289 INHIBITION OF TOLL-LIKE RECEPTOR SIGNALING IN HUMAN MACROPHAGES BY BILE ACIDS

Abstract

Background and Aims: Innate immune recognition of microbial components is dependent on pattern recognition receptors (PRRs), which detect components of foreign pathogens referred to as pathogen associated molecular patterns (PAMPs). One important group of PRRs consists of the so called Toll like receptors (TLRs), which activate a network of signal pathways leading to activation of the transcription factors IRF3, NF-uB and AP-1 and subsequent expression of pro-inflammatory cytokines and type I interferons (IFN). Methods: Peripheral blood mononuclear cells (PBMC) were isolated from human blood (healthy volunteers, man) by Ficoll Paque centrifugation. CD14 positive cells were separated by magnetically labelled beads and cultured for 9 days in the presence of M-CSF to differentiate them into macrophages. Thereafter cells were stimulated with different TLR ligands in presence or absence of hydrophobic bile acids like taurolithocholic acid (TLC), glycochenodeoxycholic acid (GCDC) and taurochenodeoxycholic acid (TCDC). Cells were lysed and mRNA and protein expression were measured by realtime PCR and immunoblot respectively. Results: Hydrophobic bile acids changed cytokine expression pattern of TLR ligand stimulated macrophages. LPS-induced TNF-a, IL-6 and IFN-bmRNA expression and expression of antiviral proteins like MxA and PKR were significantly downregulated in response to hydrophobic bile acids, however LPS-induced expression of the strong inhibitory anti-inflammatory interleukin-10 was not modulated. These effects could be mimicked by cAMP. TLC also inhibited TNF-a and IFN-b mRNA expression induced by other TLR ligands like TLR 2, 3, 7 and 9. Phosphorylation of TLR4 dependent signaling elements like p-IRF3 (Ser) and p-p65 (Ser) were not effected by bile acids. However, the inhibitory effect of bile acids on cytokine expression was accompanied by elevation of cAMP and could be reverted by blocking proteinkinase A. This suggested an involvement of the bile acid receptor TGR5 in inhibition of TLR-responses, which we detected in human macrophages by immunostaining. Conclusions: These data show that bile acids mediate immunosuppressive effects by blocking TLR-induced activation of innate immunity responses in human macrophages, thereby probably changing differentiation and activation of macrophages. This may contribute to the known clinical observations that cholestatic patients with high serum bile acid concentrations show reduced cell mediated immunity.