Squamous cell carcinoma involving the midline of the dorsum of the tongue: a case report.

Introduction: Most cases of squamous cell carcinoma (SCC) of the tongue occur on the lateral surface; however, SCC of the dorsum is extremely rare. Case presentation: The authors describe the case of a 79-year-old man with SCC involving the midline of the dorsum of the tongue. The lesion was surgically resected. The patient was followed up for 1 year and 6 months, and no recurrence was noted. Discussion: SCC of the dorsal midline is even rarer and accounts for less than 1% of tongue carcinomas. SCC involving the dorsum may have a worse prognosis than SCC of the lateral or ventral surface. This report is the first to use submental flap reconstruction to treat cancer of the midline dorsum of the tongue. Conclusion: The authors encountered a case of SCC involving the midline of the dorsum of the tongue, which has rarely been reported in the literature. The authors attained a favorable outcome through surgical intervention.

Immunoglobulin-like transcript 2 as an impaired anti-tumor cytotoxicity marker of natural killer cells in patients with hepatocellular carcinoma.

Introduction: Natural killer (NK) cells play a pivotal role in immune surveillance in the liver. We aimed to identify potential targets for NK cell-mediated immune intervention by revealing the functional molecules on NK cells in HCC patients. Methods: To evaluate the impact of aging on NK cell phenotypes, we examined NK cells from healthy volunteers (HVs) of various ages. Because ILT2 expression on CD56dim NK cells increased with increasing age, we enrolled age-matched HCC patients and HVs. We determined the NK cell phenotypes in blood mononuclear cells (PBMCs) and intrahepatic lymphocytes (IHLs) from cancerous and non-cancerous tissues. We evaluated cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC) of NK cells in vitro. Results: ILT2-positive CD56dim NK cells in PBMCs were increased in HCC patients compared with HVs. In HCC patients, ILT2-positive CD56dim NK cells were increased in cancerous IHLs compared with non-cancerous IHLs and PBMCs. We examined the impact of macrophage migration inhibitory factor (MIF) on ILT2 expression in co-cultures of HCC cells and NK cells. The enhanced expression of ILT2 on CD56dim NK cells from HCC patients was inhibited by masking antibodies against MIF and CXCR4. ILT2-positive CD56dim NK cells exhibited lower capacities for cytotoxicity and ADCC than ILT2-negative cells, which were partially restored by ILT2 blockade. Conclusions: In HCC patients, ILT2 is a signature molecule for cancerous CD56dim NK cells with impaired cytolytic capacity. The MIF-CXCR4 interaction is associated with ILT2 induction on CD56dim NK cells and ILT2 serves as a target for functional NK cell restoration.

Differences in branched-chain amino acid to tyrosine ratio (BTR) among etiologies of chronic liver disease progression compared to healthy adults.

BACKGROUND: The branched-chain amino acids (BCAAs) to tyrosine (Tyr) ratio (BTR) test is used to evaluate the progression of chronic liver disease (CLD). However, the differences across sex, age, body mass index (BMI) and etiologies are still unclear. METHODS: We retrospectively reviewed data from 2,529 CLD cases with free amino acids (FAAs) in peripheral blood from four hospitals and 16,421 general adults with FAAs data from a biobank database. In total, 1,326 patients with CLD (covering seven etiologies) and 8,086 healthy controls (HCs) were analyzed after exclusion criteria. We investigated the change of BTR in HCs by sex, age and BMI and then compared these to patients divided by modified ALBI (mALBI) grade after propensity score matching. RESULTS: BTR is significantly higher in males than females regardless of age or BMI and decreases with aging in HCs. In 20 types of FAAs, 7 FAAs including BCAAs were significantly decreased, and 11 FAAs including Tyr were significantly increased by mALBI grade in total CLD. The decreasing timings of BTR were at mALBI grade 2b in all CLD etiologies compared to HCs, however in chronic hepatitis C (CHC), chronic hepatitis B (CHB) and alcoholic liver disease (ALD), BTR started to decrease at 2a. There was a positive correlation between BCAAs and albumin among parameters in BTR and mALBI. The correlation coefficients in PBC, ALD and MASLD were higher than those of other etiologies. CONCLUSIONS: BTR varies by sex and age even among healthy adults, and decreasing process and timing of BTR during disease progression is different among CLD etiologies.

