Level Of Hepatitis B Virus Replication Research Articles

Decreased expression of HBV surface antigen (HBsAg) with sK122R and sV96A co-mutation is associated with an ineffective antibody response in a chronic hepatitis B patient.

Emergence and predominance of hepatitis B virus (HBV) variants carrying S gene mutations frequently occur in HBV-infected individuals. Here, coexistent serum anti-HBsAg antibody (HBsAb) and HBV surface antigen (HBsAg) were detected in a chronic HBV patient. The patient's HBsAg proteins possessed amino acid substitutions sK122R and sV96A. We reported this case and conducted relevant studies to investigate differences in expression levels and antibody neutralization of HBsAg proteins bearing sK122R and sV96A amino acid substitutions to explore causes of antigen-antibody coexistence in a chronic hepatitis B patient. We first sequenced the S gene from HBV present within the patient's serum. Based on the S gene sequence, we cloned wild-type and mutated S gene sequences via site-directed mutagenesis to construct expression plasmids pJW4303-WT (wild-type), pJW4303-sV96A, pJW4303-sK122R, and pJW4303-sV96A-sK122R. Plasmids were transfected into HEK 293T cells then culture supernatants and cells were collected. Collected cells and supernatants were next subjected to a series of quantitative and functional tests to assess expression and neutralization characteristics of wild-type and mutant HBsAg proteins. Based on quantification of HBsAg expression in cells transfected with the four plasmids, HBsAg-sK122R-sV96A was more intracellularly retained and less secreted than HBsAg-sV96A single-mutant protein and WT. Neutralization ability of serum from chronic HBV patient against culture supernatants containing recombinant HBsAg proteins were ranked from highest to lowest as HBsAg-sV96A, HBsAg-sV96A-sK122R, and HBsAg-sK122R. However, no significant differences of neutralization efficiency by high-potency antibodies from HBV-vaccinees against these three mutant proteins were observed. The levels of HBsAg proteins with amino acid substitutions sV96A-sK122R were greatly reduced in culture supernatants but were apparently increased in the intracellular fraction. This may account for the higher levels of HBV replication in patients. HBsAg neutralization by HBsAb in this patient may have been compromised by the HBsAg sK122R amino acid substitution, suggesting that antibodies produced by the patient had lost their HBV-neutralizing effect.

Hepatitis B virus associated cirrhosis: overview of prognostic and management

Hepatitis B virus (HBV) infection is a global public health problem ; two billion people worldwide have evidence of past or present infection with HBV, and 296 million individuals are chronic carriers (i.e., positive for hepatitis B surface antigen [HBsAg]), of whom approximately 887,000 dies annually from HBV-related liver disease: chronic hepatitis B-associated cirrhosis and hepatocellular carcinoma. There are described some risk factors to the development of cirrhosis to HBV-infected persons: alcohol consumption, HBeAg status, metabolic syndrome, HBV genotypes and variants, and the level of HBV replication. The observation of a strong association between the development and decompensation of cirrhosis as well as between the development of HCC and the level of HBV replication suggests that suppression of HBV replication by long-term antiviral treatment may decrease the risk of complications in patients with HBV-related cirrhosis. The indication for antiviral treatment is given if any HBV DNA levels are detectable in the serum of HBV-infected patients with cirrhosis. Into the new era of antiviral treatment with nucleos(t)ide analogs we can prevent the development of cirrhosis in patients with chronic HBV infection and hepatic decompensation in many patients with HBV-related cirrhosis.

Biology and molecular pathogenesis of hepatitis B virus infection

Hepatitis B virus (HBV) remains a global public health infection. Over the past several decades, the basic principles of HBV gene expression and replication as well as the viral and host determinants governing infection outcomes have been largely uncovered. The clinical manifestations of HBV infection vary from an acute and chronic form of the disease. During the acute phase of the infection, the disease manifestations vary from subclinical hepatitis to anicteric hepatitis, icteric hepatitis, and fulminant hepatitis while during the chronic infection, manifest in different from ranging from asymptomatic carrier state to chronic hepatitis, cirrhosis, and hepatocellular carcinoma. The clinical outcome of the infection depends upon the level of HBV replication, age at infection, and the immune status of the host. HBV integration into the host genome often serves as a relevant source of hepatitis B surface antigen (HBsAg) expression during chronic infection, possibly triggering an immune response. If HBV is not eliminated, a delicate balance between viral replication and immune defence prevails which may lead to chronic hepatitis and liver cirrhosis. It is generally acknowledged that the humoral antibody response contributes to the clearance of circulating virus particles and the prevention of viral spread within the host while the cellular immune response eliminates infected cells.

