Varied immune responses of HBV-specific B cells in patients undergoing pegylated interferon-alpha treatment for chronic hepatitis B.

BACKGROUND & AIMS: The changes of HBV-specific B-cells in chronic hepatitis B (CHB) patients underwent pegylated interferon-alfa (PEG-IFNα) treatment and achieved functional cure remain unclear. We aimed to evaluate the alterations in HBV-specific B-cells during treatment and therefore explored the mechanism of functional recovery of HBsAg-specific B-cells. METHODS: We included 39 nucleos(t)ide analogues-treated CHB patients who received sequential combination therapy with PEG-IFNα and 8 treatment-naive CHB patients. HBV-specific B-cells were characterized ex vivo using fluorescent labeled HBsAg and HBcAg. The frequency, phenotype, and subsets of HBV-specific B-cells and follicular helper T cells (Tfh-cells) were detected using flow cytometry. The functionality of HBV-specific B-cells was quantified through ELISpot assays. RESULTS: During treatment, the fraction of activated memory B-cells (MBCs) among HBsAg-specific B-cells and the expression of IgG, CXCR3, and CD38 increased. Antibody-secretion capacity of HBsAg-specific B-cell was restored after treatment only in patients with a functional cure and it showed a positive correlation with serum hepatitis B surface antibody levels. The phenotype and function of HBsAg-specific B-cells differed between patients with and without functional cure. Patients with functional cure exhibited IgG+ classical MBCs and plasmablasts in HBsAg-specific B-cells. HBcAg-specific B-cells displayed both attenuated antibody secretion with reduced IgG expression and an IgM+ atypical type of MBCs after treatment, irrespective of with and without functional cure. The number of CD40L+ Tfh-cells increased after PEG-IFNα treatment and positively correlated with HBsAg-specific B-cell activation. CONCLUSIONS: After PEG-IFNα treatment, HBsAg- and HBcAg-specific B-cells exhibit various changes in antibody secretion. Their functional differences are reflected in the alterations in phenotypes and subtypes. The presence of CD40L+ Tfh-cells is associated with the active recovery of HBsAg-specific B-cells. IMPACT AND IMPLICATIONS: HBV-related complications and hepatocellular carcinoma remain the leading causes of mortality from chronic liver disease worldwide, and a cure is rarely achieved with antiviral therapies. Elucidating the immunological mechanisms underlying the functional cure of CHB patients offers a promising therapeutic strategy for viral clearance, such as therapeutic vaccine. We analyzed the alterations in HBV-specific B-cells in patients treated with PEG-IFNα and identified novel pathways for immunotherapeutic boosting of B cell immunity.

Liver Stiffness Measurement can Predict Liver Inflammation in Chronic Hepatitis B Patients with Normal Alanine Transaminase.

Background and Aims: To determine whether liver stiffness measurement (LSM) indicates liver inflammation in chronic hepatitis B (CHB) with different upper limits of normal (ULNs) for alanine aminotransferase (ALT). Methods: We grouped 439 CHB patients using different ULNs for ALT: cohort I, ≤40 U/L (439 subjects); cohort II, ≤35/25 U/L (males/females; 330 subjects); and cohort III, ≤30/19 U/L (males/females; 231 subjects). Furthermore, 84 and 96 CHB patients with normal ALT (≤40 U/L) formed the external and prospective validation groups, respectively. We evaluated the correlation between LSM and biopsy-confirmed liver inflammation, and determined diagnostic accuracy using area under the curve (AUC). A noninvasive LSM-based model was developed using multivariate logistic regression. Results: Fibrosis-adjusted LSM values significantly increased with increasing inflammation. The AUCs of LSM in cohorts I, II, and III were 0.799, 0.796, and 0.814, respectively, for significant inflammation (A≥2) and 0.779, 0.767, and 0.770, respectively, for severe inflammation (A=3). Cutoff LSM values in all cohorts for A≥2 and A=3 were 6.3 and 7.5 kPa, respectively. Internal, external, and prospective validations showed high diagnostic accuracy of LSM for A≥2 and A=3, and no significant differences in AUCs among the four groups. LSM and globulin independently predicted A≥2. The AUC of an LSM-globulin model for A≥2 exceeded those of globulin, ALT, and AST, but was similar to that of LSM. Conclusions: LSM predicted liver inflammation and guided the indication of antiviral therapy for CHB in patients with normal ALT.

Effect of liver inflammation on accuracy of FibroScan device in assessing liver fibrosis stage in patients with chronic hepatitis B virus infection.

