Hepatitis B virus covalently closed circular DNA (HBV cccDNA) forms a mini-chromosome structure inside infected hepatocyte nuclei and plays an important role in chronic hepatitis B infection. Methylation of cccDNA-bound histone and the associations of heterochromatin HP1 complex related proteins with cccDNA were investigated in this thesis using transient transfection study system and chromatin immunoprecipitation assay. Di- and tri-methylation of histone H3 lysine 4 residue (H3K4), which plays an activating role in eukaryotic transcription, were found to associate with cccDNA in a way in parallel to the level of HBV replication in our system. On the other hand, tri-methylation of H3K9, which plays an inhibitory role in eukaryotic transcription, was found to associate with cccDNA during decline of HBV replication. During the decline of HBV replication, cccDNA was associated with histone methyltransferases SUV39H1 and SUV420H1 and histone demethylase PLU1. The dynamic of the association of heterochromatin protein 1 (HP1) to cccDNA was similar to that of SUV39H1. The association of cccDNA with five HP1 complex-related proteins (three DNA methyltransferases Dnmt3a, Dnmt3b and Dnmt1 and two methylated DNA binding proteins MBD1 and MeCP2) was studied, and their associations could be roughly divided into two stages. From 72 hours to 96 hours post-transfection, there was an increased association of cccDNA with Dnmt3a, Dnmt3b and MBD1, which was in parallel to the increased association of HP1 and SUV39H1with -cccDNA. From 96 hours to 120 hours after transfection, an increased association of Dnmt1 and MeCP2 with cccDNA was detected, which was correlated to that of SUV420H1. At the time when HBV replication was declining at 120 hours post-transfection, a highest association of SUV39H1, SUV420H1, HP1 and all 5 HP1 complex-related proteins with cccDNA was found. In conclusion, methylation of cccDNA-bound histone was associated with HBV replication. Activating H3K4 methylation was found to correlate with increase in HBV replication, while inhibitory H3K9 methylation correlated with decrease in HBV replication. The association of HP1 was in parallel to that of SUV39H1, indicating that HP1-SUV39H1 complex might be involved, and thereby recruiting other proteins for transcription suppression. Recruitment of DNA methyltransferases and methylated DNA binding proteins to cccDNA provided further evidence that methylation of cccDNA plays a role in transcription suppression. This study identified the associations of methylated histone and other related proteins with cccDNA and their correlations with viral replication. These results enhance our knowledge in HBV replication cycles and transcription regulation. It may show a novel area in development of antiviral drugs such as histone methyltransferase modulators. / published_or_final_version / Medicine / Master / Master of Philosophy

Background/Aim: Hepatitis C virus infection is a known risk factor of impaired glucose metabolism and diabetes mellitus. Whether hepatitis B virus (HBV) infection is also associated with impaired glucose tolerance remains uncertain. The aim of the study was to conduct a systematic review on the association between HBV infection and impaired glucose metabolism Methods: Studies reporting the association between HBV infection and markers of impaired glucose metabolism were identified through keyword search in PubMed and Google Scholar. 10 studies (out of 320) were included in this systematic review. Results were included. Majority (n=7) of the included studies were conducted among the Asian populations. Of the 10 included studies, eight studies reported a significant association between HBV infection and impaired glucose metabolism, proxied by impaired glucose tolerance, impaired fasting glucose, diabetes mellitus, insulin resistance, and metabolic syndromes. The remaining two studies using diabetes mellitus and insulin resistance as outcome measures did not find a positive association with HBV infection. Conclusions: The association between HBV and impaired glucose metabolism is suggestive from the evidence compiled from included articles. However, whether the development of glucose intolerance or diabetes mellitus is linked to an infectious cause of HBV is still inconclusive. Further studies that could improve on the current understanding of the associations between HBV infection and impaired glucose metabolism are necessary. / published_or_final_version / Public Health / Master / Master of Public Health