Novel frameshift variant of WNT10A in a Japanese patient with hypodontia.

Congenital tooth agenesis is caused by the impairment of crucial genes related to tooth development, such as Wnt signaling pathway genes. Here, we investigated the genetic causes of sporadic congenital tooth agenesis. Exome sequencing, followed by Sanger sequencing, identified a novel single-nucleotide deletion in WNT10A (NC_000002.12(NM_025216.3):c.802del), which was not found in the healthy parents of the patient. Thus, we concluded that the variant was the genetic cause of the patient's agenesis.

Toll-like receptor 7 agonist, GS-986, is an immune-stimulant inducing follicular helper T cells and expanding HBs antigen-specific B cells in vitro.

BACKGROUNDS AND AIMS: Toll-like receptor (TLR) agonists have been developed as adjuvants to efficiently induce antiviral immune responses. Specificity and potency of these compounds are essential requirements for clinical trial applications. In patients with hepatitis B virus (HBV) infections, sustained loss of hepatitis B surface antigen (HBsAg) is a therapeutic goal, which may be achievable by the sequential activation of follicular helper T cells (Tfh) and antibody-secreting B cells. We aimed to elucidate whether novel TLR7 agonist, GS-986, could activate immune responses involved in HBV elimination. METHODS: To clarify the impact of GS-986 on pDCs, we quantified the expression levels of surface markers and evaluated for Tfh induction in a culture model consisting of human pDCs with allogeneic naïve CD4+ T cells. In addition, we examined whether GS-986 could enhance HBs antibody production capacity using PBMC from CHB patients. RESULTS: pDCs from CHB patients had lower OX40L expression and as well as impaired capacity for Tfh induction compared with those from healthy donors. However, GS-986-stimulated pDCs from CHB patients expressed OX40L and produced IL-6 and IL-12, resulting in the induction of IL-21-producing Tfh cells (CXCR5+ PD-1+ CD4+ ) from naïve CD4+ T cells. The Tfh-inducing capacity of GS-986 was reduced in the presence of an anti-OX40L blocking antibody. Furthermore, GS-986 promoted HBsAg-specific antibody production in PBMCs from CHB patients. CONCLUSIONS: GS-986 is an adjuvant that stimulates pDCs to induce Tfh differentiation and antigen-specific B-cell production. This immune profile may be beneficial for therapeutic application as an immune modulator in CHB patients.

Effects of sodium glucose cotransporter 2 inhibitors and pioglitazone on FIB-4 index in metabolic-associated fatty liver disease.

BACKGROUND: The antidiabetic drugs sodium glucose cotransporter 2 inhibitors (SGLT2is) and thiazolidinediones have beneficial effects on the liver dysfunction of patients with nonalcoholic fatty liver disease and type 2 diabetes mellitus (T2DM). We aimed to determine the efficacy of these drugs for the treatment of liver disease in patients with metabolic dysfunction-associated fatty liver disease (MAFLD) and T2DM. METHODS: We undertook a retrospective study of 568 patients with MAFLD and T2DM. Of these, 210 were treating their T2DM with SGLT2is (n = 95), 86 with pioglitazone (PIO), and 29 with both. The primary outcome was the change in Fibrosis-4 (FIB-4) index between baseline and 96 weeks. RESULTS: At 96 weeks, the mean FIB-4 index had significantly decreased (from 1.79 ± 1.10-1.56 ± 0.75) in the SGLT2i group, but not in the PIO group. The aspartate aminotransferase to platelet ratio index, serum aspartate and alanine aminotransferase (ALT), hemoglobin A1c, and fasting blood sugar significantly decreased in both groups (ALT: SGLT2i group, -17 ± 3 IU/L; PIO group, -14 ± 3 IU/L). The bodyweight of the SGLT2i group decreased, but that of the PIO group increased (-3.2 kg and +1.7 kg, respectively). When the participants were allocated to two groups according to their baseline ALT (>30 IU/L), FIB-4 index significantly decreased in both groups. In patients taking pioglitazone, the addition of SGLT2i improved liver enzymes but not FIB-4 index for 96 weeks. CONCLUSIONS: Treatment with SGLT2i causes a larger improvement in FIB-4 index than PIO in patients with MAFLD over 96 weeks.

