Type 2 innate lymphoid cells (ILC2) mediate inflammatory immune responses in the context of diseases triggered by the alarmin IL-33. In recent years, IL-33 has been implicated in the pathogenesis of immune-mediated liver diseases. However, the immunoregulatory function of ILC2s in the inflamed liver remains elusive. Using the murine model of Con A-induced immune-mediated hepatitis, we showed that selective expansion of ILC2s in the liver was associated with highly elevated hepatic IL-33 expression, severe liver inflammation, and infiltration of eosinophils. CD4 T cell-mediated tissue damage and subsequent IL-33 release were responsible for the activation of hepatic ILC2s that produced the type 2 cytokines IL-5 and IL-13 during liver inflammation. Interestingly, ILC2 depletion correlated with less severe hepatitis and reduced accumulation of eosinophils in the liver, whereas adoptive transfer of hepatic ILC2s aggravated liver inflammation and tissue damage. We further showed that, despite expansion of hepatic ILC2s, 3-d IL-33 treatment before Con A challenge potently suppressed development of immune-mediated hepatitis. We found that IL-33 not only activated hepatic ILC2s but also expanded CD4 Foxp3 regulatory T cells (Treg) expressing the IL-33 receptor ST2 in the liver. This Treg subset also accumulated in the liver during resolution of immune-mediated hepatitis. In summary, hepatic ILC2s are poised to respond to the release of IL-33 upon liver tissue damage through expression of type 2 cytokines thereby participating in the pathogenesis of immune-mediated hepatitis. Inflammatory activity of ILC2s might be regulated by IL-33-elicited ST2 Tregs that also arise in immune-mediated hepatitis.

Background/Aims: In this observational study, we explored the humoral and cellular immune response to SARS-CoV-2 vaccination in patients with autoimmune hepatitis (AIH) and patients with cholestatic autoimmune liver disease (primary sclerosing cholangitis [PSC] and primary biliary cholangitis [PBC]). Methods: Anti-SARS-CoV-2 antibody titers were determined using the DiaSorin LIAISON and Roche immunoassays in 103 AIH, 64 PSC, and 61 PBC patients and 95 healthy controls >14 days after the second COVID-19 vaccination. The spikespecific T-cell response was assessed using an activation-induced marker assay (AIM) in a subset of individuals.Results: Previous SARS-CoV-2 infection was frequently detected in AIH but not in PBC/PSC (10/112 (9%), versus 4/144 (2.7%), p = 0.03). In the remaining patients, seroconversion was measurable in 97% of AIH and 99% of PBC/PSC patients, respectively. However, in 13/94 AIH patients antibody levels were lower than in any healthy control, which contributed to lower antibody levels of the total AIH cohort when compared to PBC/PSC or controls (641 vs. 1020 vs. 1200 BAU/ml, respectively). Notably, antibody levels were comparably low in AIH patients with (n = 85) and without immunosuppression (n = 9). Also, antibody titers significantly declined within 7 months after the second vaccination. In the AIM assay of 20 AIH patients, a spike-specific T-cell response was undetectable in 45% despite a positive serology, while 87% (13/15) of the PBC/PSC demonstrated a spike-specific T-cell response.Paul Duengelhoef, Johannes Hartl and Darius Rüther shared co-first author ship.

In the present study, we investigated the phenotype and plasticity of hepatic ILC2 in response to liver inflammation-induced cytokines and effector CD4 + T cells in order to identify mechanisms of ILC2 regulation in liver disease. Material and Methods Mice. C57BL/6 mice and OT-II mice were bred in the University Medical Center Hamburg-Eppendorf animal facility (Hamburg, Germany). Mouse experiments were conducted according to the German animal protection law and approved by the institutional review board (Behörde für Gesundheit und Verbraucherschutz, Hamburg, Germany; G44/15). Mice received humane care according to the national guidelines of the National Institutes of Health in Germany. Animal treatment. C57BL/6 mice received intraperitoneally recombinant murine (rm)IL-33 (0.3 μg; BioLegend, San Diego, CA) daily on four consecutive days. ConA (7 mg/kg; Sigma-Aldrich, St. Louis, MO) was injected intravenously into C57BL/6 mice that were analyzed 8 and 24 hours later. Heart blood was drawn from individual mice and liver injury was quantified by automated measurement of plasma activities of alanine transaminase (ALT) using a COBAS Mira System (Roche Diagnostic, Mannheim, Germany). Immunohistochemistry. Liver samples were fixed with 4% formalin and embedded in paraffin. 3 μm liver sections were cut, stained with H&E following standard protocols, and analyzed by light microscopy. Isolation of hepatic ILC2 and CD4 + T cells. Hepatic non-parenchymal cells were isolated from livers of naive and IL-33-treated mice by Percoll density gradient centrifugation. Single cell suspensions were stained with the Lineage Antibody Cocktail (APC; BD Pharmingen, Heidelberg, Germany) as well as anti-Sca-1 (Pacific Blue; D7; BioLegend) and anti-ST2 (PerCP-eFluor 710; RMST2-2; ThermoFisher Scientific, Waltham, MA) antibodies. Lineage-negative (lin −) cells were enriched by magnetic cell sorting and lin − Sca-1 + ST2 + ILC2 were purely isolated by FACS (BD FACSAriaTM III sorter, BD Biosciences). Ovalbumin (OVA)-specific CD4 + T cells were isolated from spleen and lymph nodes of OT-II mice. Therefore, CD4 + T cells were enriched from single cell suspensions using the CD4 + T Cell Isolation Kit (Miltenyi Biotec, Bergisch Gladbach, Germany). Subsequently, cells were stained with anti-TCRβ (PE-Cy7; H57-597) and anti-CD4 (BV421; GK1.5; both BioLegend) antibodies and TCRβ + CD4 + T cells were purified by FACS.