OBJECTIVE: To explore the diagnostic value of the serum metabolites identified by high-performance liquid chromatography-mass spectrometry (HPLC/MS) for hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). METHODS: A total of 126 patients admitted to Tianjin Third Central Hospital were enrolled, including 27 patients with HBV-related hepatitis with negative viral DNA (DNA-N), 24 with HBV-related hepatitis with positive viral DNA, 24 with HBV-related liver cirrhosis, 27 with HBV-related HCC undergoing surgeries or radiofrequency ablation, and 24 with HBV-related HCC receiving interventional therapy, with 25 healthy volunteers as the normal control group. Serum samples were collected from all the subjects for HPLC/MS analysis, and the data were pretreated to establish an orthogonal partial least- squares discriminant analysis (OPLS-DA) model. The differential serum metabolites were preliminarily screened by comparisons between the HBV groups and the control group, and the characteristic metabolites were identified according to the results of non-parametric test. The potential clinical values of these characteristic metabolites were evaluated using receiver operator characteristic curve (ROC) analysis. RESULTS: A total of 25 characteristic metabolites were identified in the HBV- infected patients, including 9 lysophosphatidylcholines, 2 fatty acids, 17α-estradiol, sphinganine, 5-methylcytidine, vitamin K2, lysophosphatidic acid, glycocholic acid and 8 metabolites with few reports. The patients with HBV- related HCC showed 22 differential serum metabolites compared with the control group, 4 differential metabolites compared with patients with HBV-related liver cirrhosis; 10 differential metabolites were identified in patients with HBV-related HCC receiving interventional therapy compared with those receiving surgical resection or radiofrequency ablation. From the normal control group to HBV-related HCC treated by interventional therapy, many metabolites underwent variations following a similar pattern. CONCLUSIONS We identified 25 characteristic metabolites in patients with HBV-related HCC, and these metabolites may have potential clinical values in the diagnosis of HBV-related HCC. The continuous change of some of these metabolites may indicate the possibility of tumorigenesis, and some may also have indications for the choice of surgical approach.
Carcinoma, Hepatocellular ; blood ; diagnosis ; virology ; Case-Control Studies ; Chromatography, High Pressure Liquid ; DNA, Viral ; blood ; Hepatitis B virus ; genetics ; Hepatitis B, Chronic ; blood ; virology ; Humans ; Liver Cirrhosis ; virology ; Liver Neoplasms ; blood ; diagnosis ; virology ; Mass Spectrometry ; Metabolome ; Metabolomics ; ROC Curve

To investigate whether transcription of hepatitis B virus (HBV) gene occurs in human sperm, total RNA was extracted from sperm of patients with chronic HBV infection (test-1), from donor sperm transfected with a plasmid containing the full-length HBV genome (test-2), and from nontransfected donor sperm (control), used as the template for reverse transcription-polymerase chain reaction (RT-PCR). Positive bands for HBV DNA were observed in the test groups but not in the control. Next, to identify the role of host genes in regulating viral gene transcription in sperm, total RNA was extracted from 2-cell embryos derived from hamster oocytes fertilized in vitro by HBV-transfected (test) or nontransfected (control) human sperm and successively subjected to SMART-PCR, suppression subtractive hybridization, T/A cloning, bacterial amplification, microarray hybridization, sequencing and the Basic Local Alignment Search Tool (BLAST) search to isolate differentially expressed genes. Twenty-nine sequences showing significant identity to five human gene families were identified, with chorionic somatomammotropin hormone 2 (CSH2), eukaryotic translation initiation factor 4 gamma 2 (EIF4G2), pterin-4 alpha-carbinolamine dehydratase 2 (PCBD2), pregnancy-specific beta-1-glycoprotein 4 (PSG4) and titin (TTN) selected to represent target genes. Using real-time quantitative RT-PCR (qRT-PCR), when CSH2 and PCBD2 (or EIF4G2, PSG4 and TTN) were silenced by RNA interference, transcriptional levels of HBV s and x genes significantly decreased (or increased) (P < 0.05). Silencing of a control gene in sperm did not significantly change transcription of HBV s and x genes (P > 0.05). This study provides the first experimental evidence that transcription of HBV genes occurs in human sperm and is regulated by host genes.
Animals ; Connectin/genetics* ; Cricetinae ; Eukaryotic Initiation Factor-4G/genetics* ; Gene Expression Regulation/genetics* ; Gene Silencing ; Growth Hormone/genetics* ; Hepatitis B Surface Antigens/genetics* ; Hepatitis B virus/genetics* ; Hepatitis B, Chronic/virology* ; Humans ; Hydro-Lyases/metabolism* ; Male ; Pregnancy-Specific beta 1-Glycoproteins/genetics* ; RNA, Viral/analysis* ; Spermatozoa/virology* ; Trans-Activators/genetics* ; Transcription, Genetic ; Transfection ; Viral Regulatory and Accessory Proteins

