Background: A reliable and inexpensive noninvasive marker of hepatic fibrosis is required in patients with nonalcoholic fatty liver disease (NAFLD). FIB4 index (based on age, aspartate aminotransferase [AST] and alanine aminotransferase [ALT] levels, and platelet counts) is expected to be useful for evaluating hepatic fibrosis. We validated the performance of FIB4 index in a Japanese cohort with NAFLD.Methods: The areas under the receiver operating characteristic curves (AUROC) for FIB4 and six other markers were compared, based on data from 576 biopsy-proven NAFLD patients. Advanced fibrosis was defined as stage 3-4 fibrosis. FIB4 index was assessed as: age (yr) × AST (IU/L)/(platelet count (10⁹/L) × √ALT (IU/L))Results: Advanced fibrosis was found in 64 (11%) patients. The AUROC for FIB4 index was superior to those for the other scoring systems for differentiating between advanced and mild fibrosis. Only 6 of 308 patients with a FIB4 index below the proposed low cut-off point (< 1.45) were under-staged, giving a high negative predictive value of 98%. Twenty-eight of 59 patients with a FIB4 index above the high cut-off point (> 3.25) were over-staged, giving a low positive predictive value of 53%. Using these cutoffs, 91% of the 395 patients with FIB-4 values outside 1.45-3.25 would be correctly classified. Implementation of the FIB4 index in the Japanese population would avoid 58% of liver biopsies.Conclusion: The FIB4 index was superior to other tested noninvasive markers of fibrosis in Japanese patients with NAFLD, with a high negative predictive value for excluding advanced fibrosis. The small number of cases of advanced fibrosis in this cohort meant that this study had limited power for validating the high cut-off point.

NK cells can destroy tumor cells without prior sensitization or immunization. Tumors often lose expression of MHC molecules and/or antigens. However, NK cells can lyse tumor cells in a non-MHC-restricted manner and independent of the expression of tumor-associated antigens. NK cells are therefore considered ideal for adoptive cancer immunotherapy; however the difficulty of obtaining large numbers of fully functional NK cells that are safe to administer deters its clinical use. This phase I clinical trial seeks to address this obstacle by first developing a novel system that expands large numbers of highly activated clinical grade NK cells, and second, determining if these cells are safe in a mono-treatment so they can be combined with other reagents in the next round of clinical trials. Patients with unresectable, locally advanced and/or metastatic digestive cancer who did not succeed with standard therapy were enrolled. NK cells were expanded ex vivo by stimulating PBMCs with OK432, IL-2, and modified FN-CH296 induced T cells. Patients were administered autologous natural killer cell three times weekly via intravenous infusions in a dose-escalating manner (dose 0.5 × 109, 1.0 × 109, 2.0 × 109 cells/injection, three patients/one cohort). Total cell population had a median expansion of 586-fold (range 95–1102), with a significantly pure (90.96 %) NK cell population. Consequently, NK cells were expanded to approximately 4720-fold (range 1372–14,116) with cells being highly lytic in vitro and strongly expressing functional markers such as NKG2D and CD16. This NK cell therapy was very well tolerated with no severe adverse events. Although no clinical responses were observed, cytotoxicity of peripheral blood was elevated approximately twofolds up to 4 weeks post the last transfer. We successfully generated large numbers of activated NK cells from small quantities of blood without prior purification of the cells. We also determined that the expanded cells were safe to administer in a monotherapy and are suitable for the next round of clinical trials where their efficacy will be tested combined with other reagents. Trial Registration: UMIN UMIN000007527

The genetic factors affecting the natural history of nonalcoholic fatty liver disease (NAFLD), including the development of nonalcoholic steatohepatitis (NASH) and NASH-derived hepatocellular carcinoma (NASH-HCC), are still unknown. In the current study, we sought to identify genetic factors related to the development of NAFLD, NASH, and NASH-HCC, and to establish risk-estimation models for them. For these purposes, 936 histologically proven NAFLD patients were recruited, and genome-wide association (GWA) studies were conducted for 902, including 476 NASH and 58 NASH-HCC patients, against 7,672 general-population controls. Risk estimations for NAFLD and NASH were then performed using the SNPs identified as having significant associations in the GWA studies. We found that rs2896019 in PNPLA3 [p = 2.3x10^-31, OR (95%CI) = 1.85 (1.67–2.05)], rs1260326 in GCKR [p = 9.6x10^-10, OR (95%CI) = 1.38(1.25–1.53)], and rs4808199 in GATAD2A [p = 2.3x10^-8, OR (95%CI) = 1.37 (1.23–1.53)] were significantly associated with NAFLD. Notably, the number of risk alleles in PNPLA3 and GATAD2A was much higher in Matteoni type 4 (NASH) patients than in type 1, type 2, and type 3 NAFLD patients. In addition, we newly identified rs17007417 in DYSF [p = 5.2x10^-7, OR (95%CI) = 2.74 (1.84–4.06)] as a SNP associated with NASH-HCC. Rs641738 in TMC4, which showed association with NAFLD in patients of European descent, was not replicated in our study (p = 0.73), although the complicated LD pattern in the region suggests the necessity for further investigation. The genetic variants of PNPLA3, GCKR, and GATAD2A were then used to estimate the risk for NAFLD. The obtained Polygenic Risk Scores showed that the risk for NAFLD increased with the accumulation of risk alleles [AUC (95%CI) = 0.65 (0.63–0.67)]. Conclusions: We demonstrated that NASH is genetically and clinically different from the other NAFLD subgroups. We also established risk-estimation models for NAFLD and NASH using multiple genetic markers. These models can be used to improve the accuracy of NAFLD diagnosis and to guide treatment decisions for patients.

Inflammatory processes have an important role in the development of hepatic steatosis and progression to nonalcoholic steatohepatitis (NASH). Interleukin-6 (IL-6) is known to be a proinflammatory cytokine, but also promotes liver regeneration and protects the liver against various forms of damage. The role of IL-6/Glycoprotein130 (GP130) in NASH remains unclear. In this study, we determined whether blocking IL-6/GP130 signaling prevents progression of steatohepatitis in a mouse NASH model. Six-week-old male C57/BL6 mice were fed either chow control or a methionine choline-deficient (MCD) diet for 8 weeks. Half of the MCD diet-fed mice were treated with 15 mg/kg rat anti-mouse IL-6 receptor antibody (MR16-1), intraperitoneally twice weekly, the remainder and chow-fed mice were injected with 15 mg/kg rat IgG as a control. Hepatic steatosis, injury, fibrosis, apoptosis, markers of lipid peroxidation/oxidant stress and IL-6-related gene expressions were evaluated. MR16-1 treatment decreased signal transducer and activator of transcription 3 activities and expression of suppressor of cytokine signaling 3 in MCD diet-treated mouse livers. Although this treatment enhanced intrahepatic lipid accumulation accompanied by increased sterol regulatory element-binding protein 1 and decreased peroxisome proliferator-activated receptor-α expression, elevated plasma alanine aminotransferase levels were improved with decreased plasma free fatty acid levels, lipid peroxidation/oxidant stress and hepatic apoptosis. Blocking IL-6/GP130 signaling by MR16-1 enhanced MCD diet-induced hepatic steatosis, but ameliorated liver injury. These findings suggest that hepatic IL-6 signaling has a protective role against the progression of hepatic steatosis but may enhance liver inflammation.