Increased sensitivity of Treg cells from patients with PBC to low dose IL-12 drives their differentiation into IFN-γ secreting cells

Abstract

IL-12 is a pro-inflammatory cytokine that induces the production of interferon-γ (IFNγ) and favours the differentiation of T helper 1 (Th1) cells. In the presence of IL-12 human Treg cells acquire a Th1-like phenotype with reduced suppressive activity in vitro. Primary biliary cholangitis (PBC) is an autoimmune cholestatic liver disease characterised by high Th1 and Th17 infiltrating cells, reduced frequencies of Treg cells, and a genetic association with IL-12 signalling. Herein, we sought to evaluate the IL-12 signalling pathway in PBC pathology, by studying human samples from patients with PBC, alongside those with primary Sjögren's syndrome (pSS)(autoimmune disease with IL-12 signalling gene association), primary sclerosing cholangitis (PSC) (cholestatic liver disease without IL-12 gene association) and healthy individuals. Our data revealed that TLR stimulation of PBC (n = 17) and pSS monocytes (n = 6) resulted in significant induction of IL12A mRNA (p < 0.05, p < 0.01, respectively) compared to PSC monocytes (n = 13) and at similar levels to HC monocytes (n = 8). PSC monocytes expressed significantly less IL-12p70 (108 pg/ml, mean) and IL-23 (358 pg/ml) compared to HC (458 pg/ml and 951 pg/ml, respectively) (p < 0.01, p < 0.05). Treg cells from patients with PBC (n = 16) and pSS (n = 3) but not PSC (n = 10) and HC (n = 8) responded to low dose (10 ng/ml) IL-12 stimulation by significant upregulation of IFNγ (mean 277 and 254 pg/ml, respectively) compared to PSC and HC Treg cells (mean 22 and 77 pg/ml, respectively)(p < 0.05). This effect was mediated by the rapid and strong phosphorylation of STAT4 on Treg cells from patients with PBC and pSS (p < 0.05) but not PSC and HC. In the liver of patients with PBC (n = 7) a significantly higher proportion of IL-12Rβ2Tregs (16% on average) was detected (p < 0.05) compared to other liver disease controls (5%)(n = 18) which also showed ex vivo high IFNG and TBET expression. CONCLUSION: Our data show an increased sensitivity of PBC and pSS Treg cells to low dose IL-12 stimulation, providing ongoing support for the importance of the IL12-IL-12Rβ2-STAT4 pathway on Treg cells in disease pathogenesis and potentially treatment.

The Pathogenesis of Primary Biliary Cholangitis: A Comprehensive Review

“…162 Liaskou et al recently showed that Treg cells from patients with PBC respond to low-dose IL-12 stimulation by significant upregulation of IFN-γ; this effect was mediated by the rapid and strong phosphorylation of STAT4 on Treg cells from patients with PBC providing ongoing support for the importance of the IL12-IL-12Rβ2-STAT4 pathway on Treg cells in disease pathogenesis and potentially treatment. 163 Finally, increased frequencies of Th17 lymphocytic infiltrates have been documented in liver sections from PBC patients, and importantly, they are primarily localized around the damaged interlobular bile ducts in PBC. Most importantly, Th17 skewing is prominent in advanced PBC patients with intensive secretion of IL-23p19 by inflamed hepatocytes around IL-23R, IL-12Rβ2, and IFN-γ expressing degenerated cholangiocytes.…”

Phenotyping multiple subsets in Sjögren’s syndrome: a salivary proteomic SWATH-MS approach towards precision medicine

“…We specifically focused on S100A proteins that have been reported to be increased in saliva and serum of pSS [32] and associated also to a high risk of vascular complications in pSS patients [33–35]. S100 A proteins converged on IL-12 signaling at the GO analysis, a biological process that has been demonstrated to play active roles in the expansion and organization of infiltrative injuries in SS, mirroring specifically DC involvement and Th1 cells differentiation in the disease pathogenesis [6, 36]. We found that S100A proteins were expressed abundantly in saliva, at local sites of inflammation, and we also observed their increase in a stage-manner from pSS-phenotype with normal flow to pSS-phenotype with low flow.…”

Constitutive Activation of Natural Killer Cells in Primary Biliary Cholangitis

“…Liaskou et al have previously shown that T reg cells from patients with PBC were highly sensitive to IL-12, but in that study IL-12 concentrations of 10 ng/ml were used (44). This is 2,000 times the concentration we used to demonstrate upregulation of CXCR6 and CD49a on NK cells from PBC patients.…”