<b>Objective:b> To investigate the effects of hepatitis B virus (HBV) genotype and mutations on the development of hepatocellular carcinoma (HCC) and to establish a new qualified HCC risk scores. <b>Methods:b> A cohort study enrolling patients with chronic HBV infection was conducted. HBV genotypes were identified by nested multiplex PCR. HBV mutations in the basic core promoter region and PreS region were sequenced after PCR amplification. Scores on risk factors were set based on nomogram. <b>Results:b> Totally, 1 525 patients were followed-up in this research. A total of 1 110 patients infected with genotype C were followed-up for 8.52 (Q(R): 5.36-11.68) years on average, of whom the incidence of HCC was 11.93/1 000 person-years. In genotype C HBV infected patients, male gender, aged 40 years and over, and four DNA mutations (T1674CG, A1762T/G1764A, A3120T, and A2962G) can increase the risk of HCC (P<0.05); interferon therapy can reduce the risk of HCC (P<0.05). A new HCC predicting model was established according to the results. After validation, the predicted disease-free survival rate was consistent with the real one. <b>Conclusions:b> Hepatitis B virus genotypes and mutations were closely associated with HCC. The new risk scoring system can well predict HCC occurrence in genotype C HBV infected patients.
Adult ; Aged ; Carcinoma, Hepatocellular/virology* ; China/epidemiology* ; Cohort Studies ; DNA, Viral/genetics* ; Female ; Genotype ; Hepatitis B virus/genetics* ; Hepatitis B, Chronic/complications* ; Humans ; Liver Neoplasms/virology* ; Male ; Middle Aged ; Mutation ; Predictive Value of Tests ; Risk Factors ; Sensitivity and Specificity

<b>Objective:b> To explore the SNP effects of patatin-like phospholipase domain which containing 3 (PNPLA3), transmembrane 6 superfamily member 2 (TM6SF2) gene, environmental effects of smoking, alcohol drinking and interaction between gene-gene, gene-environment and drinking-smoking on hepatitis B virus-associated hepatocellular carcinoma (HBV-HCC). <b>Methods:b> We collected anticoagulant peripheral blood from patients of HBV-HCC, chronic hepatitis B (CHB), liver cirrhosis (LC) and from healthy controls to detect the single nucleotide polymorphism (SNP) of patatin-like phospholipase domain containing 3 (PNPLA3) gene loci rs738409 and transmembrane 6 superfamily member 2 (TM6SF2) gene loci rs58542926, using the flight mass spectrometry method. The optimal assignment value of gene polymorphisms was defined by using the online SNP stats. Hardy-Weinberg (H-W) balance was tested for SNP. Effects of the genetic and environmental factors to HBV-HCC were analyzed by using the multiple classification logistic regression method. The gene-gene, gene-smoking and alcohol drinking interaction effects were investigated by Fork-Life analysis and binary logistic regression methods. <b>Results:b> The frequency distribution of CHB group rs738409 loci seemed not in conformity with the H-W balance (χ(2)=11.980, P<0.005). Two loci frequency distributions in the other groups were all in accordandce with the H-W balance. After adjusting for influences on age and sex and comparing to the healthy group, the rs58542926 mutation appeared as OR=1.659, 95%CI: 1.026-2.684, P=0.039, in the HBV-HCC group. When comparing to CHB group, the HBV-HCC group presented that drinking as OR=1.680, 95%CI: 1.121-2.519, P=0.012. When comparing to the LC group, the ORs of drinking and smoking were 1.539 (1.071-2.213) and 1.453 (1.005-2.099) respectively, in the HBV-HCC group. When comparing to the CHB+LC group, interactions between the HBV-HCC group were found rs738409 and rs58542926 on additive model OR=1.548 (U=1.885, P=0.029) and OR=1.658 (P=0.024) on logistic regression model while drinking was rs738409 on interaction additive model with OR=1.811(U=1.965, P=0.024). As for drinking and mutation of rs738409, the multiplication model of logistic regression showed no statistically significant differences. Interaction between smoking and drinking appeared as OR=1.756 (P<0.001) in the logistics regression multiplication model. <b>Conclusions:b> Factors as mutation of TM6SF2, smoking and drinking all appeared as risk factors for HBV-HCC. Mutations of both PNPLA3 and TM6SF2, together with smoking and drinking all served as risk factors for HBV-HCC. However, the mutation of single PNPLA3 appeared as a protective factor on HBV-HCC.
Alcohol Drinking/adverse effects* ; Carcinoma, Hepatocellular/virology* ; Case-Control Studies ; Epistasis, Genetic ; Gene-Environment Interaction ; Genetic Predisposition to Disease ; Genotype ; Hepatitis B virus ; Hepatitis B, Chronic ; Humans ; Lipase/genetics* ; Liver Cirrhosis, Alcoholic/complications* ; Liver Neoplasms/virology* ; Membrane Proteins/genetics* ; Polymorphism, Single Nucleotide ; Smoking/adverse effects*