BACKGROUND: Transient elastography (FibroScan) is a new and non-invasive test, which has been widely recommended by the guidelines of chronic hepatitis B virus (HBV) management for assessing hepatic fibrosis staging. However, some confounders may affect the diagnostic accuracy of the FibroScan device in fibrosis staging. AIM: To evaluate the diagnostic value of the FibroScan device and the effect of hepatic inflammation on the accuracy of FibroScan in assessing the stage of liver fibrosis in patients with HBV infection. METHODS: The data of 416 patients with chronic HBV infection who accepted FibroScan, liver biopsy, clinical, and biological examination were collected from two hospitals retrospectively. Receiver operating characteristic (ROC) curves were used to analyze the diagnostic performance of FibroScan for assessing the stage of liver fibrosis. Any discordance in fibrosis staging by FibroScan and pathological scores was statistically analyzed. Logistic regression and ROC analyses were used to analyze the accuracy of FibroScan in assessing the stage of fibrosis in patients with different degrees of liver inflammation. A non-invasive model was constructed to predict the risk of misdiagnosis of fibrosis stage using FibroScan. RESULTS: In the overall cohort, the optimal diagnostic values of liver stiffness measurement (LSM) using FibroScan for significant fibrosis (≥ F2), severe fibrosis (≥ F3), and cirrhosis (F4) were 7.3 kPa [area under the curve (AUC) = 0.863], 9.7 kPa (AUC = 0.911), and 11.3 kPa (AUC = 0.918), respectively. The rate of misdiagnosis of fibrosis stage using FibroScan was 34.1% (142/416 patients). The group of patients who showed discordance between fibrosis staging using FibroScan and pathological scores had significantly higher alanine aminotransferase and aspartate aminotransferase levels, and a higher proportion of moderate to severe hepatic inflammation, compared with the group of patients who showed concordance in fibrosis staging between the two methods. Liver inflammation activity over 2 (OR = 3.53) was an independent risk factor for misdiagnosis of fibrosis stage using FibroScan. Patients with liver inflammation activity ≥ 2 showed higher LSM values using FibroScan and higher rates of misdiagnosis of fibrosis stage, whereas the diagnostic performance of FibroScan for different fibrosis stages was significantly lower than that in patients with inflammation activity < 2 (all P < 0.05). A non-invasive prediction model was established to assess the risk of misdiagnosis of fibrosis stage using FibroScan, and the AUC was 0.701. CONCLUSION: Liver inflammation was an independent risk factor affecting the diagnostic accuracy of FibroScan for fibrosis stage. A combination of other related non-invasive factors can predict the risk of misdiagnosis of fibrosis staging using FibroScan.

Long-term outcomes in Chinese patients with chronic hepatitis B receiving nucleoside/nucleotide analogue therapy in real-world clinical practice: 5-year results from the EVOLVE study.

BACKGROUND: In China, the optimal management of individuals living with chronic HBV infection (CHB) remains an unmet need. The EVOLVE Study was a 5-year prospective, longitudinal, observational study that compared the clinical outcomes in treatment-naive CHB patients receiving entecavir (ETV) or lamivudine (LAM)-based therapies. METHODS: Males or females aged ≥18 years, diagnosed with CHB regardless of cirrhosis or hepatitis B e antigen (HBeAg) status were enrolled from tier 2 city hospitals (between 2012-2014). The choice of initial therapy and subsequent treatment modifications was at the discretion of treating physicians. Key outcomes included treatment modifications, virological response (HBV DNA <300 copies/ml) and HBV disease progression. RESULTS: Of the 3,408 patients enrolled, 1,807 and 628 received ETV and LAM-based therapy, respectively. The mean age was 39.5 years, 74% were male and 22.9% had cirrhosis. The rate of treatment modification was higher in the LAM-based versus ETV group (25.9% versus 13.7%); viral breakthrough was the most common reason in the LAM-based group versus financial reasons in the ETV group. At week 240, the virological response rate was 73% in both treatment groups. Compared with LAM-based therapy, ETV was associated with a significantly lower incidence of viral breakthrough (12.6% versus 2.1%) and genotypic resistance (10.1% versus 1.2%; P<0.0001 for both); significantly lower risk of HBV disease progression (14.0% versus 10.7%; P=0.0113); and lower rates of progression to decompensated cirrhosis (9.6% versus 6.4%) and hepatocellular carcinoma (1.9% versus 0.8%). CONCLUSIONS: This real-world, longitudinal study demonstrated a significantly lower risk of HBV-related disease progression, viral breakthrough and resistance with ETV versus LAM-based therapy. ClinicalTrials.gov NCT01726439.

Mechanism of inhibiting proliferation of hepatocellular carcinoma Hepa1-6 cells by embryonic stem cell-conditioned medium.

