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Expression data from human monocytes stimulated with wild-type N. Meningitidis (Nm) and recombinant IL-10 [I]

ABSTRACT: Background: Biological interpretation of gene expression data may differ depending on underlying assumptions and statistical tests used by the chosen bioinformatic tool. We used Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA) to analyze previously generated gene expression profiles induced in human monocytes by N. meningitidis and IL-10 (“the model system”), and by meningococcal sepsis plasma before and after immunodepletion of IL-10 (“the patient plasma system”). The objectives were to compare whether these two methods resulted in similar biological interpretation of the datasets; and identify whether GSEA provided additional insight about the human monocyte host response to meningococcal activation. Results: In both experimental models, GSEA and IPA associated genes induced in monocytes by N. meningitidis with pro-inflammatory innate immune activation, including TNF-signaling, Toll-like receptor signaling, JAK-STAT-signaling, type I and type II interferon. GSEA associated genes regulated by the presence of IL-10 with similar gene sets in both the model system and the patient plasma system. In the model system, GSEA and IPA identified 170 genes associated with oxidative phosphorylation/mitochondrial function to be down-regulated in monocytes stimulated with meningococci. In the patient plasma system, GSEA and IPA combined identified 122 genes associated with oxidative phosphorylation/mitochondrial dysfunction to be down-regulated by meningococcal sepsis plasma depleted for IL-10. Combining analyses from GSEA and IPA identified that some genes associated with oxidative phosphorylation/mitochondrial function that were inhibited by N. meningitidis were partially up-regulated by IL-10. Conclusions: Biological processes associated with the gene expression changes in the model system of meningococcal sepsis was comparable to the results found in the patient plasma system. By combining GSEA with IPA, we have discovered an inhibitory effect exerted by N. meningitidis on genes associated with oxidative phosphorylation, and that IL-10 possibly partially dampen this strong inhibitory effect, thereby identifying, to our knowledge, yet another area where IL-10 regulates the effect of LPS. We suggest that relying on a single bioinformatic tool together with arbitrarily chosen filtering criteria for data analysis may result in overlooking relevant biological processes and signaling pathways associated with genes differentially expressed between compared experimental conditions.

ORGANISM(S): Homo sapiens

PROVIDER: GSE82166 | GEO | 2016/06/08

SECONDARY ACCESSION(S): PRJNA324248

REPOSITORIES: GEO

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Project description:Background: Biological interpretation of gene expression data may differ depending on underlying assumptions and statistical tests used by the chosen bioinformatic tool. We used Gene Set Enrichment Analysis (GSEA) and Ingenuity Pathway Analysis (IPA) to analyze previously generated gene expression profiles induced in human monocytes by N. meningitidis and IL-10 (“the model system”), and by meningococcal sepsis plasma before and after immunodepletion of IL-10 (“the patient plasma system”). The objectives were to compare whether these two methods resulted in similar biological interpretation of the datasets; and identify whether GSEA provided additional insight about the human monocyte host response to meningococcal activation. Results: In both experimental models, GSEA and IPA associated genes induced in monocytes by N. meningitidis with pro-inflammatory innate immune activation, including TNF-signaling, Toll-like receptor signaling, JAK-STAT-signaling, type I and type II interferon. GSEA associated genes regulated by the presence of IL-10 with similar gene sets in both the model system and the patient plasma system. In the model system, GSEA and IPA identified 170 genes associated with oxidative phosphorylation/mitochondrial function to be down-regulated in monocytes stimulated with meningococci. In the patient plasma system, GSEA and IPA combined identified 122 genes associated with oxidative phosphorylation/mitochondrial dysfunction to be down-regulated by meningococcal sepsis plasma depleted for IL-10. Combining analyses from GSEA and IPA identified that some genes associated with oxidative phosphorylation/mitochondrial function that were inhibited by N. meningitidis were partially up-regulated by IL-10. Conclusions: Biological processes associated with the gene expression changes in the model system of meningococcal sepsis was comparable to the results found in the patient plasma system. By combining GSEA with IPA, we have discovered an inhibitory effect exerted by N. meningitidis on genes associated with oxidative phosphorylation, and that IL-10 possibly partially dampen this strong inhibitory effect, thereby identifying, to our knowledge, yet another area where IL-10 regulates the effect of LPS. We suggest that relying on a single bioinformatic tool together with arbitrarily chosen filtering criteria for data analysis may result in overlooking relevant biological processes and signaling pathways associated with genes differentially expressed between compared experimental conditions.

Project description:Background: Patients developing meningococcal septic shock reveal very high levels of Neisseira meningitidis and endotoxin in the circulation and organs, leading to acute cardiovascular, pulmonary and renal failure, coagulopathy and a high case fatality rate within 24 hours. Objective: To investigate transcriptional profiles in heart, lungs, kidneys, liver and spleen and immunostain key inflammatory cells and proteins in post mortem formalin-fixed, paraffin-embedded (FFPE) tissue samples from meningococcal septic shock patients. Patients and methods: Total RNA was isolated from FFPE and fresh frozen (FF) tissue samples from five patients and two controls (non-inflammatory disease). Differential expression of genes was detected using Affymetrix GeneChip Human Transcriptome 2.0 Arrays. Lung and heart tissue samples were immunostained for T-and B cells, macrophages, neutrophils and the inflammatory markers Plasminogen activator inhibitor-1 (PAI-1) and Monocyte chemoattractant protein 1 (MCP-1). Inflammatory mediators were quantified in lysates from FF tissues. Results: The transcriptional profiles showed significant regulation of pathways associated with organismal death, cell death and survival, leukocyte migration, cellular movement, proliferation of cells, cell-to-cell signaling, immune cell trafficking and inflammatory responses in an organ-specific clustering manner. The canonical pathways including acute phase response-, EIF2-, TREM1-, IL-6-, HMBG1-, PPAR signaling and LXR/RXR activation were associated with acute heart, pulmonary and renal failure and with inflammatory changes in liver and spleen were identified. The main upstream regulators were TNF, IL-1β, IL-6, RICTOR, miR-6739-3p and CD3. Increased numbers of inflammatory cells (CD68+, MPO+, CD3+ and CD20+) were found in lungs and heart. PAI-1 inhibiting fibrinolysis and MCP-1 attracting leukocyte were found significantly present in the septic tissue samples compared to the controls. Conclusions: Hundreds of transcripts were altered in lungs, heart and kidneys, all organs with an acute dysfunction. Relatively fewer transcripts were changed in the spleen. Certain transcripts were associated with the number of. N. meningitidis in the tissues. We identified specific biomarker panels which differed from organ to organ. Our results highlight the complexity of gene regulation in the different tissues. Future treatment strategies blocking or stimulating only one molecule or pathway appear unlikely to succeed.

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Expression data from human monocytes stimulated with wild-type N. Meningitidis (Nm) and recombinant IL-10 [I]

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