Recent studies suggest that liver cirrhosis is reversible after administering oral nucleos(t)ide analogue therapy to patients with hepatitis B virus (HBV) infection. However, few studies have addressed whether esophageal varices can regress after such therapy. We report a case of complete regression of esophageal varices during entecavir therapy in patients with HBV-related liver cirrhosis, suggesting that complications of liver cirrhosis such as esophageal varices can regress after the long-term suppression of HBV replication.
Abdomen/diagnostic imaging ; Antiviral Agents/*therapeutic use ; DNA, Viral/blood ; Esophageal and Gastric Varices/complications/prevention & control ; Guanine/*analogs & derivatives/therapeutic use ; Hepatitis B virus/genetics ; Hepatitis B, Chronic/complications/*drug therapy/virology ; Humans ; Liver Cirrhosis/*diagnosis/etiology ; Male ; Middle Aged ; Polymerase Chain Reaction ; Ultrasonography

BACKGROUND/AIMS: This study aimed to clarify the effect of obesity on the development of hepatocellular carcinoma (HCC) in chronic hepatitis B (CHB) patients receiving antiviral treatment. METHODS: This study applied a retrospective analysis to a historical cohort in Bundang Jesaeng Hospital. In total, 102 CHB patients were treated with entecavir as an initial treatment for CHB and checked for obesity using a body composition analyzer. Hepatic steatosis was measured semiquantitatively using Hamaguchi’s scoring system in ultrasonography. Risk factors for the development of HCC were analyzed, including obesity-related factors (body mass index [BMI], waist circumference [WC], waist-to-hip ratio [WHR], visceral fat area [VFA], and hepatic steatosis). RESULTS: The median follow-up duration of the patients was 45.2 months (interquartile range: 36.0-58.3 months). The cumulative incidence rates of HCC at 1 year, 3 years, and 5 years were 0%, 5.3%, and 9.0%, respectively. Univariable analysis revealed that the risk factors for HCC development were a platelet count of <120,000 /mm² (hazard ratio [HR]=5.21, P=0.031), HBeAg negativity (HR=5.61, P=0.039), and liver cirrhosis (HR=10.26, P=0.031). Multivariable analysis showed that the significant risk factor for HCC development was liver cirrhosis (HR=9.07, P=0.042). However, none of the obesity-related risk factors were significantly associated with HCC: BMI ≥25 kg/m² (HR=0.90, P=0.894), WC ≥90 cm (HR=1.10, P=0.912), WHR ≥0.9 (HR=1.94, P=0.386), VFA ≥100 cm² (HR=1.69, P=0.495), and hepatic steatosis (HR=0.57, P=0.602). CONCLUSION: HCC development is associated with liver cirrhosis but not obesity-related factors in CHB patients receiving entecavir.
Adult ; Antiviral Agents/*therapeutic use ; Body Mass Index ; Carcinoma, Hepatocellular/epidemiology/*etiology ; Cohort Studies ; DNA, Viral/blood ; Female ; Guanine/*analogs & derivatives/therapeutic use ; Hepatitis B virus/genetics/isolation & purification ; Hepatitis B, Chronic/complications/*drug therapy/virology ; Humans ; Incidence ; Liver Cirrhosis/complications ; Liver Neoplasms/epidemiology/*etiology ; Male ; Middle Aged ; Obesity/*complications ; Proportional Hazards Models ; Retrospective Studies ; Risk Factors ; Viral Load

