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Transcriptomic profiling of LPS-induced adrenal inflammation in WT mice

ABSTRACT: Although of the utmost importance for our survival, the activation of the adrenal gland is often impaired during sepsis. Little is known about the mechanisms involved in this process. In the present study, we have used a robust RNA Sequencing (RNA-Seq) technology in a search for novel transcriptomic signatures in adrenal glands triggered by bacterial lipopolysaccharide-induced systemic inflammation. Our results demonstrate that LPS injection to mice induce a robust change in the adrenal transcriptome, significantly altering the expression of 8458 genes as compared to saline injected animals. The principal component analysis shows a clear separation between LPS and saline treatments, as well as show low variance between biological samples from each group. The functional analysis of 4312 upregulated and 4146 downregulated genes by LPS, demonstrate a strong induction of genes involved in inflammatory and steroidogenic responses, and inhibition of genes involved in DNA repair and stem/progenitor cell pathways. Gene enrichment set analysis (GSEA) further validated the functional data, and indicated a strong grouping of genes involved in LPS-mediated induction of interferon and innate immune responses, along with previously unexplored activation of the hypoxia and inhibition of Wnt/Shh pathways in the adrenal gland. These results were further validated by quantitative PCR analysis. Collectively, this is the first study demonstrating a global alteration of adrenal gene signatures during systemic inflammation, indicating the potential pathways that are involved in the adrenal gland dysregulation during sepsis.

ORGANISM(S): Mus musculus

PROVIDER: GSE121051 | GEO | 2019/06/21

REPOSITORIES: GEO

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Project description:Background: Our understanding of the role host genetic factors play in the initiation and severity of infections caused by Gram negative bacteria is incomplete. Methods: To identify novel regulators of the host response to lipopolysaccharide (LPS), 11 inbred murine strains were challenged with LPS systemically. RNA from lung, liver, and spleen tissue was profiled on oligonucleotide microarrays to determine if unique transcripts differentiate susceptible and resistant strains of mice. Gene expression data were analyzed to identify over-represented pathways and transcription factors (TFs). The role of several TFs in innate immune response to LPS was examined by RNA interference in a mouse macrophage cell line. Mouse lines with targeted mutations were utilized to further confirm involvement of two novel genes in innate immunity. Results: In addition to two strains lacking functional TLR4 (C3H/HeJ and C57BL/6JTLR4-/-), three murine strains with functional TLR4 (C57BL/6, 129/SvlmJ, and NZW/LacJ) were found to be resistant to systemic LPS challenge; the other six strains were classified as sensitive. Gene expression analysis supports the involvement of a number of previously identified genes, molecular pathways, and TFs in the host response to LPS but also identifies Hedgehog signaling as a novel pathway activated by LPS. B6;129Ptch1+/+ wild-type mice were shown to be more sensitive to systemic LPS than B6;129Ptch1+/- heterozygote littermates further supporting the role of Hedgehog signaling in systemic LPS response. RNA interference-mediated inhibition of 6 TFs (C/EBP, Cdx-2, E2F1, Hoxa4, Nhlh1, and Tead2), out of 15 tested, was found to diminish production of IL-6 and TNF protein in murine macrophages. The role of E2F1 was confirmed by showing that B6;129E2F1-/- knockout mice are more sensitive to systemic LPS than wild-type controls. Conclusions: Our analysis of gene expression data identified novel pathways and transcription factors that regulate the host response to systemic LPS. Our results provide potential sepsis biomarkers and therapeutic targets that should be further investigated in human populations. Keywords: disease state analysis For each strain of mice, 6 mice were treated with LPS and 6 with saline. RNA of 3 animals was pooled to make two pools per condition (two biological replicates). Each pool was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Univeral Mouse Reference (dye flip techical replicates). RNA from lung, liver, and spleen tissue was profiled on oligonucleotide microarrays

Project description:Background: Our understanding of the role host genetic factors play in the initiation and severity of infections caused by Gram negative bacteria is incomplete. Methods: To identify novel regulators of the host response to lipopolysaccharide (LPS), 11 inbred murine strains were challenged with LPS systemically. RNA from lung, liver, and spleen tissue was profiled on oligonucleotide microarrays to determine if unique transcripts differentiate susceptible and resistant strains of mice. Gene expression data were analyzed to identify over-represented pathways and transcription factors (TFs). The role of several TFs in innate immune response to LPS was examined by RNA interference in a mouse macrophage cell line. Mouse lines with targeted mutations were utilized to further confirm involvement of two novel genes in innate immunity. Results: In addition to two strains lacking functional TLR4 (C3H/HeJ and C57BL/6JTLR4-/-), three murine strains with functional TLR4 (C57BL/6, 129/SvlmJ, and NZW/LacJ) were found to be resistant to systemic LPS challenge; the other six strains were classified as sensitive. Gene expression analysis supports the involvement of a number of previously identified genes, molecular pathways, and TFs in the host response to LPS but also identifies Hedgehog signaling as a novel pathway activated by LPS. B6;129Ptch1+/+ wild-type mice were shown to be more sensitive to systemic LPS than B6;129Ptch1+/- heterozygote littermates further supporting the role of Hedgehog signaling in systemic LPS response. RNA interference-mediated inhibition of 6 TFs (C/EBP, Cdx-2, E2F1, Hoxa4, Nhlh1, and Tead2), out of 15 tested, was found to diminish production of IL-6 and TNFalpha protein in murine macrophages. The role of E2F1 was confirmed by showing that B6;129E2F1-/- knockout mice are more sensitive to systemic LPS than wild-type controls. Conclusions: Our analysis of gene expression data identified novel pathways and transcription factors that regulate the host response to systemic LPS. Our results provide potential sepsis biomarkers and therapeutic targets that should be further investigated in human populations. Keywords: disease state analysis

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Transcriptomic profiling of LPS-induced adrenal inflammation in WT mice

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