After having shown that bacteria can be found in portal tracts of patients with PSC, we were interested in whether pathogen stimulation induces a proinflammatory phenotype in lymphocytes of patients with PSC. To this end, we determined whether stimulation of PBMCs with heat-inactivated bacteria or fungi may affect the expression of proinflammatory cytokines in vitro. Initially, pathogens isolated from patients’ own bile were used for stimulation of PBMCs. The results obtained were not different to those obtained Selleckchem Temsirolimus using standard pathogens, which were then used in subsequent stimulation assays. The

clinical characteristics of PSC and PBC patients and cholestatic controls included in these experiments are shown in Table 1. Stimulation with facultative pathogenic bacteria, such as E. faecalis, induced significantly more IL-17A-producing CD4+ cells in patients with PSC, as compared to HCs (E. faecalis; CD4+IL-17A+: PSC [2.22% ± 1.68%] versus HCs [0.77% ± 0.54%], P < 0.001; Fig. 3A). To investigate whether these findings are specific for PSC, we compared these results to patients with PBC as another autoimmune cholestatic liver disease also treated with UDCA as well as to

patients with different diseases leading to cholestasis, including SSC (see above). The increase in IL-17A-producing CD4+ T cells observed in PSC was not noted in patients with PBC or control cholestatic patients (E. faecalis, as compared to BAY 57-1293 concentration PSC: PBC: 0.57% ± 0.25%, P < 0.01; cholestatic controls: 0.53% ± 0.49%, P < 0.001; Fig. 3A). Because PSC-associated IBD may influence Th17 development through an impaired mucosal barrier function of the gut, patients with PSC and no evidence of IBD on colonoscopy were analyzed separately: Patients with PSC only were not different from patients with PSC and associated IBD, demonstrating that the increased frequency of IL-17A+CD4+ T cells is next an underlying feature of PSC itself (E. faecalis; PSC only: 3.24% ± 2.21% [P < 0.001],

as compared to HCs; Fig. 3A). S. aureus was found in 9% of bile samples. Stimulation with heat-killed S. aureus also led to an increased rate of IL-17A-producing CD4+ T cells in PSC patients, but not in patients with PBC or HCs (Fig. 3B). As noted above, this effect was independent from the presence of IBD (Fig. 3B). Interestingly, after stimulation with nonpathogenic heat-killed E. coli, there were no significant differences in IL-17A expression between PSC and HCs (data not shown). Also, rates of CD4+ T cells expressing IFN-γ or TNF-α after bacterial stimulation were similar between HCs and patients with PSC (Fig. 4). C. albicans was cultured from 12 of 58 individual bile samples and was previously described to have a negative effect on progression of disease, including time to liver transplantation.[5] After stimulation with C. albicans, up to 30% of CD4+ T cells produced IL-17A, which were the highest rates observed in our experiments (C.