Alinity m, a Random-Access System, for Hepatitis B Virus DNA Quantification in Plasma and Whole Blood Collected on Dried Blood Spots.

ABSTRACTThe International Liver Association recommends the use of accurate and sensitive molecular methods for determination of hepatitis B virus (HBV) DNA levels in plasma or serum of chronic HBsAg carriers. The level of HBV replication represents the strongest predictive biomarker associated with disease progression and long-term outcome of chronic HBV infection. The purpose of this study was to evaluate the ability to the new Alinity m System to detect and quantify HBV DNA in plasma and whole blood collected on dried blood spots (DBS). Paired plasma and DBS samples from patients chronically infected with various HBV genotypes were tested in parallel for HBV DNA detection and quantification. There is a linear relationship between HBV DNA levels measured in plasma samples using the Alinity m HBV assay and the Xpert HBV viral load assay, used for comparison. A slight deviation (0.03 ± 0.31 log IU/mL) was observed within the quantitative range. In DBS, HBV DNA levels closely correlated with levels measured in plasma. All patients had detectable and quantifiable HBV DNA by DBS testing, except for one patient with a plasma HBV DNA level above 2,000 IU/mL. In conclusion, the newly developed real-time PCR-based assay Alinity m HBV assay can correctly detect HBV DNA in DBS, especially for patients with blood HBV DNA levels above 2,000 IU/mL, and also accurately quantify HBV DNA in plasma samples.IMPORTANCE Hepatitis B virus is one of the most prevalent blood-borne viruses affecting the liver and causing acute and chronic hepatitis. Only a small proportion of people with HBV infection are diagnosed. HBV DNA measurement is critical in clinical practice for the diagnosis and treatment decisions of patients requiring antiviral therapy. Dried blood spot (DBS) collection provides a simple, practical, and acceptable alternative to venous blood collection, especially in community settings. We have demonstrated high sensitivity and specificity for HBV DNA detection in DBS compared to plasma samples, especially when using clinically relevant cutoffs of 2,000 and 20,000 IU/mL. Results support the use of DBS in community-based settings.

Canonical and Divergent N-Terminal HBx Isoform Proteins Unveiled: Characteristics and Roles during HBV Replication.

Hepatitis B virus (HBV) X protein (HBx) is a viral regulatory and multifunctional protein. It is well-known that the canonical HBx reading frame bears two phylogenetically conserved internal in-frame translational initiation codons at Met2 and Met3, thus possibly generating divergent N-terminal smaller isoforms during translation. Here, we demonstrate that the three distinct HBx isoforms are generated from the ectopically expressed HBV HBx gene, named XF (full-length), XM (medium-length), and XS (short-length); they display different subcellular localizations when expressed individually in cultured hepatoma cells. Particularly, the smallest HBx isoform, XS, displayed a predominantly cytoplasmic localization. To study HBx proteins during viral replication, we performed site-directed mutagenesis to target the individual or combinatorial expression of the HBx isoforms within the HBV viral backbone (full viral genome). Our results indicate that of all HBx isoforms, only the smallest HBx isoform, XS, can restore WT levels of HBV replication, and bind to the viral mini chromosome, thereby establishing an active chromatin state, highlighting its crucial activities during HBV replication. Intriguingly, we found that sequences of HBV HBx genotype H are devoid of the conserved Met3 position, and therefore HBV genotype H infection is naturally silent for the expression of the HBx XS isoform. Finally, we found that the HBx XM (medium-length) isoform shares significant sequence similarity with the N-terminus domain of the COMMD8 protein, a member of the copper metabolism MURR1 domain-containing (COMMD) protein family. This novel finding might facilitate studies on the phylogenetic origin of the HBV X protein. The identification and functional characterization of its isoforms will shift the paradigm by changing the concept of HBx from being a unique, canonical, and multifunctional protein toward the occurrence of different HBx isoforms, carrying out different overlapping functions at different subcellular localizations during HBV genome replication. Significantly, our current work unveils new crucial HBV targets to study for potential antiviral research, and human virus pathogenesis.

