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Next generation sequencing of RNA expression on total splenic B cells from NOD mice fed variable diets

ABSTRACT: Purpose: The goals of this study is to examine if feeding NOD mice with diets supplemented with SCFAs causes altered gene expression in Splenic B cells Methods: B cell RNA profiles from NOD.LT mice fed fed different diets, a non-purified diet (n=4), a high amylose diet (n=4), a high amylose diet supplemented with acetate (n=4) or a high amylose diet supplemented with butyrate (n=4) were generated by deep sequencing, in triplicate, by Illumina 100 - base HT mode the sequence reads that passed quality filters were summarized into gene models using featureCounts from the Rsubread package, and differential gene expression for the various contrasts between dietary treatments was identified using the EdgeR analysis in R Results: Using an optimized data analysis workflow, we mapped about 20 million sequence reads per sample to the mouse genome (build GRCm38) and identified 16,257 transcripts present in B cells from NOD.LT mice fed different diet. Conclusions: Our study investigates how differing diets alter the expression of important genes in B cells, allowing us to further understand the role diets can play in modulating T1D development.

ORGANISM(S): Mus musculus

PROVIDER: GSE94774 | GEO | 2017/03/10

SECONDARY ACCESSION(S): PRJNA374020

REPOSITORIES: GEO

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Project description:How signals from fatty acid metabolism are translated into changes in food intake remains unclear. Previously we reported that mice with a genetic inactivation of Acads (short-chain acyl-CoA dehydrogenase), encoding the enzyme responsible for mitochondrial beta-oxidation of C4-C6 short-chain fatty acids (SCFAs), shift consumption away from fat and toward carbohydrate when offered a choice. This finding demonstrated that the loss of a specific enzyme in fatty acid oxidation alters the choice of diet intake. To our knowledge, there are no reports of studies on the effects of dietary fat on the brain transcriptome in genetic models of fatty acid oxidation deficiency. The current study aimed to identify molecular mediators underlying the effects of SCFA oxidation deficiency on food intake. The current study aimed to identify molecular mediators underlying the effects of SCFA oxidation deficiency on food intake. We performed a transcriptional screen for gene expression in brain tissue of Acads-/- and Acads+/+ mice fed either high-fat (HF) or low-fat (LF) diet for 2 d. Ingenuity Pathway Analysis revealed three top-scoring pathways significantly modified by genotype or diet: oxidative phosphorylation, mitochondrial dysfunction, and CREB signaling in neurons. A comparison of statistically significant responses in HF Acads-/- vs. HF Acads+/+ (3917) and Acads+/+ HF vs. LF Acads+/+ (3879) revealed 2551 genes or approximately 65% in common between the two experimental comparisons. All but one of these genes were expressed in opposite direction with similar magnitude, demonstrating that Acads-deficient mice fed HF diet display transcriptional responses that mimic those of wildtype Acads+/+ mice fed LF diet. Intriguingly, genes involved in energy sensing and metabolism followed this pattern. Quantitative RT-PCR in hypothalamus confirmed the dysregulation of several genes in these pathways. Western blotting showed that the combination of Acads deficiency and HF diet increased hypothalamic AMP-kinase, a key protein in an energy-sensing cascade that responds to depletion of ATP. Our results suggest that the decreased beta oxidation of short-chain fatty acids in Acads-deficient mice fed HF diet produces a state of energy deficiency in the brain and that AMP-kinase is the cellular energy-sensing mechanism linking fatty acid oxidation to feeding behavior in this model. Twelve-week old male BALB/cByJ (Acads-/-) and BALB/cByKZ (Acads+/+) mice were fed a high-fat (D12331; Research Diets) and low-fat (D12329) diet for two days. Total RNA from whole brain tissue was obtained from three mutant (Acads-/-) and three wild-type (Acads+/+) mice of each diet group. All mice had similar body weights before diets were initiated. Gene expression in brain was compared between strains and between diets. Twelve-week old male BALB/cByJ (Acads-/-) and BALB/cByKZ (Acads+/+) mice were fed a high-fat (D12331; Research Diets) and low-fat (D12329) diet for two days. Total RNA from liver was obtained from three mutant (Acads-/-) and three wild-type (Acads+/+) mice of each diet group. All mice had similar body weights before diets were initiated. Gene expression in liver was compared between strains and diets.

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Next generation sequencing of RNA expression on total splenic B cells from NOD mice fed variable diets

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