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Genome-wide comparison between IL-17 and combined TNF-alpha/ IL-17 induced genes in primary murine hepatocytes

ABSTRACT: Cytokines such as TNF-alpha and IL-1beta are known for their contribution to inflammatory processes in liver . In contrast, the cytokine IL-17 has not yet been assigned a role in liver diseases. IL-17 can cooperate with TNF-alpha to induce a synergistic response on several target genes in different cell lines, but no data exist for primary hepatocytes. To enhance our knowledge on the impact of IL-17 alone and combined with TNF-alpha in primary murine hepatocytes a comprehensive microarray study was designed. IL-1beta was included as this cytokine is suggested to act in a similar manner as the combination of TNF-alpha and IL-17, especially with respect to its role in mRNA stabilization. Results: The present microarray analysis demonstrates that primary murine hepatocytes responded to IL-17 stimulation by upregulation of chemokines and genes, which are functionally responsible to increase and sustain inflammation. Cxcl2, Nfkbiz and Zc3h12a were strongly induced, whereas the majority of the genes were only very moderately upregulated. Promoter analysis revealed involvement of NF-kappaB in the activation of many genes. Combined stimulation of TNF-alpha/IL-17 resulted in enhanced induction of gene expression, but significantly synergistic effects could be applied only to a few genes, such as Nfkbiz, Cxcl2, Zc3h12 and Steap4. Comparison of the gene expression profile obtained after stimulation of TNF-alpha/IL-17 versus IL-1 proposed a IL-1beta-like effect of the latter cytokine combination. Moreover, evidence was provided that modulation of mRNA stability may be a major mechanism by which IL-17 regulates gene expression in primary hepatocytes. This assumption was exemplarily proven for Nfkbiz mRNA for the first time in hepatocytes. Our studies also suggest that RNA stability can partially be correlated to the existence of AU rich elements, but further mechanisms like the RNase-activity of the upregulated Zc3h12a have to be considered. Conclusions: Our microarray analysis gives new insights in IL-17 induced gene expression in primary hepatocytes highlighting the crosstalk with the NF-kappaB signalling pathway. Gene expression profile suggests IL-17 a role in sustaining liver inflammatory processes most likely by RNA stabilization. Altogether, our results provide evidence that IL-17 alone and in concert with TNF-alpha may play a role in inflammatory liver diseases.

ORGANISM(S): Mus musculus

PROVIDER: GSE19272 | GEO | 2010/03/01

SECONDARY ACCESSION(S): PRJNA120949

REPOSITORIES: GEO

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Project description:Cytokines such as TNF-alpha and IL-1beta are known for their contribution to inflammatory processes in liver . In contrast, the cytokine IL-17 has not yet been assigned a role in liver diseases. IL-17 can cooperate with TNF-alpha to induce a synergistic response on several target genes in different cell lines, but no data exist for primary hepatocytes. To enhance our knowledge on the impact of IL-17 alone and combined with TNF-alpha in primary murine hepatocytes a comprehensive microarray study was designed. IL-1beta was included as this cytokine is suggested to act in a similar manner as the combination of TNF-alpha and IL-17, especially with respect to its role in mRNA stabilization. Results: The present microarray analysis demonstrates that primary murine hepatocytes responded to IL-17 stimulation by upregulation of chemokines and genes, which are functionally responsible to increase and sustain inflammation. Cxcl2, Nfkbiz and Zc3h12a were strongly induced, whereas the majority of the genes were only very moderately upregulated. Promoter analysis revealed involvement of NF-kappaB in the activation of many genes. Combined stimulation of TNF-alpha/IL-17 resulted in enhanced induction of gene expression, but significantly synergistic effects could be applied only to a few genes, such as Nfkbiz, Cxcl2, Zc3h12 and Steap4. Comparison of the gene expression profile obtained after stimulation of TNF-alpha/IL-17 versus IL-1 proposed a IL-1beta-like effect of the latter cytokine combination. Moreover, evidence was provided that modulation of mRNA stability may be a major mechanism by which IL-17 regulates gene expression in primary hepatocytes. This assumption was exemplarily proven for Nfkbiz mRNA for the first time in hepatocytes. Our studies also suggest that RNA stability can partially be correlated to the existence of AU rich elements, but further mechanisms like the RNase-activity of the upregulated Zc3h12a have to be considered. Conclusions: Our microarray analysis gives new insights in IL-17 induced gene expression in primary hepatocytes highlighting the crosstalk with the NF-kappaB signalling pathway. Gene expression profile suggests IL-17 a role in sustaining liver inflammatory processes most likely by RNA stabilization. Altogether, our results provide evidence that IL-17 alone and in concert with TNF-alpha may play a role in inflammatory liver diseases. Primary murine hepatocytes of three animals stimulated for 1 or 4h by TNF-alpha, IL-1beta, IL-17 or TNF-alpha followed by IL-17 were used for microarray analysis.

Project description:Resistance to erythropoietin (EPO) treatment is observed in a considerable proportion of anemic patients with chronic kidney disease. Previous reports suggest that inflammation is one of the major independent predictors of this resistance, and pro-inflammatory cytokines have been shown to inhibit erythropoiesis. The aim of this study was to investigate EPO-induced modification to gene expression in primary cultured leukocytes. Microarray experiments were performed on ex vivo Peripheral Blood Mononuclear Cells (PBMCs) from pooled ten healthy donors (n=9), primed or not with TNF-alpha, and treated with recombinant human erythropoietin alpha (EPO-alpha). Real-time PCR experiments were used to validate expression of the molecular targets considered most relevant for inflammation. Data analysis suggested that EPO-alpha treatment mainly modulated genes involved in cell movement and cell-cell interaction in primed PBMCs. Notably, EPO-alpha exerts anti-inflammatory effects inhibiting the expression of pro-inflammatory cytokine IL-8 and its receptor CXCR2; by contrast, EPO-alpha further increases expression of genes relating to promotion of inflammation encoding for IL-1beta and CCL8, and induces de novo synthesis of IL-1alpha, CXCL1 and CXCL5 in primed PBMCs. MAPK p38alpha?reducing activity is involved in modulation of IL-1beta and IL-8 expression. In conclusion, our findings confirm the anti-inflammatory role for EPO, but also suggest a plausible in vivo scenario, in which the positive correlation found between EPO-resistance and elevated levels of some pro-inflammatory mediators is due to treatment with EPO itself. One future objective will be to perform in vivo measurement of the molecular targets highlighted, in order to identify any additional markers involved in EPO-resistance among hypo-responsive patients. In this study, we analyzed the expression profiles of pooled blood PBMCs from 10 healthy donors primed or not with TNF-alpha and then treated with EPO-alpha. All RNAs from no stimulated PBMCs were hybridized against RNAs from PBMCs treated with TNF-alpha or TNF-alpha and EPO-alpha. We performed a total of 9 technical replicates.

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Genome-wide comparison between IL-17 and combined TNF-alpha/ IL-17 induced genes in primary murine hepatocytes

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