Novel WNT10A variant in a Japanese case of nonsyndromic oligodontia.

Congenital tooth agenesis is one of the most common anomalies in humans. Many genetic factors are involved in tooth development, including MSX1, PAX9, WNT10A, and LRP6. Thus, mutations in these genes can cause congenital tooth agenesis in humans. In this study, we identified a novel nonsense WNT10A variant, NM_025216.3(WNT10A_v001):c.1090A > T, which produces a C-terminal truncated gene product, p.(Lys364*), in a sporadic form of congenital tooth agenesis. The variant was not found in the healthy parents and thus was considered to cause congenital tooth agenesis in the case.

No increased risk of hepatocellular carcinoma after eradication of hepatitis C virus by direct-acting antivirals, compared with interferon-based therapy.

It is well-known that sustained virological response (SVR) by interferon (IFN)-based therapy against hepatitis C virus (HCV) infection reduced the incidence of hepatocellular carcinoma (HCC). However, whether IFN-free direct-acting antivirals reduce the risk of HCC is controversial. Therefore, this study aims to compare the incidence of HCC after the achievement of SVR between sofosbuvir combined with ledipasvir (SOF/LDV) and simeprevir with pegylated interferon plus ribavirin (Sim+IFN). Japanese patients with HCV infection (genotype 1) who achieved SVR between January 2013 and December 2014 by SOF/LDV (NCT01975675, n = 320) or Sim+IFN (000015933, n = 289) therapy in two nationwide, multicenter, phase III studies were prospectively monitored for the development of HCC by ultrasonography for 5 years after the end of treatment (EOT). No HCC was detected before the treatment. HCC was detected in 9 and 7 patients in the SOF/LDV and the Sim+IFN group in 5 years, respectively. The cumulative incidences of HCC rates 1, 3, and 5 years after EOT were similar between the two groups (1.5%, 2.7%, and 3.2% for the SOF/LDV and 1.8%, 2.8%, and 3.0% for the Sim+IFN group, respectively). No HCC was developed 3.5 years after EOT. Interestingly, a retrospective careful review of imaging taken before therapy revealed hepatic nodules in 50% of HCC patients, suggesting HCC was pre-existed before therapy. In conclusion, we could not find any differences in the incidence of HCC after the HCV eradication between the two therapeutic regimens, suggesting no enhancement of HCC development by DAA.

Novel MSX1 frameshift mutation in a Japanese family with nonsyndromic oligodontia.

Congenital tooth agenesis is a common anomaly in humans. We investigated the etiology of human tooth agenesis by exome analysis in Japanese patients, and found a previously undescribed heterozygous deletion (NM_002448.3(MSX1_v001):c.433_449del) in the first exon of the MSX1 gene. The deletion leads to a frameshift and generates a premature termination codon. The truncated form of MSX1, namely, p.(Trp145Leufs*24) lacks the homeodomain, which is crucial for transcription factor function.

A novel LRP6 variant in a Japanese family with oligodontia.

Congenital tooth agenesis is a common anomaly in human development. We performed exome sequence analysis of genomic DNA collected from Japanese patients with tooth agenesis and their relatives. We found a novel single-nucleotide insertion in the LRP6 gene, the product of which is involved in Wnt/ß-catenin signaling as a coreceptor for Wnt ligands. The single-nucleotide insertion results in a premature stop codon in the extracellular region of the encoded protein.

Increased Frequency of Dysfunctional Siglec-7-CD57+PD-1+ Natural Killer Cells in Patients With Non-alcoholic Fatty Liver Disease.

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 CD56dim NK cell populations compared with HVs, and the CD56dim 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+CD56dim NK cells were reduced in NAFLD patients. Notably, Siglec-7 levels on CD56dim 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+CD56dim 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.

Blood angiopoietin-2 predicts liver angiogenesis and fibrosis in hepatitis C patients.

