A dual role for hepatocyte-intrinsic canonical NF-κB signaling in virus control.

BACKGROUND & AIMS: Hepatic innate immune control of viral infections has largely been attributed to Kupffer cells, the liver-resident macrophages. However, hepatocytes, the parenchymal cells of the liver, also possess potent immunological functions in addition to their known metabolic functions. Owing to their abundance in the liver and known immunological functions, we aimed to investigate the direct antiviral mechanisms employed by hepatocytes. METHODS: Using lymphocytic choriomeningitis virus (LCMV) as a model of liver infection, we first assessed the role of myeloid cells by depletion prior to infection. We investigated the role of hepatocyte-intrinsic innate immune signaling by infecting mice lacking canonical NF-κB signaling (IkkßΔHep) specifically in hepatocytes. In addition, mice lacking hepatocyte-specific interferon-α/ß signaling-(IfnarΔHep), or interferon-α/ß signaling in myeloid cells-(IfnarΔMyel) were infected. RESULTS: Here, we demonstrate that LCMV activates NF-κB signaling in hepatocytes. LCMV-triggered NF-κB activation in hepatocytes did not depend on Kupffer cells or TNFR1 signaling but rather on Toll-like receptor signaling. LCMV-infected IkkßΔHep livers displayed strongly elevated viral titers due to LCMV accumulation within hepatocytes, reduced interferon-stimulated gene (ISG) expression, delayed intrahepatic immune cell influx and delayed intrahepatic LCMV-specific CD8+ T cell responses. Notably, viral clearance and ISG expression were also reduced in LCMV-infected primary hepatocytes lacking IKKß, demonstrating a hepatocyte-intrinsic effect. Similar to livers of IkkßΔHep mice, enhanced hepatocytic LCMV accumulation was observed in livers of IfnarΔHep mice, whereas IfnarΔMyel mice were able to control LCMV infection. Hepatocytic NF-κB signaling was also required for efficient ISG induction in HDV-infected dHepaRG cells and interferon-α/ß-mediated inhibition of HBV replication in vitro. CONCLUSIONS: Together, these data show that hepatocyte-intrinsic NF-κB is a vital amplifier of interferon-α/ß signaling, which is pivotal for strong early ISG responses, immune cell infiltration and hepatic viral clearance. LAY SUMMARY: Innate immune cells have been ascribed a primary role in controlling viral clearance upon hepatic infections. We identified a novel dual role for NF-κB signaling in infected hepatocytes which was crucial for maximizing interferon responses and initiating adaptive immunity, thereby efficiently controlling hepatic virus replication.

Investigating the epigenetic profile of the inflammatory gene IL-6 in late-life depression.

BACKGROUND: It is well established that there is a link between inflammation and depression, with several studies reporting increased circulating levels of the pro-inflammatory cytokine, interleukin-6 (IL6), in depressed individuals. Peripheral epigenetic marks, including DNA methylation, hold promise as biomarkers for a range of complex conditions, with potential to inform diagnosis and tailor interventions. The aim of this study was to determine whether individuals with depression display differential methylation of the IL6 gene promoter compared to individuals without depression. METHODS: The ESPRIT study of later life neuropsychiatric disorders used a random sampling framework to select non-institutionalised participants aged ≥65 years and over living in the Montpellier region of France. Major depressive disorder (MDD) was assessed using the Mini International Neuropsychiatric Interview (MINI) according to DSM-IV criteria. High levels of depressive symptoms were defined as a score of ≥16 on the Centre for Epidemiologic Studies Depression Scale (CES-D). IL6 promoter DNA methylation was measured on a sub-sample of 380 participants who provided buccal samples. RESULTS: Individuals with depression (current MDD or high depressive symptoms) had lower IL6 methylation levels at one of the four sites investigated, however the effect size was small (∆ 2.4%, SE 0.009, p = 0.006). Interestingly, antidepressant use was independently associated with higher IL-6 methylation at the same site (∆ 4.6%, SE 0.019, p = 0.015). In multivariate linear regression analyses adjusting for covariates, including sex and smoking status, these associations remained. There was no effect modification when considering IL6 genotype. CONCLUSION: This study presents evidence that IL6 methylation may be a marker of depression status in older individuals, however further work is now needed to replicate these findings and to assess the association with inflammatory status of individuals.

Control of APOBEC3B induction and cccDNA decay by NF-κB and miR-138-5p.

BACKGROUND & AIMS: Immune-mediated induction of cytidine deaminase APOBEC3B (A3B) expression leads to HBV covalently closed circular DNA (cccDNA) decay. Here, we aimed to decipher the signalling pathway(s) and regulatory mechanism(s) involved in A3B induction and related HBV control. METHODS: Differentiated HepaRG cells (dHepaRG) knocked-down for NF-κB signalling components, transfected with siRNA or micro RNAs (miRNA), and primary human hepatocytes ± HBV or HBVΔX or HBV-RFP, were treated with lymphotoxin beta receptor (LTßR)-agonist (BS1). The biological outcomes were analysed by reverse transcriptase-qPCR, immunoblotting, luciferase activity, chromatin immune precipitation, electrophoretic mobility-shift assay, targeted-bisulfite-, miRNA-, RNA-, genome-sequencing, and mass-spectrometry. RESULTS: We found that canonical and non-canonical NF-κB signalling pathways are mandatory for A3B induction and anti-HBV effects. The degree of immune-mediated A3B production is independent of A3B promoter demethylation but is controlled post-transcriptionally by the miRNA 138-5p expression (hsa-miR-138-5p), promoting A3B mRNA decay. Hsa-miR-138-5p over-expression reduced A3B levels and its antiviral effects. Of note, established infection inhibited BS1-induced A3B expression through epigenetic modulation of A3B promoter. Twelve days of treatment with a LTßR-specific agonist BS1 is sufficient to reduce the cccDNA pool by 80% without inducing significant damages to a subset of cancer-related host genes. Interestingly, the A3B-mediated effect on HBV is independent of the transcriptional activity of cccDNA as well as on rcDNA synthesis. CONCLUSIONS: Altogether, A3B represents the only described enzyme to target both transcriptionally active and inactive cccDNA. Thus, inhibiting hsa-miR-138-5p expression should be considered in the combinatorial design of new therapies against HBV, especially in the context of immune-mediated A3B induction. LAY SUMMARY: Immune-mediated induction of cytidine deaminase APOBEC3B is transcriptionally regulated by NF-κB signalling and post-transcriptionally downregulated by hsa-miR-138-5p expression, leading to cccDNA decay. Timely controlled APOBEC3B-mediated cccDNA decay occurs independently of cccDNA transcriptional activity and without damage to a subset of cancer-related genes. Thus, APOBEC3B-mediated cccDNA decay could offer an efficient therapeutic alternative to target hepatitis B virus chronic infection.