The present study aimed to investigate the antiproliferative effect of embryonic stem cell-conditioned medium (ESC-CM) on the mouse liver cancer Hepa1-6 cells in vitro. Furthermore, in order to elucidate the underlying molecular mechanism, the microRNAs (miRNAs) in ESC-CM associated with the inhibition of Hepa1-6 proliferation were identified. Following the co-culture of ESC-CM and Hepa1-6 in Transwell chambers, the proliferation, cell cycle, apoptosis and associated protein expression were determined in Hepal-6 cells. Moreover, miRNA array analysis was employed to identify differentially expressed miRNAs. Based on the differentially expressed miRNAs, the target genes and potential associated signaling pathways were determined. Finally, RT-qPCR was conducted to confirm the above results. The ESC-CM inhibited Hepal-6 cell proliferation and increased the percentage of cells at G1 phase and decreased the percentage of cells at the G2/M phase of the cell cycle. The expression of cyclin D1/cyclin-dependent kinase (CDK)4/CDK6 was decreased following co-culture, with no effect on cell apoptosis. Six significantly regulated miRNAs were identified and 423 putative target genes of these regulated miRNAs were predicted. Gene ontology analysis revealed the putative target genes to be associated with the 'DNA replication (GO: 0006260)' GO term, 'apoptosis' and 'signal transduction'. The Kyoto Encyclopedia of Genes and Genomes analysis indicated that deregulated miRNAs were enriched in the Wnt signaling (KEGG entry: Map 04310) and Hippo signaling pathways (KEGG entry: Map 04390), pathways associated with cancer. Overall, the present study demonstrated the inhibition of Hepa1-6 cell line proliferation upon treatment with ESC-CM, by decreasing cell cycle-associated protein cyclin D1/CDK4/CDK6 expression and arresting cells in G1 phase of the cell cycle, with no effect on cell apoptosis. Furthermore, the inhibition of proliferation by ESC-CM may be mediated by miRNAs that affect cell cycle-associated mRNAs and the Wnt signaling pathway.

Expression and Clinical Significance of Decoy Receptor 3 in Acute-on-Chronic Liver Failure.

AIMS: To explore the expression level and clinical significance of decoy receptor 3 (DcR3) in patients with acute-on-chronic liver failure (ACLF). METHODS: Serum DcR3 levels were measured by enzyme-linked immunosorbent assay (ELISA) in 76 patients with ACLF and 41 non-ACLF patients with chronic liver disease. Blood routine and liver functions were accessed for their correlations with DcR3. RESULTS: Serum DcR3 in ACLF patients was significantly higher than that in non-ACLF patients. It was positively correlated with neutrophilic granulocyte, aspartate aminotransferase, prothrombin time, and international standardized ratio, but negatively correlated with platelet and serum albumin. At the early stage, the level of DcR3 was not significantly different between the survival and nonsurvival group of ACLF. However, at the late stage, DcR3 increased in nonsurvival and gradually decreased in survivals. The baseline DcR3 could not sufficiently predict the outcome of ACLF, while the change of DcR3 within the first week displayed a better predictive value than model for end-stage liver disease (MELD) score. CONCLUSIONS: DcR3 was highly expressed in patients with ACLF and correlated with several clinical indices. Dynamic change of DcR3 might predict the prognosis of ACLF.

Ceruloplasmin, a reliable marker of fibrosis in chronic hepatitis B virus patients with normal or minimally raised alanine aminotransferase.

AIM: To develop a non-invasive model to evaluate significant fibrosis and cirrhosis by investigating the association between serum ceruloplasmin (CP) levels and liver fibrosis in chronic hepatitis B (CHB) patients with normal or minimally raised alanine aminotransferase (ALT). METHODS: Serum samples and liver biopsy were obtained from 193 CHB patients with minimally raised or normal ALT who were randomly divided into a training group (n = 97) and a validation group (n = 96). Liver histology was evaluated by the METAVIR scoring system. Receiver operator characteristic curves were applied to the diagnostic value of CP for measuring liver fibrosis in CHB patients. Spearman rank correlation analyzed the relationship between CP and liver fibrosis. A non-invasive model was set up through multivariate logistic regression analysis. RESULTS: Serum CP levels individualized various fibrosis stages via area under the curve (AUC) values. Multivariate analysis revealed that CP levels were significantly related to liver cirrhosis. Combining CP with serum GGT levels, a CG model was set up to predict significant fibrosis and liver cirrhosis in CHB patients with normal or minimally raised ALT. The AUC, sensitivity, specificity, positive predictive value, and negative predictive value were 0.84, 83.1%, 78.6%, 39.6%, and 96.5% to predict liver cirrhosis, and 0.789, 80.26%, 68.38%, 62.25%, and 84.21% to predict significant fibrosis. This model expressed a higher AUC than FIB-4 (age, ALT, aspartate aminotransferase, platelets) and GP (globulin, platelets) models to predict significant fibrosis (P = 0.019 and 0.022 respectively) and revealed a dramatically greater AUC than FIB-4 (P = 0.033) to predict liver cirrhosis. CONCLUSION: The present study showed that CP was independently and negatively associated with liver fibrosis. Furthermore, we developed a novel promising model (CG), based on routine serum markers, for predicting liver fibrosis in CHB patients with normal or minimally raised ALT.