Knockdown of Virus Antigen Expression Increases Therapeutic Vaccine Efficacy in High-Titer Hepatitis B Virus Carrier Mice

Hepatitis B virus (HBV) infection persists because the virus-specific immune response is dysfunctional. Therapeutic vaccines might be used to end immune tolerance to the virus in patients with chronic infection, but these have not been effective in patients so far. In patients with chronic HBV infection, high levels of virus antigens might prevent induction of HBV-specific immune responses. We investigated whether knocking down expression levels of HBV antigens in liver might increase the efficacy of HBV vaccines in mice. We performed studies with male C57BL/6 mice that persistently replicate HBV (genotype D, serotype ayw)-either from a transgene or after infection with an adeno-associated virus that transferred an overlength HBV genome-and expressed HB surface antigen at levels relevant to patients. Small hairpin or small interfering (si)RNAs against the common 3'-end of all HBV transcripts were used to knock down antigen expression in mouse hepatocytes. siRNAs were chemically stabilized and conjugated to N-acetylgalactosamine to increase liver uptake. Control mice were given either entecavir or non-HBV-specific siRNAs and vaccine components. Eight to 12 weeks later, mice were immunized twice with a mixture of adjuvanted HBV S and core antigen, followed by a modified Vaccinia virus Ankara vector to induce HBV-specific B- and T-cell responses. Serum and liver samples were collected and analyzed for HBV-specific immune responses, liver damage, and viral parameters. In both models of HBV infection, mice that express hepatocyte-specific small hairpin RNAs or that were given subcutaneous injections of siRNAs had reduced levels of HBV antigens, HBV replication, and viremia (1-3 log10 reduction) compared to mice given control RNAs. Vaccination induced production of HBV-neutralizing antibodies and increased numbers and functionality of HBV-specific, CD8+ T cells in mice with low, but not in mice with high, levels of HBV antigen. Mice with initially high titers of HBV and knockdown of HBV antigen expression, but not mice with reduced viremia after administration of entecavir, developed polyfunctional, HBV-specific CD8+ T cells, and HBV was eliminated. In mice with high levels of HBV replication, knockdown of HBV antigen expression along with a therapeutic vaccination strategy, but not knockdown alone, increased numbers of effector T cells and eliminated the virus. These findings indicate that high titers of virus antigens reduce the efficacy of therapeutic vaccination. Anti-HBV siRNAs and therapeutic vaccines are each being tested in clinical trials-their combination might cure chronic HBV infection.