BACKGROUND: Pathological angiogenesis is involved in the development of hepatocellular carcinoma. In patients with chronic hepatitis C (CHC), the level of angiogenic factor angiopoietin (ANGP)-2 is reported to be increased in the blood, correlating with fibrosis. In this study, we aimed to clarify whether blood ANGP-2 is useful as a biomarker for liver angiogenesis and fibrosis in CHC patients and to further reveal the relationship between such pathology in a carbon tetrachloride (CCl4)-treated liver fibrosis mouse model. METHODS: Plasma levels of ANGP-2, expression of a liver sinusoidal endothelial cell (LSEC) marker (CD31), collagen deposition (Sirius Red staining) in the liver, clinical fibrosis markers (Mac-2 binding protein glycosylation isomer, virtual touch quantification, and liver stiffness measurement), and liver function (albumin bilirubin score) were examined in CHC patients. To determine the effects of an anti-angiogenic agent on liver fibrosis in vivo, sorafenib was administered to the CCl4-treated mice (BALB/c male). RESULTS: The plasma levels of ANGP-2 were increased in CHC patients compared to healthy volunteers and decreased by the eradication of hepatitis C with direct-acting antivirals. In addition, plasma ANGP-2 levels were correlated with CD31 expression, collagen deposition, clinical fibrosis markers, and liver function. Sorafenib inhibited liver angiogenesis and fibrosis in the CCl4-treated mice and was accompanied by decreased ANGP-2 expression in LSECs. CONCLUSIONS: ANGP-2 may serve as a useful biomarker for liver angiogenesis and fibrosis in CHC patients. In addition, angiogenesis and fibrosis may be closely related.

Serum soluble sialic acid-binding immunoglobulin-like lectin-7 concentration as an indicator of liver macrophage activation and advanced fibrosis in patients with non-alcoholic fatty liver disease.

AIM: Non-alcoholic fatty liver disease (NAFLD) is a leading cause of liver disease worldwide. Because liver fibrosis is associated with the long-term prognosis of patients with NAFLD, there is an urgent need for non-invasive markers of liver fibrosis. Sialic acid-binding immunoglobulin-like lectin-7 (Siglec-7) is an immunomodulatory molecule expressed on various immune cells, including macrophages, which plays a key role in liver inflammation and fibrosis in NAFLD. We aimed to determine whether serum levels of soluble Siglec-7 (sSiglec-7) could have utility at a marker of fibrosis in this patient population. METHODS: We examined serum samples from 93 NAFLD patients and 19 healthy donors for macrophage-associated protein, including sSiglec-7, soluble CD163, and YKL-40, and examined their correlation with liver fibrosis scores, tissue elastography, and histological findings. Independent factors associated with advanced fibrosis were analyzed using a logistic regression model and a decision tree. To clarify the source of sSiglec-7, we examined its expression in liver tissue-derived macrophages and cultured monocyte-derived macrophages. RESULTS: Serum sSiglec-7 levels were significantly higher in NAFLD patients compared with healthy donors, and correlated positively with sCD163 and YKL-40 levels. Serum sSiglec-7 was an independent diagnostic marker with high specificity (96.3%) for advanced fibrosis (F3 and F4) in NAFLD patients. Siglec-7 was mainly expressed on CCR2+ macrophages in the liver, and sSiglec-7 production by monocyte-derived macrophages in vitro was increased after stimulation by pro-inflammatory factors. CONCLUSIONS: Elevated serum sSiglec-7 could serve as an independent marker with high specificity for advanced liver fibrosis in patients with NAFLD.

Immune Determinants in the Acquisition and Maintenance of Antibody to Hepatitis B Surface Antigen in Adults After First-Time Hepatitis B Vaccination.