BACKGROUND/AIMS: Before tenofovir (TDF) become available in South Korea, combination therapy with entecavir (ETV) and adefovir (ADV) was the most potent regimen for chronic hepatitis B (CHB) patients who fail to respond to rescue therapy for drug resistance. We analyzed the efficacy of ETV-ADV combination therapy and investigated the clinical and clonal results of TDF-based rescue therapy in CHB patients refractory to this combination. METHODS: We retrospectively reviewed the medical records of CHB patients treated for up to 3 years with ETV-ADV combination therapy as a rescue therapy for drug resistance. In cases refractory to this combination, clinical and clonal analyses were performed for TDF-based rescue therapy. RESULTS: The analysis was performed on 48 patients. Twelve patients achieved a virological response (VR) within 3 years. A VR was subsequently achieved in nine of the ten patients without a VR who switched to TDF monotherapy. A VR was also achieved in six of the seven patients who switched to lamivudine-TDF combination therapy, and in two of the two patients who switched to ETV-TDF combination therapy. In an in vitro susceptibility test, viral replication was detected with TDF monotherapy but not with ETV-TDF combination therapy. CONCLUSIONS: The efficacy of ETV-ADV combination therapy was insufficient in CHB patients who were refractory to rescue therapy. A more potent regimen such as ETV-TDF combination therapy may be considered in such refractory cases.
Adenine/*analogs & derivatives/therapeutic use ; Adult ; Aged ; Antiviral Agents/*therapeutic use ; DNA, Viral/analysis/metabolism ; Drug Resistance, Viral ; Drug Therapy, Combination ; Female ; Guanine/*analogs & derivatives/therapeutic use ; Hepatitis B virus/genetics/isolation & purification ; Hepatitis B, Chronic/*drug therapy/virology ; Humans ; Male ; Middle Aged ; Organophosphonates/*therapeutic use ; Polymerase Chain Reaction ; Republic of Korea ; Retrospective Studies ; Tenofovir/*therapeutic use ; Treatment Outcome

BACKGROUND/AIMS: Before tenofovir (TDF) become available in South Korea, combination therapy with entecavir (ETV) and adefovir (ADV) was the most potent regimen for chronic hepatitis B (CHB) patients who fail to respond to rescue therapy for drug resistance. We analyzed the efficacy of ETV-ADV combination therapy and investigated the clinical and clonal results of TDF-based rescue therapy in CHB patients refractory to this combination. METHODS: We retrospectively reviewed the medical records of CHB patients treated for up to 3 years with ETV-ADV combination therapy as a rescue therapy for drug resistance. In cases refractory to this combination, clinical and clonal analyses were performed for TDF-based rescue therapy. RESULTS: The analysis was performed on 48 patients. Twelve patients achieved a virological response (VR) within 3 years. A VR was subsequently achieved in nine of the ten patients without a VR who switched to TDF monotherapy. A VR was also achieved in six of the seven patients who switched to lamivudine-TDF combination therapy, and in two of the two patients who switched to ETV-TDF combination therapy. In an in vitro susceptibility test, viral replication was detected with TDF monotherapy but not with ETV-TDF combination therapy. CONCLUSIONS: The efficacy of ETV-ADV combination therapy was insufficient in CHB patients who were refractory to rescue therapy. A more potent regimen such as ETV-TDF combination therapy may be considered in such refractory cases.
Adenine/*analogs & derivatives/therapeutic use ; Adult ; Aged ; Antiviral Agents/*therapeutic use ; DNA, Viral/analysis/metabolism ; Drug Resistance, Viral ; Drug Therapy, Combination ; Female ; Guanine/*analogs & derivatives/therapeutic use ; Hepatitis B virus/genetics/isolation & purification ; Hepatitis B, Chronic/*drug therapy/virology ; Humans ; Male ; Middle Aged ; Organophosphonates/*therapeutic use ; Polymerase Chain Reaction ; Republic of Korea ; Retrospective Studies ; Tenofovir/*therapeutic use ; Treatment Outcome