The regulation of HBx in hepatitis B virus replication

Hepatocellular carcinoma (HCC) is one of the most aggressive human malignancies in the world, which has been proven to be highly associated with a background of chronic and persistent infection of hepatitis B virus(HBV). HBV is an enveloped, partially double-stranded DNA virus that belongs to the Hepadnaviridae family of viruses. The level of HBV replication in a chronically HBV-infected individual can vary throughout the infection, and the rise and fall of HBV replication during chronic infection may affect disease progression. In the process of HBV infection, the replication of HBV DNA, the synthesis of viral proteins (HBp, HBc, HBs, HBx), and the packaging and releasing of new virus particles are performed. HBx protein is translated from HBx mRNA which transcribed by cccDNA and has a total length of 154 amino acids. Based on a brief description of the structure and distribution of HBx protein, the effects of HBx on HBV replication are reviewed. Once HBV infection of hepatocytes cell, the viruses first bind to receptors on the cell membrane, following by entering the cytoplasm in a manner similar to transportation of vesicles, then the viral removed the envelope and genome DNA is transferred into the nucleus. The DNA through the repairing mechanism generates cccDNA, which is transcribed and translated into viral mRNAs and proteins whose finally are packaged together to form new viral particles. It can be seen that the generation of cccDNA is the key for the replication of viral DNA and the formation of persistent infection. cccDNA exists in the nucleus independently of genomic DNA, but it is similar to genomic DNA and binds a large number of histones and non-histones, which are regulated by a variety of epigenetics such as methylation, acetylation and ubiquitination, to affect the transcription efficiency of cccDNA. Until now, the molecular mechanisms related to HBV replication and associated liver diseases have not been well understood. Currently, cumulative evidence indicates that the methylation, acetylation, and ubiquitination have vital roles in HBV replication as well as virus-related pathogenesis. This review aims to summarize advances in the study of HBx in terms of regulating the HBV replication through its mediating protein methylation, acetylation, deacetylation and ubiquitination. At the same time, miRNAs also play an important role in the regulation of HBx on HBV DNA replication. miRNAs are a type of small regulatory RNAs that consist of about 22 nt nucleotides and can not be translated into proteins. In organisms, miRNA involves in a variety of life activities and plays a role of the regulation of complementary mRNA at the post-transcription level. It is identified that HBx can regulate viral DNA replication by influencing the expression of target genes through miR-122, miR-15b, miR-125a, miR-192-3p, miR-539 and so on. However, HBx, acting as trans-activating factors, interacts with transcription factors to regulate the transcription of miRNAs. These transcription factors mainly include TBP, NF-κB, P53, c-Myc, CREB1 and so on. To sum up, we review the mechanism of HBx on HBV replication to provide ideas for the accurate diagnosis and treatment of HBV-induced hepatocellular carcinoma.

ЛАТЕНТНАЯ ФОРМА ИНФЕКЦИИ, ВЫЗВАННАЯ ВИРУСОМ ГЕПАТИТА B

This paper reviews the literature on occult hepatitis B virus (HBV) infection (OBI), discussing its definition, pathogenesis, diagnosis and prevention. OBI is characterized by a low level of HBV replication, when the viral DNA is detected in the liver at low concentrations (< 200 IU/ml) and may be undetectable in serum, while antibodies to the HBV core protein (anti-HBc) and/or to its surface protein (anti-HBs) may be present. In most cases, OBI is caused by the virus whose genome is replicatively competent and comparable to the genomes isolated from individuals with HBsAgpositive infection. Host factors are strongly involved in the induction and maintenance of an occult form of infection. Clinical recovery from HBV infection does not imply complete eradication of the virus, but only the ability of the immune system to keep reproduction of the remaining virus in the liver under control. Numerous clinical studies have shown that any immunosuppressive factors may trigger HBV reactivation producing a typical serological profile of active infection. The clinical significance of OBI is summarized by the following three key points: 1) the “occult” virus can be transmitted, mainly with blood transfusions or liver transplantation, followed by development of acute hepatitis B in the recipient; 2) HBV can get reactivated during immunosuppression, and 3) latent HBV infection contributes to the progression of other chronic liver diseases and plays a role in hepatocarcinogenesis. Testing for anti-HBc is a simple precautionary measure to prevent transmission of HBV during blood transfusion, especially in immunocompromised patients. This review is based on 60 publications, 3 Russian and 57 foreign, retrieved from by the following databases: PubMed, Google Scholar, Scopus, Springer, Cochrane Library, Wiley Online Library, and Russian Science Citation Index.

MiR-146 promotes HBV replication and expression by targeting ZEB2

Hepatitis B virus (HBV) is associated with the development of a wide spectrum of liver diseases. The involvement of miRNAs in HBV replication is being gradually identified. Among these miRNAs, miR-146a expression was found to be positively correlated with HBV replication levels. However, the regulatory relationship between miR-146a and HBV replication is still unclear. In the present study, miR-146a was upregulated in HBV-expressing HepG2.2.15 cells compared with HepG2 cells. Overexpression of miR-146a or knockdown of Zinc finger E-box-binding homeobox 2 (ZEB2) promoted HBV replication and expression, while downregulation of miR-146a or overexpression of ZEB2 suppressed HBV replication and expression. In addition, miR-146a was demonstrated to directly target ZEB2. Furthermore, ZEB2 silencing abated anti-miR-146a-induced inhibition on HBV replication and expression. These findings suggested that miR-146a promoted HBV replication by targeting ZEB2, providing a new antiviral strategy for HBV infection.