Global implementation of a birth-dose hepatitis B (HB) vaccine has significantly reduced the prevalence of hepatitis B virus (HBV) carriers. Durable and sufficient titers of antibodies to hepatitis B surface antigen (anti-HBs) are desirable for vaccinees to gain resistance to HBV exposure. However, the existence of primary nonresponders and vaccinees who lost anti-HBs over time remains a challenge for the strategy of HBV elimination. We thus aim to clarify the mechanisms of acquisition and maintenance of vaccine-induced anti-HBs in healthy adults. We retrospectively analyzed the vaccination records of 3,755 first-time HB-vaccinated students and also traced the acquired antibody transition of 392 first-time vaccinees for 10 consecutive years. To understand the cellular and humoral immune response, we prospectively examined peripheral blood from 47 healthy first-time HB-vaccinated students, 62 booster-vaccinated health care workers, and 20 individuals who maintained their anti-HBs. In responders, a significant increase of follicular helper T (Tfh) cells, activated plasmablasts, and plasma cells was observed in first-time-vaccinated but not booster-vaccinated persons. We also discovered memory B cells and antibody-secreting cells were more abundant in individuals who maintained anti-HBs. According to vaccination records, higher anti-HBs antibody titer acquisition was related to the longer term maintenance of anti-HBs, the level of which was positively correlated with prevaccination levels of serum interferon-γ and related chemokines. The second series of vaccination as a booster provided significantly higher anti-HBs antibody titers compared to the initial series. Conclusion: Coordinated activation of Tfh and B-cell lineages after HB vaccination is involved in the acquisition and maintenance of anti-HBs. Our findings support the rationale of preconditioning the immune status of recipients to ensure durable vaccine responses.

Bone morphogenetic protein 4 provides cancer-supportive phenotypes to liver fibroblasts in patients with hepatocellular carcinoma.

BACKGROUND: Cancer-associated fibroblasts (CAFs) are essential constituents of cancer-supportive microenvironments. The high incidence of hepatocellular carcinoma (HCC) in advanced fibrosis patients implies that fibroblasts have a promoting effect on HCC development. We aimed to explore the regulators of phenotypes and function of CAFs in the liver. METHODS: We established primary cancer-associated fibroblasts (CAFs) and non-cancerous liver fibroblasts (NFs) from 15 patients who underwent HCC resection. We compared phenotypes, capacity of cytokine/chemokine production and gene expression profiles between pairs of CAFs and NFs from the same donors. We examined resected tissue from additional 50 patients with HCC for immunohistochemical analyses. RESULTS: The CAFs expressed more ACTA2 and COL1A1 than the NFs, suggesting that CAFs are more activated phenotype. The CAFs produced larger amounts of IL-6, IL-8 and CCL2 than the NFs, which led to invasiveness of HuH7 in vitro. We found that Bone Morphogenetic Protein-4 (BMP4) is up-regulated in CAFs compared to NFs. The CAF phenotype and function were gained by BMP4 over-expression or recombinant BMP4 given to fibroblasts, all of which decreased with BMP4 knockdown. In tissues obtained from the patients, BMP4-positive cells are mainly observed in encapsulated fibrous lesions and HCC. Positive expression of BMP4 in HCC in resected tissues, not in fibroblasts, was associated with poorer postoperative overall survival in patients with HCC. CONCLUSION: Endogenous and exogenous BMP4 activate liver fibroblasts to gain capacity of secreting cytokines and enhancing invasiveness of cancer cells in the liver. BMP4 is one of the regulatory factors of CAFs functioning in the microenvironment of HCC.

Pro-angiogenic TIE-2-expressing monocytes/TEMs as a biomarker of the effect of sorafenib in patients with advanced hepatocellular carcinoma.

Sorafenib, a multi-kinase inhibitor, inhibits tumor angiogenesis and is the first-line systemic therapy for patients with advanced hepatocellular carcinoma (HCC). However, due to its limited effects and frequent occurrence of side effects, biomarkers are needed to predict the effects of sorafenib. We considered the possibility of using TIE-2-expressing monocytes (TEMs) to predict the response in sorafenib-treated patients with advanced HCC. TEMs serve as a diagnostic marker of HCC and are related to angiogenesis. We analyzed 25 advanced HCC patients and prospectively evaluated TEMs before (Pre TEMs) and at 1 month after initial therapy (T1m TEMs). The radiologic response was evaluated by modified Response Evaluation Criteria in Solid Tumors (mRECIST). Median survival time (MST) was significantly longer in the partial response/stable disease (PR/SD) group (21.8 months) than in the PD group (8.7 months). ΔTEMs (changes of T1m TEMs compared to Pre TEMs) were significantly lower in the PR/SD group than in the PD group. MST of the ΔTEMs low group (14.2 months) was significantly longer than that of the high group (8.7 months). Univariate and multivariate Cox regression analyses showed that ΔTEMs [hazard ratio (HR) = 8.53, 95% confidence interval (CI) = 1.51-48.16, p = 0.015] and Child-Pugh class (HR = 5.59, 95% CI = 1.06-29.63, p = 0.043) were independently associated with overall survival. Our results suggest that ΔTEMs could serve as a biomarker for predicting radiologic response and overall survival in sorafenib-treated patients with advanced HCC.

