Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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The effects of terbutaline or GW9508 on TNF-alpha and IFN gamma (TNF-alpha + IFN gamma) stimulation by HaCaT

ABSTRACT: G-protein coupled receptors (GPCRs) have diverse roles in physiological processes, including immunity. Gs-coupled GPCRs increase while Gi-coupled ones decrease intracellular cAMP. Previous studies suggest that, in epithelial cells, Gs-coupled GPCRs enhance whereas Gi-coupled GPCRs suppress pro-inflammatory immune responses. In order to examine the issue, we chose beta2 adrenergic receptor and GPR40 as representatives of Gs- and Gi- coupled GPCRs, respectively, and examined their effects on TNF-alpha and IFN-gamma-(TNF-alpha + IFN-gamma) induced gene expression by HaCaT. We used microarrays to detail the global changes of gene expression induced by a beta2 adrenergic receptor agonist terbutaline or GPR40 agonist GW9508 pre-treatment in TNF-alpha + IFN-gamma - stimulated HaCaT cells. HaCaT cells were pre-treated with terbutaline or GW9508, TNF-alpha + IFN-gamma were then added, and cultured for another 24 h. Cells were then used for RNA extraction and hybridization on Affymetrix microarrays. We sought to clarify changes in gene expression after 1) TNF-alpha + IFN-gamma, 2) TNF-alpha + IFN-gamma + terbutaline, and 3) TNF-alpha + IFN-gamma + GW9508 treatment. To this end, we set 4 groups of samples; 1) unstimulated group, 2) TNF-alpha + IFN-gamma-stimulated group, 3) TNF-alpha + IFN-gamma + terbutaline-stimulated group, and 4) TNF-alpha + IFN-gamma + GW9508-stimulated group. In each group, HaCaT cells were stimulated in triplicate wells (n=3).

ORGANISM(S): Homo sapiens

SUBMITTER: Tomoko Fujita

PROVIDER: E-GEOD-20297 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Approximately 50% of patients with chronic hepatitis C (CHC) have a sustained virologic response (SVR) to treatment with pegylated interferon (pegINF)-α and ribavirin. Non-response to treatment is associated with constitutively increased expression of IFN-stimulated genes (ISGs) in the liver. Treatment of patients with acute hepatitis C (AHC) is more effective, with SVR rates >90%. We investigated mechanisms of the different responses of patients with CHC and AHC to pegIFN-α therapy. We analyzed IFN signaling and ISG expression in liver samples from patients with acute hepatitis C (AHC), patients with chronic hepatitis (CHC), and individuals without hepatitis C (controls) using microarray, immunohistochemical, and protein analyses. Findings were compared with those from primary human hepatocytes stimulated with IFN-α or IFN-γ, as reference sets. Expression levels of 100s of genes, primarily those regulated by IFN-γ, were altered in liver samples from patients with AHC compared with controls. Expression of IFN-γ–stimulated genes was induced in liver samples from patients with AHC, whereas expression of IFN-α–stimulated genes was induced in samples from patients with CHC. In an expression analysis of negative regulators of IFN-α signaling, we did not observe differences in expression of SOCS1 or SOCS3 between liver samples from patients with AHC and those with CHC. However, USP18 (another negative regulator of IFN-α signaling), was upregulated in liver samples of patients with CHC that did not respond to therapy, but not in AHC. In conclusion, differences in expression of ISGs might account for the greater response of patients with AHC, compared to those with CHC, to treatment with pegINF-α and ribavirin. Specifically, USP18 is upregulated in liver samples of patients with CHC that do not respond to therapy, but not in patients with AHC. (Interferon-γ Stimulated Genes, but not USP18, are Expressed in Livers of Patients with Acute Hepatitis C; Dill MT, Makowska Z et al, Gastroenterology 2012 (in press)) Primary human hepatocytes from 2 donors were analyzed. From each donor there are 5 samples: untreated cells, cells treated with interferon alpha (1000 IU/ml) for 6 and 24 hours and cells treated with interferon gamma (1000 IU/ml) for 6 and 24 hours.

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The effects of terbutaline or GW9508 on TNF-alpha and IFN gamma (TNF-alpha + IFN gamma) stimulation by HaCaT

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