Immunotherapy for Chronic Hepatitis B Virus Infection.

While new therapies for chronic hepatitis C virus infection have delivered remarkable cure rates, curative therapies for chronic hepatitis B virus (HBV) infection remain a distant goal. Although current direct antiviral therapies are very efficient in controlling viral replication and limiting the progression to cirrhosis, these treatments require lifelong administration due to the frequent viral rebound upon treatment cessation, and immune modulation with interferon is only effective in a subgroup of patients. Specific immunotherapies can offer the possibility of eliminating or at least stably maintaining low levels of HBV replication under the control of a functional host antiviral response. Here, we review the development of immune cell therapy for HBV, highlighting the potential antiviral efficiency and potential toxicities in different groups of chronically infected HBV patients. We also discuss the chronic hepatitis B patient populations that best benefit from therapeutic immune interventions.

Hepatitis B virus covalently closed circular DNA homeostasis is independent of the lymphotoxin pathway during chronic HBV infection.

Current treatment options for patients with chronic hepatitis B virus (HBV) infection are not curative as they are not effective in eliminating covalently closed circular DNA (cccDNA). cccDNA is a stable template for HBV transcription in the nucleus of hepatocytes and is thought to be one of the main factors responsible for HBV persistence. Recently, activation of the lymphotoxin beta receptor (LTβR) has been shown to trigger degradation of cccDNA through induction of cytidine deaminases of the APOBEC3 family in HBV cell culture model systems. To assess the presence and relevance of such mechanisms in the liver of chronically HBV-infected patients, we compared intrahepatic cccDNA levels with the expression levels of lymphotoxins and some of their target genes (eg APOBEC deaminases) in liver biopsy tissue. Our results confirm elevated gene expression levels of components of the lymphotoxin pathway including lymphotoxin alpha (LTα), lymphotoxin beta (LTβ), APOBEC3B (A3B) and APOBEC3G (A3G) in the chronically HBV-infected liver compared to uninfected liver. Furthermore, expression levels of the genes of the APOBEC deaminase family were correlated with those of LTα and LTβ gene expression, consistent with lymphotoxin-mediated upregulation of APOBEC gene expression. However, intrahepatic cccDNA and HBV replication levels were not correlated with LTα, LTβ and APOBEC gene expression. In conclusion, these results suggest that although the lymphotoxin pathway is activated in the chronically HBV-infected liver, it has no major impact on HBV cccDNA metabolism in chronic HBV infection.

Hepatitis B Virus-Associated Hepatocellular Carcinoma.

Liver cancer is the fifth most common cancer worldwide in men and the ninth in women. It is also the second most common cause of cancer mortality. Hepatocellular carcinoma (HCC) is the most common type of liver cancer. About 350 million people globally are chronically infected with HBV. Chronic hepatitis B virus (HBV) infection accounts for at least 50% cases of HCC worldwide. Other non-HBV factors may increase HCC risk among persons with chronic HBV infection. Both indirect and direct mechanisms are involved in HCC oncogenesis by HBV. HCC-promoting HBV factors include long-lasting infection, high levels of HBV replication, HBV genotype, HBV integration, specific HBV mutants, and HBV-encoded oncoproteins (e.g., HBx and truncated preS2/S proteins). Recurrent liver inflammation caused by host immune responses during chronic HBV infection can lead to liver fibrosis and cirrhosis and accelerate hepatocyte turnover rate and promote accumulation of mutations. Major breakthroughs have been achieved in the prevention of HBV-associated HCC with HBV vaccines and antiviral therapies.