The impact of PNPLA3 and JAZF1 on hepatocellular carcinoma in non-viral hepatitis patients with type 2 diabetes mellitus.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is an established independent risk factor for hepatocellular carcinoma (HCC). T2DM is associated with non-alcoholic steatohepatitis (NASH), which is a major cause of non-HBV and non-HCV-related HCC; nevertheless, it has been difficult to identify those patients with T2DM who have a high risk of developing HCC. The aim of this study was to identify genetic determinants that predispose T2DM patients to HCC by genotyping T2DM susceptibility loci and PNPLA3. METHODS: We recruited 389 patients with T2DM who satisfied the following three criteria: negative for HBs-Ag and anti-HCV Ab, alcohol intake <60 g/day, and history of T2DM >10 years. These patients were divided into two groups: T2DM patients with HCC (DM-HCC, n = 59) or those without HCC (DM-non-HCC, n = 330). We genotyped 51 single-nucleotide polymorphisms (SNPs) previously reported as T2DM or NASH susceptibility loci (PNPLA3) compared between the DM-HCC and DM-non-HCC groups with regard to allele frequencies at each SNP. RESULTS: The SNP rs738409 located in PNPLA3 was the greatest risk factor associated with HCC. The frequency of the PNPLA3 G allele was significantly higher among DM-HCC individuals than DM-non-HCC individuals (OR 2.53, p = 1.05 × 10(-5)). Among individuals homozygous for the PNPLA3 G allele (n = 115), the frequency of the JAZF1 rs864745 G allele was significantly higher among DM-HCC individuals than DM-non-HCC individuals (OR 3.44, p = 0.0002). CONCLUSIONS: PNPLA3 and JAZF1 were associated with non-HBV and non-HCV-related HCC development among Japanese patients with T2DM.

Appropriate use of virtual touch quantification and FibroScan M and XL probes according to the skin capsular distance.

BACKGROUND: Appropriate utilization of different diagnostic modalities is essential for the accurate liver stiffness measurements (LSM) in patients with chronic liver diseases. The aim of this study was to evaluate the efficacy of Virtual Touch Quantification (VTQ) and the FibroScan M and XL probes in term of accurate LSM and to identify factors associated with inadequate measurements in obese and non-obese Japanese patients. METHODS: A total of 664 consecutive patients with chronic liver disease were prospectively enrolled. LSM were evaluated concurrently with VTQ and the FibroScan M and XL probes. LSM quality was categorized as inadequate (success rate <60% and/or interquartile range/median value of ≥30%) or adequate. RESULTS: No significant differences in the rate of inadequate LSM were observed among the three diagnostic modalities. In multivariate analysis, skin capsule distance (SCD) was strongly associated with inadequate rates obtained with VTQ and the M probe [odds ratio (OR) 1.28, P < 0.0001 and OR 1.20, P < 0.0001, respectively]. Inadequate LSM rates with both VTQ and the M probe increased with longer SCD, with a significant difference between subgroups at an SCD of ≥22.5 mm (VTQ 54.0%; M probe 51.1%; XL probe 25.2%; P < 0.0001). The rates of inadequate LSM rates with VTQ were significantly lower than those with the XL probe at an SCD of <17.5 mm. A total of 15 liver biopsy specimens obtained from nonalcoholic fatty liver disease patients confirmed the diagnostic accuracy and high applicability of the XL probe. CONCLUSIONS: Long SCD reduced the diagnostic performance of the FibroScan® M probe and VTQ. LSM modalities should be selected according to SCD.