Noninvasive models for assessment of liver fibrosis in patients with chronic hepatitis B virus infection.

There are approximately 240 million patients with chronic hepatitis B virus (HBV) infection worldwide. Up to 40% of HBV-infected patients can progress to liver cirrhosis, hepatocellular carcinoma or chronic end-stage liver disease during their lifetime. This, in turn, is responsible for around 650000 deaths annually worldwide. Repeated hepatitis flares may increase the progression of liver fibrosis, making the accurate diagnosis of the stage of liver fibrosis critical in order to make antiviral therapeutic decisions for HBV-infected patients. Liver biopsy remains the "gold standard" for diagnosing liver fibrosis. However, this technique has recently been challenged by the development of several novel noninvasive tests to evaluate liver fibrosis, including serum markers, combined models and imaging techniques. In addition, the cost and accessibility of imaging techniques have been suggested as additional limitations for invasive assessment of liver fibrosis in developing countries. Therefore, a noninvasive assessment model has been suggested to evaluate liver fibrosis, specifically in HBV-infected patients, owing to its high applicability, inter-laboratory reproducibility, wide availability for repeated assays and reasonable cost. The current review aims to present the status of knowledge in this new and exciting field, and to highlight the key points in HBV-infected patients for clinicians.

A Retrospective Study on the Significance of Liver Biopsy and Hepatitis B Surface Antigen in Chronic Hepatitis B Infection.

To investigate changes in the HBV replication level along with the natural course of chronic HBV infection and to examine the accuracy of the immune tolerant phase defined by the serological profile.A total of 390 chronic HBV-infected patients were retrospectively recruited for this study. They were classified into immune-tolerance (IT), immune-clearance (IC), low-replicative (LR), and HBeAg-negative hepatitis (ENH) phases according to serological profiles (single-standard, SS) or dual-standard (DS) with the inclusion of liver histology. Serum HBV DNA and HBsAg were quantitatively measured, and liver histology was quantitatively analyzed.The accuracy of the SS-defined IT phase was low, and active pathological changes were detected in 56 of 112 SS-defined IT patients. DS-defined IT patients had higher HBsAg levels (P = 0.0002) than the SS-defined patients. The quantitative HBsAg level can help identify SS-defined IT patients with potential liver injury. The area under the received operating characteristic curve for predicting the DS-defined IT phase was 0.831 (HBsAg 4.398 log IU/mL; sensitivity 87.5%; specificity 73.2%). HBV DNA was reduced by 4 logs, whereas HBsAg was only decreased by 2 logs with HBeAg positive to negative phase conversion.Approximately half of IT patients defined by SS may have medium or severe liver injury. Quantitative measurement of the HBsAg level can help identify SS-defined IT patients with potential liver injury.

[Effects of fenofibrate on hepatocyte apoptosis in nonalcoholic fatty liver].

OBJECTIVE: To use a rat model of nonalcoholic liver disease (NAFLD) to observe effects of the peroxisome proliferator-activated receptor-a (PPAR-a) agonist fenofibrate on hepatic steatosis in nonalcoholic fatty liver and to investigate the underlying mechanism. METHODS: Sixty-six Sprague-Dawley rats were given adaptive feeding for 1 week and then randomly allocated into the following three groups: unmodeled control (group C,n =18), untreated NAFLD model (group M, n =24), and fenofibrate-treated NAFLD model (group F, n =24).Group C rats were given a normal diet, while group M and group F rats were given a high-fat diet. After model establishment, the group F rats were treated with fenofibrate (10 mg/kg/d, intraperitoneal) and the group C and group M rats were given sham-treatment with cosolvent (5 mL/kg/d, intraperitoneal). At the end of treatment weeks 4, 6 and 8, one-third of rats in each group were euthanized.Liver tissues were assessed by hematoxylin-eosin (HE) staining to determine level of steatosis and inflammaion activity, and by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling to measure changes in hepatocyte apoptosis index. Changes in expression levels of the PPAR-a receptor and apoptosis factors (bcl-2, bax and caspase-3) were assessed by reverse transcription-PCR and immunohistochemistry. RESULTS: The NAFLD modeled rats showed appropriate induction of hepatic steatosis, hepatic inflammation, and hepatocyte apoptosis. Compared to the group M rats, the group F rats showed lower expression of PPAR-and bcl-2 and higher expression of bax and caspase-3 at both the mRNA and protein level. CONCLUSION: Fenofibrate can ameliorate hepatic steatosis in an experimental rat model of NAFLD, and the mechanism may be associated with inhibition of hepatocyte apoptosis.

[Validation study of prediction models of hepatitis B virus-related hepatocellular carcinoma].