Zinc finger E-box-binding homeobox 2 inhibits hepatitis B virus replication and expression

Objective: To investigate the effect of zinc finger E-box-binding homeobox 2 (ZEB2) on hepatitis B virus (HBV) replication and expression. Methods: HepG2, HepG2.2.15, and HepAD38 cells were cultured separately, and Western blot was used to measure the expression of ZEB2. HepG2.2.15 cells were cultured and transfected with ZEB2 expression plasmids or shRNA targeting ZEB2. Western blot was used to measure the expression of ZEB2 and HBV core proteins, quantitative real-time PCR was used to measure HBV 3.5 kb RNA and HBV DNA, Southern blot was used to measure HBV replicative intermediate, and ELISA was used to measure the expression of HBsAg and HBeAg, in order to clarify the effect of ZEB2 on HBV replication and expression. The dual-luciferase reporter system was used to analyze the effect of ZEB2 on HBV promoter, and the chromatin immunoprecipitation assay was used to detect the binding of ZEB2 to HBV promoter. The t-test was used for comparison of means between groups. Results: The expression of ZEB2 was inhibited in the cells with HBV replication. Overexpression of ZEB2 reduced the level of HBV replication and expression by about 50% (P< 0.05). After ZEB2 was downregulated by shZEB2-1 or shZEB2-2, the level of HBV replicative intermediate increased from 58.53 ± 3.43 to 112.80 ± 5.03, and 128.30 ± 2.31, the relative expression level of HBV 3.5 kb RNA increased from 1.00 ± 0.01 to 2.03 ± 0.02 and 2.32 ± 0.03, the level of HBsAg increased from 35.63% ± 1.57% to 81.87% ± 0.43% and 100.00% ± 2.18%, and HBeAg increased from 37.00% ± 0.70% to 88.00% ± 2.60% and 100.00% ± 0.75%. Furthermore, ZEB2 could bind to HBV core promoter and inhibit its transcriptional activity. Conclusion: ZEB2 inhibits HBV replication and expression through binding to HBV core promoter and inhibiting its transcriptional activity.

Hepatitis B virus and its sexually transmitted infection - an update.

Epidemiology: incidence and prevalence: About 5% of the world’s population has chronic hepatitis B virus (HBV) infection, and nearly 25% of carriers develop chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). The prevalence of chronic HBV infection in human immunodeficiency virus (HIV)-infected individuals is 5%-15%; HIV/HBV coinfected individuals have a higher level of HBV replication, with higher rates of chronicity, reactivation, occult infection, and HCC than individuals with HBV only. The prevalence of HBV genotype A is significantly higher among men who have sex with men (MSM), compared with the rest of the population. Molecular mechanisms of infection, pathology, and symptomatology: HBV replication begins with entry into the hepatocyte. Sodium taurocholate cotransporting polypeptide was identified in 2012 as the entry receptor of HBV. Although chronic hepatitis B develops slowly, HIV/HBV coinfected individuals show more rapid progression to cirrhosis and HCC. Transmission and protection: The most common sources of HBV infection are body fluids. Hepatitis B (HB) vaccination is recommended for all children and adolescents, and all unvaccinated adults at risk for HBV infection (sexually active individuals such as MSM, individuals with occupational risk, and immunosuppressed individuals). Although HB vaccination can prevent clinical infections (hepatitis), it cannot prevent 100% of subclinical infections. Treatment and curability: The goal of treatment is reducing the risk of complications (cirrhosis and HCC). Pegylated interferon alfa and nucleos(t)ide analogues (NAs) are the current treatments for chronic HBV infection. NAs have improved the outcomes of patients with cirrhosis and HCC, and decreased the incidence of acute liver failure.

Sodium taurocholate cotransporting polypeptide inhibition efficiently blocks hepatitis B virus spread in mice with a humanized liver.

Sodium taurocholate cotransporting polypeptide (NTCP) is a recently discovered hepatitis B virus (HBV) receptor. In the present study, we used TK-NOG mice with a humanized liver to examine the impact of endogenous NTCP expression on HBV infection. Upon inoculation with HBV, these mice exhibited clear viremia in 2 weeks, and serum HBV DNA levels gradually increased. The frequency of HBsAg-positive hepatocytes in the liver was 5.1 ± 0.6% at 2 weeks and increased with increasing HBV DNA levels, reaching 92.9 ± 2.8% at 10 to 12 weeks. In vivo siRNA-mediated NTCP knockdown before and after HBV inoculation significantly suppressed the levels of HBV replication and the frequency of HBsAg-positive hepatocytes at 2 weeks, whereas NTCP knockdown 13 weeks after infection did not affect these parameters. Similar to the humanized mouse livers in the early phase of HBV infection, human liver samples from chronic hepatitis B patients, especially those treated with nucleos(t)ide analogues, contained a considerable number of hepatocytes that were negative for the anti-HBs antibody. In conclusion, NTCP inhibition prevents the spread of HBV-infected hepatocytes in mice with a humanized liver. NTCP-targeted therapy has potential for regulating HBV infection in patients with chronic hepatitis B.

