BACKGROUND. The clearance of hepatitis B surface antigen (HBsAg) loss, defined as functional cure, is a clinical target in patients with chronic hepatitis B (CH). To understand the immune responses underlying functional cure, we evaluated cytokine and chemokine expression profiles from patients with resolving and nonresolving acute hepatitis B (AH). METHODS. We cross-sectionally evaluated 41 chemokines and cytokines at the peak of hepatitis in the sera from 41 self-limited AH patients who achieved HBsAg seroconversion, 8 AH patients who failed to clear HBsAg within 1 year after the diagnosis, 8 CH patients with hepatic flare, and 14 healthy volunteers. We longitudinally examined 41 chemokines and cytokines in the sera from 4 self-limited AH patients, 3 chimpanzees inoculated with hepatitis B virus (HBV), and 2 CH patients treated with nucleotide analogs and PEG–IFN- α , one resulting in functional cure. RESULTS. In AH patients and HBV-inoculated chimpanzees with HBsAg loss, CXCL9, CXCL10, CXCL11, CXCL13, and IL-21 were elevated at hepatitis with subsequent decline of HBsAg. Interestingly, IL-21 elevation was observed only in resolving AH patients but not in nonresolvers. CXCL13 and IL-21 elevation was not observed in CH patients who failed to attain HBsAg loss, even at hepatic flare. A concomitant increase of CXCL13 and IL-21 was significant in CH patients who attained HBsAg seroconversion with a sequential therapy. CONCLUSION. Elevation of serum CXCL9, CXCL10, CXCL11, CXCL13, and IL-21 might be a hallmark of functional cure of AH or CH patients.

BCL2 family proteins including pro-survival proteins, BH3-only proteins and BAX/BAK proteins control mitochondria-mediated apoptosis to maintain cell homeostasis via the removal of damaged cells and pathogen-infected cells. In this study, we examined the roles of BCL2 proteins in the induction of apoptosis in cells upon infection with flaviviruses, such as Japanese encephalitis virus, Dengue virus and Zika virus. We showed that survival of the infected cells depends on BCLX_L, a pro-survival BCL2 protein due to suppression of the expression of another pro-survival protein, MCL1. Treatment with BCLX_L inhibitors, as well as deficient BCLX_L gene expression, induced BAX/BAK-dependent apoptosis upon infection with flaviviruses. Flavivirus infection attenuates cellular protein synthesis, which confers reduction of short-half-life proteins like MCL1. Inhibition of BCLX_L increased phagocytosis of virus-infected cells by macrophages, thereby suppressing viral dissemination and chemokine production. Furthermore, we examined the roles of BCLX_L in the death of JEV-infected cells during in vivo infection. Haploinsufficiency of the BCLX_L gene, as well as administration of BH3 mimetic compounds, increased survival rate after challenge of JEV infection and suppressed inflammation. These results suggest that BCLX_L plays a crucial role in the survival of cells infected with flaviviruses, and that BCLX_L may provide a novel antiviral target to suppress propagation of the family of Flaviviridae viruses. The genus Flavivirus including Japanese encephalitis virus, Dengue virus, and Zika virus all of which are mosquito-borne human pathogen and cause serious diseases in humans. Therefore, the development of effective vaccines and antivirals against several flaviviruses is still needed. BCL2 family proteins control mitochondria-mediated apoptosis to maintain cell homeostasis via the removal of damaged cells and pathogen-infected cells, deregulation of which leads to severe diseases including cancer and autoimmune diseases. Here, we showed that BCLX_L is a critical cell survival factor during infection with flaviviruses, and that inhibition of BCLX_L by treatment with BH3 mimetics restricts the production of infectious particles and the expression of chemokines in vitro and in vivo. Inhibition of BCLX_L induces apoptosis in cells infected with flaviviruses and these cells are quickly removed by engulfment of phagocytes, which leads to inhibition of virus dissemination without any inflammatory reaction. Based on these data, BCLX_L would appear to be a suitable target for the development of novel antivirals against a broad range of flavivirus infections.

Frailty including physical inactivity is associated with the survival of patients with hepatocellular carcinoma (HCC). We aimed to investigate the effects of in-hospital exercise on frailty in patients with HCC. This was a multi-center observational study. Patients with HCC were classified into exercise (n = 114) and non-exercise (n = 67) groups. The exercise group was treated with a mixture of aerobic and resistance exercises (20–40 min/day, median four days). Frailty was assessed using the liver frailty index (LFI). Factors for changes in LFI were examined by multivariate and decision-tree analyses. The factors were also examined after propensity score matching. During hospitalization, LFI was significantly improved in the exercise group compared to the non-exercise group (ΔLFI −0.17 vs. −0.02, p = 0.0119). In multivariate analysis, exercise (odds ratio (OR) 2.38, 95% confidence interval (CI) 1.240–4.570, p = 0.0091) and females (OR 2.09; 95%CI, 1.062–4.109; p = 0.0328) were identified as independent factors for the improvement of LFI. In the decision-tree analysis, exercise was identified as an initial classifier associated with the improvement of LFI. Similar findings were also seen in the propensity score matching analyses. We demonstrated that in-hospital exercise improved frailty in patients with HCC. Thus, in-hospital exercise may be beneficial for improving physical function in patients with HCC.

Non-alcoholic fatty liver disease (NAFLD) is a progressive disorder that can develop into liver fibrosis and hepatocellular carcinoma. Natural killer (NK) cells have been shown to protect against liver fibrosis and tumorigenesis, suggesting that they may also play a role in the pathogenesis of NAFLD. Sialic acid-binding immunoglobulin-like lectins (Siglecs) are a family of inhibitory and activating receptors expressed by many cell types, including NK cells. Here, we investigated the phenotypic profiles of peripheral blood and intrahepatic NK cells, including expression of Siglecs and immune checkpoint molecules, and their association with NK cell function in patients with NAFLD. Immune cells in the peripheral blood of 42 patients with biopsy-proven NAFLD and 13 healthy volunteers (HVs) were identified by mass cytometry. The function of various NK cell subpopulations was assessed by flow cytometric detection of intracellular IFN-γ and CD107a/LAMP-1, a degranulation marker, after in vitro stimulation. We found that peripheral blood from NAFLD patients, regardless of fibrosis stage, contained significantly fewer total CD56⁺ NK cell and CD56^dim NK cell populations compared with HVs, and the CD56^dim cells from NAFLD patients were functionally impaired. Among the Siglecs examined, NK cells predominantly expressed Siglec-7 and Siglec-9, and both the expression levels of Siglec-7 and Siglec-9 on NK cells and the frequencies of Siglec-7⁺CD56^dim NK cells were reduced in NAFLD patients. Notably, Siglec-7 levels on CD56^dim NK cells were inversely correlated with PD-1, CD57, and ILT2 levels and positively correlated with NKp30 and NKp46 levels. Further subtyping of NK cells identified a highly dysfunctional Siglec-7⁻CD57⁺PD-1⁺CD56^dim NK cell subset that was increased in patients with NAFLD, even those with mild liver fibrosis. Intrahepatic NK cells from NAFLD patients expressed elevated levels of NKG2D and CD69, suggesting a more activated phenotype than normal liver NK cells. These data identify a close association between NK cell function and expression of Siglec-7, CD57, and PD-1 that could potentially be therapeutically targeted in NAFLD.