OBJECTIVE: To validate two previously published models (REACH-B score and CU-HCC score) for predicting the risk of developing hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). METHODS: In-patients of the Liver Center of First Affiliated Hospital Fujian Medical University who tested positive for hepatitis B surface antigen (HBsAg;more than 6 months) and were admitted for treatment between October 1,2004 and May 1,2014 were enrolled for study. The 627 study participants were grouped according to presence of HCC (151 in the HCC case group, and 476 in the non-HCC control group). Relevant clinical data from 3 and 5 years prior to the current hospital admission were collected retrospectively and assessed using the REACH-B and CU-HCC scoring systems. A subset of the study participants (65 HCC cases, and 94 non-HCC controls) was used for the verification analysis of prediction for 5-year risk of HBV-related HCC.T-test, rank sum test, chisquare test and the receiver operating characteristic curve were used for statistical analyses. RESULTS: For the REACH-B score,prediction of 3-year risk of developing HCC had an area under the curve (AUC) of 0.78,a sensitivity of 73.00% and a specificity of 78.70%.In male patients with alanine aminotransferase (ALT) more than or equal to 45 U/L, the REACH-B score prediction of 3-year risk of developing HCC had an AUC of 0.89, a sensitivity of 87.09% and a specificity of 83.86%. The REACH-B score prediction of 5-year risk of HCC had an AUC of 0.79,a sensitivity of 73.60% and a specificity of 75.53%; the CU-HCC score prediction of 5-year risk of HCC had an AUC of 0.76, a sensitivity of 78.40% and a specificity of 77.40%. CONCLUSION: Both the REACH-B and CUHCC scoring systems can be used for HCC prediction among patients at the First Affiliated Hospital Fujian Medical University. For male patients with ALT more than or equal to 45 U/L,the REACH-B score may be a more sensitive predictor for 3-year risk of developing HBV-related HCC.

HBeAg-negative chronic hepatitis patients should be monitored more strictly: a cross-sectional retrospective study on antiviral treatment-naïve patients.

AIM: To investigate the prevalence and characteristics of hepatitis B e antigen (HBeAg) negative/treatment naïve subjects with low hepatitis B virus (HBV) DNA levels (<10(4) copies/ml) and low alanine aminotransferase (ALT) levels (<2 × upper limit of normal) in patients with HBV-related hepatocellular carcinoma (HCC). METHODS: A total of 226 treatment naïve patients diagnosed with HBV-related HCC, divided into five Barcelona Clinic Liver Cancer (BCLC) stages, were enrolled and retrospectively analyzed. Virological parameters including hepatitis B surface antigen (HBsAg), HBeAg, HBV DNA levels, and laboratory parameters including ALT and aspartame aminotransferase were accessed at the time of HCC was diagnosed. Comparison between HBeAg positive patients and HBeAg negative patients was performed using a χ(2) test. RESULTS: While laboratory parameters correlated with BCLC stages, virological parameters did not. HBeAg negative patients were more prevalent than HBeAg positive patients (160, 70.8% vs. 66, 29.2%). HBsAg and HBV DNA levels in HBeAg negative patients were significantly lower than that in HBeAg positive patients (all P < 0.001). Of the 160 HBeAg negative patients, 74 (46.25%) had low HBsAg, 76 (47.5%) had low DNA levels, and 35 (21.9%) patients had low DNA and normal ALT levels. CONCLUSIONS: In treatment naïve patients with HBV-related HCC, the majority (70.8%) were HBeAg negative patients. More than one fifth of HBeAg negative patients had low HBV DNA levels and normal ALT levels, indicating more strict monitoring for HBeAg negative patients is needed.

Interleukin-6 as an early diagnostic marker for bacterial sepsis in patients with liver cirrhosis.