Rethinking the pathogenesis of hepatitis B virus (HBV) infection.

Chronic hepatitis B virus (HBV) infection affects approximately 375 million people worldwide. Current antiviral treatment effectively controls, but rarely clears chronic HBV infection. In addition, a significant portion of chronic HBV infected patients are not suitable for currently available antiviral therapy, and still face higher risk for cirrhosis and hepatocellular carcinoma. The poorly understood pathogenesis of HBV infection is the main barrier for developing more effective treatment strategies. HBV has long been viewed as non-cytopathic and the central hypothesis for HBV pathogenesis lies in the belief that hepatitis B is a host specific immunity-mediated liver disease. However, this view has been challenged by the accumulating experimental and clinical data that support a model of cytopathic HBV replication. In this article we systematically review the pathogenic role of HBV replication in hepatitis B and suggest possible HBV replication related mechanisms for HBV-mediated liver injury. We propose that a full understanding of HBV pathogenesis should consider the following elements. I. Liver injury can be caused by high levels of HBV replication and accumulation of viral products in the infected hepatocytes. II. HBV infection can be either directly cytopathic, non-cytopathic, or a mix of both in an individual patient depending upon accumulation levels of viral products that are usually associated with HBV replication activity in individual infected hepatocytes.

Technical standards for hepatitis B virus X protein (HBx) research.

Chronic infection with hepatitis B virus (HBV) is a risk factor for developing hepatocellular carcinoma (HCC). The life cycle of HBV is complex and has been difficult to study because HBV does not infect cultured cells. The HBV regulatory X protein (HBx) controls the level of HBV replication and possesses an HCC cofactor role. Attempts to understand the mechanism(s) that underlie HBx effects on HBV replication and HBV-associated carcinogenesis have led to many reported HBx activities that are likely influenced by the assays used. This review summarizes experimental systems commonly used to study HBx functions, describes limitations of these experimental systems that should be considered, and suggests approaches for ensuring the biological relevance of HBx studies.

Dynamic correlation between induction of the expression of heme oxygenase-1 and hepatitis B viral replication.

Heme oxygenase‑1 (HO‑1) possesses significant potential as a drug target for hepatitis B, which may be transferable to patient therapy. The aim of the present study was to clarify the dynamic correlation between the hepatitis B virus (HBV) and HO‑1. The levels of HBV replication and expression of HO‑1 were investigated in HepG2.2.15 hepatoma cells following exposure to 5‑50µMhemin for 1‑6days. The mRNA expression levels of HO‑1 were then detected using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). HBV replication levels were determined by enzyme‑immunoassay and a PCR‑fluorescence quantitation assay. The results of the present study demonstrated that the mRNA expression levels of HO‑1 increased in a dose‑dependent manner in the HepG2.2.15 cells, following exposure to 5‑50µM hemin. The mRNA expression levels of HO‑1 reached a peak following exposure of the cells to hemin for threedays, subsequently the expression of HO‑1 decreased. Following exposure to hemin at an optimal concentration of 50µM for 1‑6days, the levels of the hepatitisB surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in the cells were significantly reduced. This marked reduction in the expression of HBsAg and HBeAg reached its peak on the first day, following which the inhibition weakened as the duration of exposure increased. In addition, the inhibition of HBV DNA replication increased with the a longer duration of exposure. Furthermore, HBV DNA levels were significantly decreased following exposure to hemin for 3‑6days. In conclusion, the present study demonstrated that induced expression of HO‑1 interfered with HBV replication in a dose and time‑dependent manner, implying that a reduction of the HBV viral load may contribute to upregulation in the expression of HO‑1.