BACKGROUND/AIMS: Tenofovir disoproxil fumarate (TDF) plays a pivotal role in the management of drug-resistant chronic hepatitis B. However, it remains unclear whether TDF-nucleoside analogue combination therapy provides better outcomes than TDF monotherapy. This study aimed to compare the efficacy of TDF monotherapy with that of TDF-nucleoside analogue combination therapy in patients with drug-resistant chronic hepatitis B. METHODS: This retrospective cohort study included 76 patients receiving TDF-based rescue therapy for more than 12 months. Suboptimal response was defined as serum HBV-DNA level of >60 IU/mL during prior rescue therapy. Multi-drug resistance was defined as the presence of two or more drug resistance-related mutations confirmed by mutation detection assay. The relationship between baseline characteristics and virologic response (HBV DNA <20 IU/mL) at 12 months were evaluated using logistic regression analysis. RESULTS: Fifty-five patients (72.4%) were suboptimal responders to prior rescue therapy, and 26 (34.2%) had multi-drug resistance. Forty-two patients (55.3%) received combination therapy with nucleoside analogues. Virologic response at 12 months was not significantly different between the TDF monotherapy group and TDF-nucleoside analogue combination therapy group (p=0.098). The serum HBV DNA level was reduced to -4.49+/-1.67 log10 IU/mL in the TDF monotherapy group and to -3.97+/-1.69 log10 IU/mL in the TDF-nucleoside analogue combination therapy group at 12 months (p=0.18). In multivariate analysis, female sex (p=0.032), low baseline HBV-DNA level (p=0.013), and TDF monotherapy (p=0.046) were predictive factors for virologic response at 12 months. CONCLUSIONS: TDF monotherapy showed similar efficacy to that of TDF-nucleoside analogue combination therapy in patients with drug-resistant chronic hepatitis B.
Adult ; Aged ; Antiviral Agents/pharmacology/*therapeutic use ; Cohort Studies ; DNA, Viral/blood ; Drug Resistance, Viral ; Drug Therapy, Combination ; Female ; Hepatitis B virus/drug effects/genetics/isolation & purification ; Hepatitis B, Chronic/*drug therapy/virology ; Humans ; Logistic Models ; Male ; Middle Aged ; Multivariate Analysis ; Nucleosides/chemistry/therapeutic use ; Retrospective Studies ; Sex Factors ; Tenofovir/*therapeutic use ; Treatment Outcome ; Young Adult

In order to investigate the mutation of HBV polymerase gene reverse transcription conserved region (P region) in chronic hepatitis B (CHB) patients, 212 CHB patients who took antiretroviral treatment with nucleotide analogues were chosen. The drug resistance mutations of HBV P region and HBV genotype were detected by Pyrosequencing. Sequence analysis showed that the drug resistance sites of HBV P region located at sites 173; 180; 181; 184; 204; 236 and 250. The main site of HBV P region drug resistance was 204 and 180, accounting for 35.8% and 23.5%, respectively. There were significant differences in the mutation rate of site 180 among different age groups. There were also significant differences in the mutation rate of site 204 among younger than 30 age group, 41 to 50 age group and 51 to 60 age group. (P < 0.05, P < 0.01). The mutation rate of site 180 combined with site 204 was 66.6%. The mutation rate of site 181 combined with site 236 was 23.3%. The age of C genotype infected patients was significantly older than B genotype infected patients (P < 0.01). M204V/I mutation mostly existed in the form of joint L180M mutation, the mutation rate was age-related. The detection of HBV genotypes and drug resistance sites of HBV P region have important clinical implications for the treatment and prognosis of patients with CHB.
Adult ; Aged ; Antiviral Agents ; pharmacology ; China ; Drug Resistance, Viral ; Female ; Gene Products, pol ; genetics ; Genotype ; Hepatitis B virus ; classification ; drug effects ; enzymology ; genetics ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Male ; Middle Aged ; Mutation, Missense ; Young Adult