OBJECTIVE: Liver cirrhosis is associated with frequent bacterial infections that increase the mortality rate. However, the early diagnosis and treatment of these infections are often difficult. In this retrospective-prospective observational study, the serum levels of interleukin-6 (IL-6) and procalcitonin (PCT) were measured in 233 cirrhotic patients to evaluate the early diagnostic and prognostic values of IL-6 and PCT for cirrhotic patients. METHODS: Cirrhotic patients admitted to the Liver Research Center of the First Affiliated Hospital of Fujian Medical University between 1 October 2012 and 30 June 2014 were enrolled. They showed no evidence of infection on admission, and all had first onset of fever and met the systemic inflammatory response syndrome criteria 72 hours after admission. The serum IL-6 and PCT levels were determined on admission, at the onset of fever (0 hour) and 24 and 48 hours after fever onset. RESULTS: A total of 233 cirrhotic patients, including 183 men and 50 women, with a median age of 56 (46-65) years were enrolled. A training group of 159 patients was retrospectively enrolled from 1 October 2012 to 31 December 2013, and a validation group of 74 patients was prospectively enrolled from 1 January 2014 to 30 June 2014. Among these patients, 134 were diagnosed with bacterial sepsis, 96 of whom were in the training group and 38 of whom were in the validation group; infections were ultimately ruled out in 99 patients: 63 training patients and 36 validation patients. At 0 hour, the IL-6 and PCT levels as well as the proportion of neutrophils were much higher in septic patients than in nonseptic ones. The IL-6 level and proportion of neutrophils peaked upon the onset of fever, 24 hours before the PCT levels and white blood cell count, and then sharply declined. The area under the receiver operating characteristic curve of IL-6 for diagnosing sepsis was largest at the onset of fever (area under the receiver operating characteristic curve, 0.983; 95% confidence interval, 0.967-0.999). The threshold of IL-6 for diagnosis was 135 pg/mL, with a sensitivity of 94.8% and a specificity of 93.7%. These diagnostic values were also confirmed in the validation group, with a sensitivity of 97.4% and specificity of 80.6%. Eleven (11.5%) patients died, and 85 (88.5%) patients recovered in the sepsis group of training patients after a 4-week follow-up. The IL-6 level was significantly higher in the nonsurvival group than that in the survival group (1813.00 vs 472.10 pg/mL, P = .004) at the onset of sepsis. The cutoff value for predicting prognosis was 1105 pg/mL, with a sensitivity of 81.8% and a specificity of 76.5%. CONCLUSIONS: The serum IL-6 levels increased earlier than the PCT in septic cirrhotic patients. The direct measurement of the serum IL-6 level can help to rapidly detect bacterial infection, thus allowing for early therapeutic decisions and prognostic predictions.

Hepatitis B surface antigen in late hepatitis B infection.

Hepatitis B surface antigen (HBsAg) levels are used to evaluate and monitor clinical phases of chronic hepatitis B infection but their clinical significance is unclear in the late complications, cirrhosis of the liver and hepatocellular carcinoma. This study aimed to evaluate HBsAg levels across the whole natural history of hepatitis B virus infection, including late complications. This retrospective, cross-sectional study enrolled 838 treatment-naive patients diagnosed with chronic hepatitis B infection at First Affiliated Hospital of Fujian Medical University between 2009 and 2012. Patients were classified into six groups: immunotolerance, immunoclearance, low replicative, negative hepatitis e (HBeAg) phases, liver cirrhosis, and hepatocellular carcinoma. Main outcome measures were serum HBsAg, HBeAg, HBV DNA, total bilirubin, albumin, alanine and aspartate aminotransferase, and quantitative correlation of HBsAg with HBV DNA. HBsAg levels declined significantly between clinical phases of infection (all P < 0.001) and were significantly lower in decompensated than in compensated cirrhosis (2.90 vs. 3.30, P < 0.001) but not significantly different between early versus advanced hepatocellular carcinoma. Significant positive correlations were observed between serum HBsAg and HBV DNA at immunoclearance and HBeAg negative phases, compensated and decompensated liver cirrhosis and advanced but not early hepatocellular carcinoma (all P < 0.001). HBsAg and HBV DNA were significantly higher in HBeAg positive patients with advanced hepatocellular carcinoma (P < 0.001). HBsAg levels differ significantly between chronic hepatitis B infection phases, decreasing progressively from chronic infection to cirrhosis and hepatocellular carcinoma. Significant correlations are found between serum HBsAg and HBV DNA.

Alpha-fetoprotein level as a biomarker of liver fibrosis status: a cross-sectional study of 619 consecutive patients with chronic hepatitis B.

BACKGROUND: Hepatitis B virus (HBV) infection is a serious public health problem worldwide. This study aimed to investigate the relationship between serum alpha-fetoprotein (AFP) levels and pathological stages of liver biopsy in patients with chronic hepatitis B (CHB). METHODS: The study included 619 patients who were diagnosed with CHB from March 2005 to December 2011. AFP levels were measured by electrochemiluminescence. Liver biopsy samples were classified into five levels of inflammation (G) and fibrosis (S) stages, according to the Chinese guidelines for prevention and treatment of viral hepatitis. Two multivariable ordinal regression models were performed to determine associations between AFP, GGT, and APRI (AST/PLT ratio) and stages of inflammation and fibrosis. RESULTS: Significant positive and moderate correlations were shown between AFP levels and inflammation stages and between AFP levels and fibrosis stages (ρ = 0.436 and 0.404, p < 0.001). Median values of AFP at liver fibrosis stages S0-1, S2, S3, and S4 were 3.0, 3.4, 5.4, and 11.3 ng/ml, respectively, and median APRI (AST/PLT ratio) was 0.41. Receiver operating characteristic (ROC) curve analyses revealed that the areas under the curves (AUCs) were 0.685, 0.727, and 0.755 (all p <0.001) for judging inflammation stages of G ≥ 2, G ≥ 3, G = 4 by AFP; and 0.691, 0.717, and 0.718 (all p <0.001) for judging fibrosis stages of S ≥ 2, S ≥ 3, and S = 4 by AFP. APRI levels showed significant positive and moderate correlations with inflammation stages (ρ = 0.445, p < 0.001). AST, GGT, and APRI levels showed significant positive but very weak to weak correlations with fibrosis stages (ρ = 0.137, 0.237, 0.281, p < 0.001). CONCLUSIONS: Serum AFP levels increased as pathological levels of inflammation and fibrosis increased in CHB patients. Our data showed the clinical significance of serum AFP levels in diagnosing liver inflammation and fibrosis. Assessment of liver pathology may be improved by creating a predictive mathematical model by which AFP levels with other biomarkers.