Association of serum IFN-λ3 with inflammatory and fibrosis markers in patients with chronic hepatitis C virus infection.

BACKGROUND: Hepatitis C virus (HCV) is one of the major causes of liver cancer. The single nucleotide polymorphisms within the IFNL3 gene, which encodes interferon (IFN)-λ(3), are strongly associated with the response to pegylated IFN-α (PEG-IFN-α) plus ribavirin (RBV) therapy in chronic hepatitis C (C-CH) patients. However, the roles of IFN-λ(3) in chronic HCV infection are still elusive. In this study, we aimed to identify clinical and immunological factors influencing IFN-λ(3) and evaluated whether serum IFN-λ(3) levels are involved or not involved in the response to PEG-IFN-α plus RBV therapy. METHODS: We enrolled 119 C-CH patients with HCV genotype 1 infection who underwent 48 weeks of PEG-IFN-α plus RBV therapy. As controls, 23 healthy subjects and 56 patients with non-HCV viral hepatitis were examined. Serum IFN-λ(3) was quantified by chemiluminescence enzyme immunoassay, and 27 cytokines or chemokines were assayed by the multiplexed BioPlex system. RESULTS: Serum IFN-λ(3) levels were higher in C-CH patients or acute hepatitis E patients than in healthy volunteers. Such levels did not differ between the IFNL3 genotypes. In C-CH patients, serum IFN-λ(3) was positively correlated with aspartate aminotransferase, alanine aminotransferase, α-fetoprotein, histological activity, fibrosis index, IFN-γ-inducible protein 10, and platelet-derived growth factor. Multivariate analysis showed that IFNL3 single nucleotide polymorphisms, fibrosis score, and macrophage inflammatory protein 1α were involved in the sustained viral clearance in PEG-IFN-α plus RBV therapy; however, serum IFN-λ(3) levels were not involved. CONCLUSION: Serum IFN-λ(3) levels are increased in C-CH patients regardless of the IFNL3 genotype. IFN-λ(3) is a biomarker reflecting the activity and fibrosis of liver disease, but is not correlated with the responsiveness to PEG-IFN-α plus RBV therapy.

Ex vivo induction of IFN-λ3 by a TLR7 agonist determines response to Peg-IFN/ribavirin therapy in chronic hepatitis C patients.

BACKGROUND: Genetic variation around interleukin-28B (IL28B), encoding IFN-λ3, predict non-responders to pegylated interferon-α/ribavirin (Peg-IFN/RBV) therapy in chronic hepatitis C (CHC). However, it remains unclear the expression and the role of IL28B itself. The aim of this study is to develop easy and useful methods for the prediction of treatment outcomes. METHODS: The mRNA and protein levels of IFN-λ3 induced by ex vivo stimulation of peripheral blood mononuclear cells (PBMC) or magnetically selected dendritic cells (DCs) with toll-like receptor agonists (TLR3; poly I:C, TLR7; R-837) were measured by the quantitative real-time polymerase chain reaction and our newly developed chemiluminescence enzyme immunoassays, respectively, and compared with the clinical data. RESULTS: We found that BDCA-4(+) plasmacytoid and BDCA-3(+) myeloid DCs were the main producers of IFN-λs when stimulated with R-837 and poly I:C, respectively. Detectable levels of IFN-λs were inducible even in a small amount of PBMC, and IFN-λ3 was more robustly up-regulated by R-837 in PBMC of CHC patients with favorable genotype for the response to Peg-IFN/RBV (TT in rs8099917) than those with TG/GG. Importantly, the protein levels of IFN-λ3 induced by R-837 clearly differentiated the response to Peg-IFN/RBV treatment (p = 1.0 × 10(-10)), including cases that IL28B genotyping failed to predict the treatment response. The measurement of IFN-λ3 protein more accurately predicted treatment efficacies (95.7 %) than that of IL28B genotyping (65.2 %). CONCLUSIONS: Genetic variations around IL28B basically affect IFN-λ3 production, but different amounts of IFN-λ3 protein determines the outcomes of Peg-IFN/RBV treatment. This study, for the first time, presents compelling evidence that IL28B confer a functional phenotype.