Kung-Hao Liang

Automated biphasic morphological assessment of hepatitis B-related liver fibrosis using second harmonic generation microscopy

Liver fibrosis assessment by biopsy and conventional staining scores is based on histopathological criteria. Variations in sample preparation and the use of semi-quantitative histopathological methods commonly result in discrepancies between medical centers. Thus, minor changes in liver fibrosis might be overlooked in multi-center clinical trials, leading to statistically non-significant data. Here, we developed a computer-assisted, fully automated, staining-free method for hepatitis B-related liver fibrosis assessment. In total, 175 liver biopsies were divided into training (n = 105) and verification (n = 70) cohorts. Collagen was observed using second harmonic generation (SHG) microscopy without prior staining, and hepatocyte morphology was recorded using two-photon excitation fluorescence (TPEF) microscopy. The training cohort was utilized to establish a quantification algorithm. Eleven of 19 computer-recognizable SHG/TPEF microscopic morphological features were significantly correlated with the ISHAK fibrosis stages (P < 0.001). A biphasic scoring method was applied, combining support vector machine and multivariate generalized linear models to assess the early and late stages of fibrosis, respectively, based on these parameters. The verification cohort was used to verify the scoring method, and the area under the receiver operating characteristic curve was >0.82 for liver cirrhosis detection. Since no subjective gradings are needed, interobserver discrepancies could be avoided using this fully automated method.

Resistin reinforces interferon λ-3 to eliminate hepatitis C virus with fine-tuning from RETN single-nucleotide polymorphisms.

The effect of resistin (RETN) on the response to anti-HCV therapy remains unclear. A prospective cohort study was performed using 655 consecutive HCV patients, of whom 513 had completed a course of interferon-based therapy. Multivariate and GEE analyses revealed four RETN single-nucleotide polymorphisms (SNPs), rs34861192, rs3219175, rs3745367 and rs1423096, to be synergistically associated with resistin levels. After adjusting for co-factors such as interferon λ-3 (IFNL3)-rs12979860, the resistin level and the hyper-resistinemic genotype at the 4 RETN SNPs were positively and negatively associated with a sustained virological response (SVR), respectively. RETN-rs3745367 was in linkage disequilibrium with IFNL3-rs12979860. Compared to non-SVR patients, SVR patients had higher levels of pre-therapy resistin, primarily originating from intrahepatic lymphocytes, stellate cells, Kupffer cells, hepatic progenitor cells and hepatocytes. This difference diminished over the course of therapy, as only SVR patients exhibited a 24-week post-therapy decrease in resistin. Both resistin and IFNL3 mRNAs were upregulated, but only resistin mRNA was upregulated by recombinant resistin in peripheral blood mononuclear cells with and without hyper-resistinemic genotypes of the 4 RETN SNPs, respectively. Fine-tuned by RETN SNPs, intrahepatic, multi-cellular resistin reinforced IFNL3 in eliminating HCV via immunomodulation to counteract pro-inflammation. These results encourage the development of novel resistin-targeted anti-viral agents.

Diabetes, hepatocellular carcinoma, and mortality in hepatitis C-infected patients: a population-based cohort study

The effect of diabetes mellitus (DM) on the development of hepatocellular carcinoma (HCC) and all‐cause mortality after HCC development in chronic hepatitis C virus (HCV)‐infected patients remains inconclusive. This cohort study aimed to investigate these issues using the Taiwanese National Health Insurance Research Database.

A Circulating MicroRNA Signature Capable of Assessing the Risk of Hepatocellular Carcinoma in Cirrhotic Patients

With the availability of potent antiviral therapies, complete suppression of hepatitis B virus (HBV) replication and total eradication of hepatitis C virus (HCV) can now be achieved. Despite these advances, hepatocellular carcinoma (HCC) still develops in a substantial proportion of cirrhotic patients, suggesting that host factors remain critical. Dysregulation of miRNAs is noted in many cancers, and circulating miRNAs can be readily assayed. In this study, we aimed to develop a circulating miRNA signature to assess the risk of HCC in cirrhotic patients. We first discovered that HBV- and HCV-related cirrhotic patients had distinguishable circulating miRNA profiles. A cohort of 330 cirrhotic patients was then compared against a cohort of 42 early HCC patients with complete remission. A score comprising 5 miRNAs and a binary etiology variable was established that was capable of differentiating between these two groups (AUC = 72.5%, P < 0.001). The 330 cirrhotic patients were further stratified into high- and low-risk groups, and all patients were longitudinally followed for 752 (11–891) days. Of them, 19 patients developed HCC. The high-risk group had significantly higher cumulative HCC incidence (P = 0.038). In summary, a circulating miRNA-based score was developed that is capable of assessing HCC risks in cirrhotic patients.