A new model for predicting liver cirrhosis in chronic hepatitis B virus carriers with low serum alanine transaminase activity.

OBJECTIVES: To develop a cirrhosis-predicted model in chronic hepatitis B virus carriers with alanine transarninase (ALT) less than two times the upper limit of normal (ULN). METHODS: Treatment-naive patients (n=278), who had undergone liver biopsies, were randomly divided into two groups - a training group and a validation group. Thirteen bio-clinical parameters were analyzed. A liver cirrhosis-predicting model (PPT model) was constructed using multivariate analysis. The diagnostic value of the model was analyzed by the receiving operating characteristics (ROC) method and compared with other available models. RESULTS: A PPT model to predict liver cirrhosis was derived from three independent predictors of liver fibrosis [platelet count (PLT), prothrombin time (PT) and total bile acid (TBA)]. PPT model predicted cirrhosis with an area under the ROC (AUROC) curve of 0.83, a positive predictive value of 86.7% and a negative predictive value of 95.2%. Compared with APRI, FIB-4, age-AST model, AP index and APGA model, PPT model had the highest correlation coefficient (r=0.49) and greater predictive performance (AUROC of 0.83). CONCLUSIONS: The PPT model was accurate in predicting cirrhosis and may reduce the need for liver biopsy in chronic hepatitis B virus carriers with ALT less than two times ULN.

Serum cytokeratin-18 in the diagnosis of non-alcoholic steatohepatitis: A meta-analysis.

AIM: Identifying patients with non-alcoholic steatohepatitis (NASH), a more aggressive form with a worse prognosis than for simple steatosis, is highly important. Liver biopsy still remains the gold standard for diagnosing NASH, but with limitations. The diagnostic value of serum cytokeratin-18 (CK-18) in predicting NASH is still indefinite. METHODS: We selected relevant studies by a literature search of the PubMed, Ovid Medline and Cochrane Library databases up to January 2012. A DerSimonian-Laird random effects model was used to compute the pooled estimates of sensitivity (SEN), specificity (SPE), and diagnostic odds ratio (DOR), and a summary receiver operating characteristic (SROC) curve was constructed. Stratified analysis was performed to explore the heterogeneity in test accuracy. Funnel plot and Egger's regression were performed to assess publication bias. RESULTS: A total of 10 studies with 838 patients were included (nine CK-18 fragments and five total CK-18 studies) in this meta-analysis. Among nine CK-18 fragment studies with a significant publication bias, the pooled results on SEN, SPE and DOR were 0.83 (95% CI, 0.80-0.86), 0.71 (95% CI, 0.66-0.76) and 11.90 (95% CI, 6.05-23.40), respectively, and age and body mass index were most strongly associated with the observed heterogeneity. Among five total CK-18 studies with homogeneity, the pooled results of SEN, SPE and DOR were 0.77% (95% CI, 0.70-0.83), 0.71 (95% CI, 0.65-0.77) and 7.99 (95% CI, 4.09-15.62), respectively. The area under the ROC curve (± SE) of CK-18 fragments and total CK-18 were 0.8445 (± 0.0306) and 0.8170 (± 0.0429), respectively. CONCLUSION: Both CK-18 fragments and total CK-18 have a clinically meaningful benefit in noninvasive diagnosing of NASH, though total CK-18 has a relatively low diagnostic accuracy. CK-18 fragments may be a useful biomarker for screening rather than identifying NASH.

Distribution and clinical correlates of viral and host genotypes in Chinese patients with chronic hepatitis C virus infection.

