Evaluation of Fucosylated Haptoglobin and Mac-2 Binding Protein as Serum Biomarkers to Estimate Liver Fibrosis in Patients with Chronic Hepatitis C

<div><p>Fucosylated haptoglobin (Fuc-Hpt) and Mac-2 binding protein (Mac-2 bp) are identified as cancer biomarkers, based on the results from a glyco-proteomic analysis. Recently, we reported that these glyco-biomarkers were associated with liver fibrosis and/or ballooning hepatocytes in patients with nonalcoholic fatty liver disease (NAFLD). We evaluated the ability of these glycoproteins to estimate liver fibrosis in 317 patients with chronic hepatitis C. We measured the serum Fuc-Hpt and Mac-2 bp levels using a lectin-antibody ELISA and ELISA, respectively. The serum levels of both Fuc-Hpt and Mac-2 bp increased with the progression of liver fibrosis. The multivariate analysis revealed that Mac-2 bp was an independent factor associated with moderate liver fibrosis (F ≥ 2). In contrast, Fuc-Hpt was an independent factor associated with advanced liver fibrosis (F ≥ 3). In terms of evaluating liver fibrosis, the serum levels of these glycomarkers were correlated with well-known liver fibrosis indexes, such as the aspartate aminotransferase to platelet ratio index (APRI) and Fibrosis-4 (FIB4) index. An assay that combined the APRI or FIB4 index and the Fuc-Hpt or Mac-2 bp levels increased the AUC value for diagnosing hepatic fibrosis. Interestingly, the cumulative incidence of hepatocellular carcinoma (HCC) was significantly higher in the patients with elevated serum levels of Fuc-Hpt and Mac-2 bp. In conclusion, both Fuc-Hpt and Mac-2 bp could be useful glyco-biomarkers of liver fibrosis and predictors of HCC in patients with chronic hepatitis C.</p></div

Clinical course of the 317 patients after liver biposy.

<p>Clinical course of the 317 patients after liver biposy.</p

Factors associated with advanced liver fibrosis (F ≥ 3).

<p>Factors associated with advanced liver fibrosis (F ≥ 3).</p

The diagnostic abilities of the serum Fuc-Hpt and Mac-2 bp levels for assessing the stage of liver fibrosis.

<p>The ROC curves and AUROC values of serum Fuc-Hpt and Mac-2 bp in F ≥ 2, F ≥ 3 and F4 stage liver fibrosis were shown in the figure.</p

Box-and-whisker plot of the serum Fuc-Hpt (A) and Mac-2 bp (B) levels for each liver fibrosis stage.

<p>The top and bottom of each box represent the first and third quartiles, respectively, with the height of the box representing the interquartile range, covering 50% of the values. The line across each box represents the median. The error bars show the minimum and maximum values. The statistical analysis was performed with the Wilcoxon rank sum test. * P < 0.05. ** P < 0.001.</p

The diagnostic abilities of combinations with these glycoproteins.

<p>The diagnostic abilities of combinations with these glycoproteins.</p

Factors associated with moderate liver fibrosis (F ≥ 2).

<p>Factors associated with moderate liver fibrosis (F ≥ 2).</p

The correlation coefficients between each of the liver fibrosis markers.

<p>The correlation coefficients between each of the liver fibrosis markers.</p

The cumulative hepatocellular carcinoma incidence rate.

<p>The median follow-up period after liver biopsy was 2.4 years (Interquartile range; 1.0–4.9 years). HCC was detected in 19 patients during the follow-up period. The incidence rates at 1, 3 and 5 years in all patients were 1.8%, 4.3% and 9.2%, respectively, using the Kaplan-Meier method. (A) We divided these findings into two groups by the liver histology grade, F0-1 and F2-4, and the median (B) Fuc-Hpt (C) and Mac-2 bp (D) levels. These analysis were performed using the log-rank test.</p

Frequency and role of NKp46 and NKG2A in hepatitis B virus infection

<div><p>Background and Aim</p><p>Natural Killer (NK) cells are involved in the control of viral infection. However, the role of NK cells in chronic hepatitis B (CHB) remains unclear. This study investigated the frequencies and roles of NK cells in CHB, with a focus on activating receptor NKp46 and inhibitory receptor NKG2A.</p><p>Patients/Method</p><p>Peripheral blood lymphocytes were obtained from 71 CHB patients and 37 healthy subjects (HS). The expressions of NKp46 and NKG2A were analyzed using flow cytometry. The role of NKp46-ligand was assessed using an in vitro co-culture system. Cytotoxicity and IFN-γ production in NK cells were evaluated using RT-PCR and flow cytometry.</p><p>Results</p><p>CHB patients were classified into treatment-naïve patients with low HBV DNA titer (CHB-L; n = 28), high HBV DNA titer (CHB-H; n = 24) by the cut-off level of serum HBV DNA 4 log copies/ml, and patients receiving nucleos(t)ide analogue (CHB-NA; n = 19). The expressions of NKp46 and NKG2A were higher in CHB-H than in HS/CHB-L/CHB-NA. HepG2.2.15 had higher NKp46-ligand expression than HepG2. When NK cells from HS were co-cultured with HepG2.2.15, inhibition of the NKp46 and NKp46-ligand interaction by anti-NKp46 antibody significantly reduced cytolysis of HepG2.2.15 and IFN-γ production. However, those reductions were not observed in co-culture with HepG2. Additionally, NK cells that highly expressed NKp46 also highly expressed NKG2A (NKp46^highNKG2A^high subset). The frequencies of NKp46^highNKG2A^high subset in CHB-H were higher than those in HS/CHB-L/CHB-NA. Among treatment-naïve CHB patients, the frequencies of NKp46^highNKG2A^high subset were positively correlated with serum ALT (P<0.01, r = 0.45) and HBV DNA (P<0.01, r = 0.59) levels. The expressions of Fas-L, STAT1, TRAIL and CD107a were higher and IFN-γ expression was lower in the NKp46^highNKG2A^high subset than in the other subsets.</p><p>Conclusion</p><p>The NKp46 and NKp46-ligand interaction contributes to NK cell activation. A novel NK cell subset, the NKp46^highNKG2A^high subset, may be associated with liver injury and HBV replication.</p></div