Introduction: China has about 120 million hepatitis B virus (HBV) carriers and a 7.2% hepatitis B surface antigen (HBsAg) prevalence in 2006.This creates a huge disease burden and also leads to significant economic losses. Since 2002, a free universal infant hepatitis B vaccination programme has provideda 3-dose primary vaccination for all infants. Although some economic evaluations of this programme have been conducted, a comprehensive cost-effectiveness analysis (CEA) to estimate long-term benefit using mathematical modeling would aid understanding of population strategies for hepatitis B control in large populations. Moreover, the most common mode of infection is perinataltransmission at birth. However the more effective immunization programme involving screening women during pregnancy for HBV-carrier status and providing passive-active vaccination for newborns has not been implemented in China. Aims: To identify the most cost-effective universal infant hepatitis B vaccination strategy for China. Method: A hospital-based survey was conducted during 2010-2011 in a general hospital in Shenzhen, China, in order to obtain costing data to estimate the economic burden of chronic hepatitis B patients. Annual direct and indirect costs from this study were used as cost parameters in the CEA models. Mathematical models were developed to simulate perinatal transmission, vaccination programmes and disease progression using Markov modeling and decision trees. Quality-adjusted life year (QALYs) as well as health and monetary outcomes were also assessed. Univariate sensitivity analysis and probabilistic sensitivity analysis using Monte Carlo simulation were performed to test parameter uncertainty. Two programmes of screening of pregnant women for both HBsAg and/or HBeAg and the infant passive-active vaccination were compared with the current vaccine-only programme in one CEA, while the other CEA estimated the effect of the current infant programme compared with no vaccination. Findings: The estimated total economic burden including annual direct and indirect cost among hepatitis B patients of RMB 43104.5 (US$6340.8). The economic burdens of associated disease states of hepatitis B infection were highest for hepatocellular carcinoma (HCC) (RMB 77297.1), decompensated cirrhosis (RMB 50725.7), chronic active hepatitis B (CAH) (RMB 37449.5) and finally compensated cirrhosis (RMB 37276.9). The average total economic burden per hepatitis B patient amounted to 46% of Shenzhen GDP per capitain 2010, and 5.4% of the city’s annual per capita income. The current vaccine-only infant vaccination programme was justified by costsavings, from both a societal and health care payer’s perspective, reducing new HBV infections by about 76%. This has produced a gain of 743,000 life-years and 620,000 QALYs given current numbers and savings of US$2~3billion saved over the lifetime of a national cohortof 10,000,000 newborns. A universal control programme involving the screening of pregnant women for HBsAg and passive-active vaccination, would reduce new infections by 13%, saving 436,000 life years and gaining 121,000 QALYs for a saving of about US$546 million compared with current vaccine-only programme. Implications: The universal infant hepatitis B vaccination programme is currently a cost-effective strategy for hepatitis B control in China.A beneficial amendment to the current strategy wouldinclude screening of all pregnant women for HBsAg and vaccinating newborns in a passive-active way. / published_or_final_version / Public Health / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / toc / Medicine / Master / Doctor of Medicine

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / Paediatrics / Master / Master of Philosophy

No description available.

The human hepatitis B virus (HBV) is a small hepatotropic virus, which affects approximately 350 million chronic sufferers worldwide. It has a compact 3.2 kbp dsDNA genome encoding four major overlapping genes namely core, polymerase, surface and X required for its replication. The virus synthesises a pregenomic RNA (pgRNA) which functions both as an RNA intermediate for reverse transcription into the DNA genome and as the mRNA for the translation of the core (C) and polymerase (P) proteins. The core overlaps the polymerase gene and is translated at a 10 to 1 ratio. The polymerase gene translated from the P AUG codon is preceded by at least 4 upstream AUG codons (uAUGs), namely C AUG, C1 AUG, J AUG and C2 AUG. Various mechanisms have been implicated in the synthesis of the polymerase protein. This led to the currently accepted model which involves leaky scanning and a reinitiation mechanism in polymerase synthesis. However, multiple sequence alignment of the pgRNA revealed a short upstream open reading frame (uORF) highly conserved at the nucleotide level in all HBV subtypes and mammalian hepadnaviruses. This previously unreported uORF, designated as C0 ORF in this study is also conserved in its position and length. Past studies have either omitted this uORF in their test constructs or ignored its potential role. The C0 ORF has a conserved weak initiation context and is located within the epsilon structure within the 5� leader of the pgRNA, required for viral encapsidation. Importantly, the C0 ORF precedes and overlaps the core ORF, which may suggest an alternative model in which the core and polymerase may be translated and coordinately regulated. Fusion of the C0 ORF to luciferase showed for the first time that this uORF is translated through the detection of reporter activity (~20% of C) and also visualisation of the fusion protein via western analysis using anti-C0 and anti-luciferase antibodies. Subsequent removal of the C0 ORF implicated a role in repressing downstream core fusion protein synthesis in HepG2 cells. A similar repression was observed on J expression. To study the effect of C0 on downstream polymerase translation, a pgRNA-like DNA construct was made and subsequent mutations introduced. Mutation of the C0 AUG led to an increase in initiation at the downstream P AUG. Alteration of the existing weak initiation context to an optimal context which favours stronger initiation consistently showed a potential role for C0 ORF in facilitating reinitiation at certain downstream initiation codons including P AUG. Mutations of other uAUGs preceding the P AUG were also done to better understand their roles in regulating polymerase synthesis. The removal of the C AUG markedly increased expression from the P AUG. This study revealed other internal uAUGs in-frame to the C AUG, namely the C1 and C2 AUGs are also effectively translated, further reducing availability of translating ribosomes to downstream P AUG. Indeed the removal of the C1 and C2 AUGs led to a corresponding increase in initiation from the P AUG. Initiation at the internal J AUG was also reported and its removal showed a significant decrease in expression from the P AUG, consistent with the previous model implicating reinitiation at the P initiation site after translation of the short J ORF. The inhibitory role of the 5 uAUGs prior to the P AUG were confirmed when all were removed, giving rise to translation almost equal to that at C AUG. Taken together, these results suggest a new model in which the HBV C0 ORF plays a key role in controlling core and polymerase synthesis by repressing core translation and making available more ribosomes to downstream AUGs possibly facilitating translation reinitiation. In addition, the translation of the C0 ORF across the [epsilon] region may also preclude encapsidation, potentially acting as a switch discriminating the pgRNA template between encapsidation and translation. Therefore, the highly conserved [epsilon] region and C0 ORF present an excellent target for molecular based antiviral drugs (antisense oligonucleotides, aptamers, ribozymes) potentially providing new anti HBV drugs.

Thesis (M Nursing)--University of South Australia, 1996

Diss. (sammanfattning) Stockholm : Karol. inst. / Härtill 6 uppsatser.