ovary/testicles, spleen, and stomach - equal amounts from males and females). This combination of source tissues provided a high diversity of genes expressed in the midrange of the detection system for the AECOM mouse cDNA microarrays. Briefly, 60μg total RNA, extracted in Trizol® (Invitrogen, Carlsbad, CA) from brains of three infected (I) and three control (C) mice, purified with RNeasy® mini kit (Qiagen, Valencia, CA), were reverse transcribed into cDNA incorporating fluorescent Cy3-dUTP. The composite reference was reverse transcribed to incorporate Cy5-dUTP. Each of the six Cy3-labeled brain extracts was co-hybridized overnight at 50°C against the Cy5-labeled reference with AECOM 32k Mouse oligonucleotide arrays, MO3 printing series (platform described in http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GPL5371). After hybridization, the slides were washed at room temperature, using solutions containing 0.1% sodium dodecyl sulfate (SDS) and 1% SSC (3M NaCl + 0.3M sodium citrate) to remove the non-hybridized cDNAs.

ORGANISM(S): Mus musculus

SUBMITTER: Dumitru Iacobas

PROVIDER: E-GEOD-24086 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:cDNA microarrays have been shown to be useful for monitoring global gene expression patterns in normal and disease states and in response to various environmental stimuli. In this study we have used a cattle cDNA microarray containing 7653 elements to analyze expression profiles in 19 different cattle tissues. Signal intensities from all tissue sample RNAs were compared to a reference standard RNA created from different tissues and cell lines. Data analysis identified a subset of genes significantly differentially expressed between tissues and the reference standard that were further subdivided according to fold change. Log transformed ratios were normalized using the intensity-based regional Lowess algorithm. A global error model, to account for the dependence of variation on signal intensities, was used to identify lists of genes for effect of tissue on gene expression taking into account an experiment-wise significance of 0.05, using either a Bonferroni correction (663 genes) or Benjamini and Hochbergâ??s False Discovery Rate (3350 genes). Non-supervised cluster analysis revealed groups of genes common to nerve, muscle, immune or digestive tissues. Discriminant analysis was used to support physiological functional categories and embryonic origin of tissues. Unique profiles were constructed with genes preferentially expressed in specific tissues or tissue groups in order to define gene expression for individual tissues. Global expression along a large collection of tissues revealed tissue specific expression of enzyme isomers and utilization in specific metabolic pathways. A comprehensive matrix of all possible pair-wise comparisons for individual genes among tissues was constructed to further identify genes with tissue-specific behavior and possibly unique function. A reference design was used to compare 19 cattle tissues. All tissues were compared to a universal control consisting of a mix of cattle cell lines. All samples were duplicated with a dye swap.

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Alterations in the Brain Transcriptome in Plasmodium Berghei ANKA Infected Mice

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