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IFN-b priming exacerbates LPS-driven proinflammatory cytokines in macrophages.

ABSTRACT: Purpose: We characterize the changes global expression profile in peripheral macrophages induced by LPS, IFN-b, and IFN-B+LPS treatment, using bulk RNA-sequencing. Methods: mRNA profiles of bulk macrophages obtained from WT mice following treatment with either saline, IFN-b, LPS, or IFN-b+LPS. Each treatment was performed in triplicate, and RNA-sequencing was performed on the Illumina 2500. Reads passing quality metric were aligned to the mm10 genome using annotations produced by ENSEMBL, and were analyzed at the transcript level using one-way ANOVA and fold change requirements. Results: We mapped approximately 20 million reads per sample to the mm10 genome, yielding 11385 reasonably-expressed transcripts. We identified 4372 transcripts with a p-value<0.05 and fold change>2 when compared between LPS v NS, IFNb v NS, and LPS+IFNb v NS. Of these, we identified 142 genes where priming by IFNb+LPS augmented the changes in expression induced by LPS or IFNb independently. This gene set indicated that IFNb exacerbates LPS_driven proinflammatory cytokin production. Conclusion: Here we show that IFNb priming sensitizes to TLD-driven inflammation through LPS stimulation.

ORGANISM(S): Mus musculus

PROVIDER: GSE82087 | GEO | 2021/06/01

REPOSITORIES: GEO

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Project description:Purpose: Virus infections of the CNS cause important diseases of humans and animals. As in other tissues, innate antiviral responses mediated by type I interferons (IFN) are crucially important in controlling CNS virus infections. Semliki Forest virus (SFV) infection of the mouse provides a well-characterised and tractable model to study the pathogenesis of virus encephalitis. The maturity of neuronal populations is an established critical factor determining the outcome of CNS virus infection. Using primary cultures of mouse cortical neurons, we investigated the relationships between neuronal maturation, type I IFN responses and the outcome of SFV infection. Methods: mRNA profiles of immature and mature of primary mouse cortical neurons that were mock or iFNβ treated and mock or SFV infected were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500. All 100-base-pair single-end reads were mapped to the reference mouse genome (GRCm38/mm10) using Salmon ultrafast aligner. Mapped reads were quantitated to the gene level estimates and genecount data was imported using the tximport package and differential expression analysis of RNA sequencing data was carried out using the edgeR and limma-voom packages in Bioconductor. Results: Using an optimized data analysis workflow, we mapped the 100 bp single end reads to the mouse genome (build mm10) and identified 14,933 genes in the immature and mature primary neurons with different treatments. Transcriptomic profiling revealed 3,666 genes differentially expressed between resting immature and mature cells. Multidimensional scaling plots revealed key differences between immature and mature neurons with different treatments. Immature neurons pre-treated with type I IFN and mock-infected or infected without prior IFN, clustered close to the basal (resting) control samples,indicating limited changes in gene expression under either condition. This was also the case for mature neurons treated with IFNβ. Whereas, immature neurons pre-treated with IFN and then infected, as well as mature neurons pre-treated or not with IFN and then infected underwent more dynamic changes in gene expression Conclusions: Complete transcriptome analysis demonstrated that resting immature neurons lacked the transcriptional competence to mount antiviral responses. They had no detectable transcription of the genes Ddx58 and Ifih1 which encode the key RNA virus cytoplasmic sensors RIG-I and MDA5 and no, or very low, expression of genes encoding key regulators of downstream and associated signalling pathways. Upon infection, immature neurons failed to mount an antiviral response as evidenced by their failure to produce chemokines, IFNs and other cytokines. Treatment of immature neurons with exogenous IFNb prior to infection resulted in antiviral responses and lower levels of virus replication and infectious virus production. In contrast, resting mature neurons expressed Ddx58 and Ifih1 and upon infection generated a robust antiviral response. This was augmented by pre-treatment with IFNb. Infection of mature neurons derived from IFNAR-/- mice did not make an antiviral response and replicated virus to high levels.

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IFN-b priming exacerbates LPS-driven proinflammatory cytokines in macrophages.

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