UGT2B28 genomic variation is associated with hepatitis B e-antigen seroconversion in response to antiviral therapy.

Seroconversion of hepatitis B virus (HBV) e-antigen (HBeAg) is a critical but often-missed therapeutic goal in standard antiviral treatments. An extreme-phenotype genome-wide association study was performed, comparing untreated spontaneous recoverers (with seroconversion of HBV surface antigen) versus entecavir-treated patients failing to achieve HBeAg seroconversion. A single-nucleotide-polymorphism rs2132039 on the UGT2B28 gene, alongside an adjacent copy number polymorphism (CNP605), manifested the strongest clinical associations (P = 3.4 × 10−8 and 0.001, respectively). Multivariate analysis showed that rs2132039-TT genotypes, but not CNP605 copy numbers, remained associated to spontaneous recoverers (P = 0.009). The clinical association of rs2132039 was validated successfully in an independent cohort (n = 302; P = 0.002). Longitudinal case-only analyses revealed that the rs2132039-TT genotype predicted shorter time-to-HBeAg-seroconversion in all antiviral-treated patients (n = 380, P = 0.012), as well as the peginterferon-treated subgroup (n = 123; P = 0.024, Hazard ratio [HR] = 2.104, Confidence interval [CI] = 1.105–4.007). In the entecavir-treated subgroup, the predictive effect was restricted by pretreatment alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, with effective prediction observed in patients with ALT < 200 IU/ml and ALT/AST ratio <2 (n = 132; P = 0.013, HR = 10.538, CI = 1.420–78.196).

Combinations of single nucleotide polymorphisms WWOX‐rs13338697, GALNT14‐rs9679162 and rs6025211 effectively stratify outcomes of chemotherapy in advanced hepatocellular carcinoma

A genome‐wide association study (GWAS) had identified a single nucleotide polymorphism (SNP), GALNT14‐rs9679162, capable of predicting chemotherapy responses in advanced hepatocellular carcinoma (HCC). Here, we revisited the GWAS database to search for necessary SNPs that could improve our outcome prediction.

Platelet counts modulate the quantitative relationship between hepatitis B viral DNA and surface antigen concentrations: a cross-sectional study of hematological, histological and viral factors

AbstractBackgroundThe concentrations of hepatitis B virus (HBV) DNA and surface antigen (HBsAg) are two critical virological variables to be monitored in chronic hepatitis B. HBsAg is derived from the HBV genome. Thus, higher HBV-DNA concentrations should implicate higher HBsAg levels. Nevertheless, the two variables do not manifest a simple linear relationship due to elusive host factor involvements. The aim of this study was to address the discrepancy of HBV DNA and HBsAg levels by a quantitative modeling of HBsAg concentrations.MethodsPretreatment hematological, histological and virus serological records of 327 chronic hepatitis B patients were reviewed. Two independent patient cohorts were used for validation.ResultsUnivariate/multivariate analysis showed that ISHAK fibrosis stages, HBV-DNA levels and hepatitis e-antigen status were independently associated with HBsAg concentrations. In agreement with the natural history of chronic hepatitis B, HBsAg concentrations were negatively correlated with ISHAK fibrosis stages (adjusted P = 0.002). Subgroup analysis showed that significant HBsAg-DNA correlation existed in high-viral-titer patients with HBV-DNA > 6 log10 IU/mL (P < 0.001), but not in low-viral-titer patients with HBV-DNA ≤ 6 log10 IU/mL (P = 0.076). A backward stepwise linear regression analysis in the low-viral-titer subgroup revealed a significant correlation between HBsAg levels and a linear combination of HBV-DNA levels and platelet counts. A biphasic model was thus established to accommodate patients with high and low HBV-DNA titers:HBsAg=0.538∗HBV-DNA+0.001∗platelet∗(|6-HBV-DNA|+6-HBVDNA)-0.321

Phenotypic and Genotypic Shifts in Hepatitis B Virus in Treatment-Naive Patients, Taiwan, 2008–2012

\mathrm{HB}\mathrm{sAg}=0.538\ast \mathrm{H}\mathrm{B}\mathrm{V}{\textstyle \hbox{-}}\mathrm{D}\mathrm{N}\mathrm{A}+0.001\ast \mathrm{platelet}\ast \left(\left|6{\textstyle \hbox{-}}\mathrm{H}\mathrm{B}\mathrm{V}{\textstyle \hbox{-}}\mathrm{D}\mathrm{N}\mathrm{A}\right|+6{\textstyle \hbox{-}}\mathrm{HB}\mathrm{V}\mathrm{D}\mathrm{N}\mathrm{A}\right){\textstyle \hbox{-} }0.321