BACKGROUND AND AIM: Chronic hepatitis C virus (HCV) infection is relatively frequent in China. This study investigated the clinical, demographic, and viral and host genetic characteristics that may influence disease manifestations and clinical management. METHODS: In this cross-sectional observational study, treatment-naïve Han ethnic adults with recently confirmed chronic HCV infection were enrolled at 28 hospitals across China. HCV genotype and host interleukin 28B (IL28B) genotypes were determined and compared with patient demographic parameters and medical status. RESULTS: Among the 997 HCV-positive patients analyzed, 56.8% were infected with HCV genotype 1b, followed in prevalence by genotypes 2, 3, and 6, with substantial regional variation. Overall, 84.1% of patients were IL28B genotype CC (rs12979860), with little regional variation. Cirrhosis was reported in 10.1% of patients and was significantly associated with hepatitis B virus coinfection, low HCV viral load, low serum alanine aminotransferase, high serum aspartate aminotransferase, diabetes, and high pickled food consumption. Medical procedures were common transmission risk factors; however, lifestyle-associated risk factors, including intravenous drug abuse and tattoos or piercings, were more common in patients with HCV genotype 3 or 6. CONCLUSIONS: Most HCV-infected Han Chinese patients were IL28B genotype CC (rs12979860). HCV genotypes varied by geographic region, and disease characteristics differed according to HCV genotype. Relatively frequent detection of advanced liver disease may reflect limitations on access to antiviral therapy, and suggests that greater awareness of factors that influence HCV-associated disease may help avoid clinical complications and improve patient outcomes.

Serum ceruloplasmin levels correlate negatively with liver fibrosis in males with chronic hepatitis B: a new noninvasive model for predicting liver fibrosis in HBV-related liver disease.

AIMS: This study aimed to investigate associations between ceruloplasmin (CP) levels, inflammation grade and fibrosis stages in patients with chronic hepatitis B (CHB) and to establish a noninvasive model to predict cirrhosis. METHODS: Liver biopsy samples and sera were collected from 198 CHB patients randomized into a training group (n=109) and a validation group (n=89). CP levels were determined using nephelometric immunoassays. Relationships between CP and liver inflammation and fibrosis were analyzed by Spearman rank correlation. Receiver operator characteristic (ROC) curves were used to evaluate the diagnostic value of CP for determining liver fibrosis in CHB. The liver pathology-predictive model was built using multivariate logistic regression analysis to identify relevant indicators. RESULTS: CP levels were lower in males than in females, lower in patients with inflammation stage G4 compared to other stages and lower in cirrhotic compared to non-cirrhotic patients. Using area under the curve (AUC) values, CP levels distinguished different stages of inflammation and fibrosis. Multivariate analysis showed that CP levels were all significantly associated with cirrhosis in males. A model was developed combining routine laboratory markers APPCI (alpha-fetoprotein [AFP], prothrombin time, and platelets [PLT] with CP) to predict fibrosis in CHB patients. The APPCI had a significantly greater AUC than FIB-4 (aspartate aminotransferase [AST]/ alanine aminotransferase [ALT]/PLT/age), APRI (AST/PLT ratio index), GPI (globin/PLT), and APGA (AST/PLT/gammaglutamyl transpeptidase [GGT]) models (all P-values<0.001). CONCLUSIONS: CP levels correlate negatively and indirectly with inflammation and fibrosis stages in male CHB patients. The APPCI model uses routine laboratory variables with CP to accurately predict liver fibrosis in CHB.

[Hepatitis B surface antigen affects the expression of lipid metabolism-related genes in HepG2 cells].

OBJECTIVE: To investigate the influence of hepatitis B virus (HBV)-encoded small surface protein (SHBs) on hepatic cell expression of host genes related to lipid metabolism. METHODS: The full-length SHBs gene was amplified from HBV genotype C by polymerase chain reaction (PCR) and cloned into the pcDNA3.1(+) expression vector for stable transfection into HepG2 cells (selected by G418 screening); cells transfected with empty vector served as control. The SHBs mRNA and protein levels were detected by reverse transcription-PCR and enzyme-linked immunosorbent assay. SHBs effects on expression of genes and proteins related to lipid metabolism were detected by real-time quantitative (q)PCR and western blotting, respectively. RESULTS: The stably transfected cell line HepG2-pn3.1-SHBs was established successfully. qPCR showed that the HepG2-pn3.1-SHBs cells had significantly down-regulated transcription of the ECHS1, APOA1 and LPL genes (0.161+/-0.043 vs. control cells: 0.210+/-0.022, t = 2.479; 0.031+/-0.007 vs. 0.094+/-0.055, t = 2.752; 0.770+/-0.036 vs. 0.982+/-0.031, t = 10.914), but significantly up-regulated ACC and SREBP-1c genes (0.113+/-0.027 vs. 0.059+/-0.022, t = -3.757; 0.019+/-0.002 vs. 0.015+/-0.001, t = -4.330). The CPT1a and PPARa genes' expression was slightly, but not significantly, down-regulated in the HepG2-pn3.1-SHBs cells (0.028+/-0.005 vs. 0.030+/-0.004, t = 1.022; 0.014+/-0.004 vs. 0.015+/-0.002, t = 0.758). Western blotting showed similar expression trends for the corresponding proteins. CONCLUSION: SHBs alters the expression of some host genes with known functions in fatty acid synthesis and decomposition; however, it remains unclear whether the hepatitis B surface antigen can directly contribute to development of hepatic steatosis.