Abstract
BackgroundPathogens stimulate immune functions of macrophages. Macrophages are a key sentinel cell regulating the response to pathogenic ligands and orchestrating the direction of the immune response. Our study aimed at investigating the early transcriptomic changes of bovine macrophages (Bomacs) in response to stimulation with CpG DNA or polyI:C, representing bacterial and viral ligands respectively, and performed transcriptomics by RNA sequencing (RNASeq). KEGG, GO and IPA analytical tools were used to reconstruct pathways, networks and to map out molecular and cellular functions of differentially expressed genes (DE) in stimulated cells.ResultsA one-way ANOVA analysis of RNASeq data revealed significant differences between the CpG DNA and polyI:C-stimulated Bomac. Of the 13,740 genes mapped to the bovine genome, 2245 had p-value ≤0.05, deemed as DE. At 6 h post stimulation of Bomac, poly(I:C) induced a very different transcriptomic profile from that induced by CpG DNA. Whereas, 347 genes were upregulated and 210 downregulated in response to CpG DNA, poly(I:C) upregulated 761 genes and downregulated 414 genes. The topmost DE genes in poly(I:C)-stimulated cells had thousand-fold changes with highly significant p-values, whereas in CpG DNA stimulated cells had 2–5-fold changes with less stringent p-values. The highest DE genes in both stimulations belonged to the TNF superfamily, TNFSF18 (CpG) and TNFSF10 (poly(I:C)) and in both cases the lowest downregulated gene was CYP1A1. CpG DNA highly induced canonical pathways that are unrelated to immune response in Bomac. CpG DNA influenced expression of genes involved in molecular and cellular functions in free radical scavenging. By contrast, poly(I:C) highly induced exclusively canonical pathways directly related to antiviral immune functions mediated by interferon signalling genes. The transcriptomic profile after poly(I:C)-stimulation was consistent with induction of TLR3 signalling.ConclusionCpG DNA and poly(I:C) induce different early transcriptional landscapes in Bomac, but each is suited to a specific function of macrophages during interaction with pathogens. Poly(I:C) influenced antiviral response genes, whereas CpG DNA influenced genes important for phagocytic processes. Poly(I:C) was more potent in setting the inflammatory landscape desirable for an efficient immune response against virus infection.
Highlights
IntroductionLewandowska-Sabat et al [5] have reported the early phase transcriptional program of bovine monocyte-derived MΦ infected with Staphylococcus aureus and show that S. aureus induces both, alternative and classical MΦ activation pathways
As shown in Additional file 1, the bovine macrophages (Bomacs) can phogocytose Staphylococcus aureus labelled with FITC, albeit at very low level compared to freshly isolated bovine monocytes
Using the data derived from gene expression analysis, Ingenuity Pathway Analysis (IPA), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene ontology (GO) were used to reconstruct cardinal networks, pathways, and cellular function groups of genes differentially expressed in the Bomac cell line stimulated with CpG DNA or poly(I:C)
Summary
Lewandowska-Sabat et al [5] have reported the early phase transcriptional program of bovine monocyte-derived MΦ infected with Staphylococcus aureus and show that S. aureus induces both, alternative and classical MΦ activation pathways. They concluded that activation of MΦ through the alternative pathway possibly contributes to intracellular persistence of S. aureus during mastitis in dairy cattle. This study revealed a novel iron assimilation system for carboxymycobactin Another RNASeq study of MAP infection [7] of monocyte-derived MΦ showed expression of genes that account for protective host immunity and those that might support MAP survival and proliferation in MΦ
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