Role of occult hepatitis B virus infection in chronic hepatitis C.

The development of sensitive assays to detect small amounts of hepatitis B virus (HBV) DNA has favored the identification of occult hepatitis B infection (OBI), a virological condition characterized by a low level of HBV replication with detectable levels of HBV DNA in liver tissue but an absence of detectable surface antigen of HBV (HBsAg) in serum. The gold standard to diagnose OBI is the detection of HBV DNA in the hepatocytes by highly sensitive and specific techniques, a diagnostic procedure requiring liver tissue to be tested and the use of non-standardized non-commercially available techniques. Consequently, in everyday clinical practice, the detection of anti-hepatitis B core antibody (anti-HBc) in serum of HBsAg-negative subjects is used as a surrogate marker to identify patients with OBI. In patients with chronic hepatitis C (CHC), OBI has been identified in nearly one-third of these cases. Considerable data suggest that OBI favors the increase of liver damage and the development of hepatocellular carcinoma (HCC) in patients with CHC. The data from other studies, however, indicate no influence of OBI on the natural history of CHC, particularly regarding the risk of developing HCC.

State of the art: hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the most common primary liver cancer, mainly occurring in cirrhotic livers (in 80–90% of cases) after years of chronic inflammation [1,2]. The pathogenesis of HCC is a multistep process associated with changes in the host gene expression of multiple redundant negative-growth regulatory pathways that protect cells against transformation. An imbalance between the proliferation and apoptosis of liver cells is thought to promote tumor development [3]. Annually, 3–6% of cirrhotic patients will develop HCC, mainly men with advanced liver disease. Recent trends in the USA show that men are affected three-times more frequently than women, Asians are affected twice as often as blacks and Hispanics, and blacks are affected twice as often as whites [4]. One explanation for the disproportionate effects of HCC in men may be due to the higher estrogen concentrations present in women, which suppress IL-6 production and inhibit chemically induced liver carcinogenesis [1]. Epidemiology & etiology HCC is the principal cause of death in patients with compensated cirrhosis worldwide. It is largely a problem of less developed regions, where 83% (50% in China alone) of the estimated 782,000 new cases of liver cell cancer worldwide occur (based on 2012 data) [5]. HCC is the fifth most common cancer in men (554,000 cases; 7.5% of the total) and the ninth in women (228,000 cases; 3.4% of the total) [5]. Regions of high incidence (age adjusted rates per 100,000 inhabitants per year in men) are eastern and south-eastern Asia (31.9 and 22.2, respectively). Intermediate rates occur in southern Europe (9.5) and North America (9.3), and the lowest rates are in northern Europe (4.6) and south-central Asia (3.7) [5]. The global age distribution of HCC cases is related to the dominant viral hepatitis in the underlying population and the age at which it was acquired. For example, the disease burden is highest in areas with endemic hepatitis B virus (HBV) infection, such as in sub-Saharan Africa, where the virus is mainly transmitted at birth (with >90% of these cases becoming chronic HBV carriers). However, the diagnosis of HCC often occurs later in North America and southern Europe, where the most common etiology is HBV or hepatitis C virus (HCV) acquired later in life [2]. Several factors have been reported to increase HCC risk among HBV carriers, including male sex, older age, Asian or African ethnicity, family history of HCC, viral (higher levels of HBV repli cation, HBV genotype, longer duration of infection or coinfection with HCV, HIV or hepatitis D virus), clinical (cirrhosis) and environmental or lifestyle factors (exposure to aflatoxin, chronic excessive alcohol consumption or tobacco smoking). Other risk factors that increase the risk in HCV-infected patients include coinfection with HIV, HBV, HCV genotype 1b, old age, the presence of diabetes and obesity and a high level of chronic alcohol consumption. However, both HBV and HCV are highly amenable to preventive measures with antivirals or vaccination (for HBV). In Italy, for example, the Italian Liver Cancer (ITA.LI.CA) database reveals a decline in HCV infection with a concomitant increase in alcohol consumption as the primary etiology for HCC [6].