Summary Chronic infection with the hepatitis B virus (HBV) affects an estimate of 240 million people worldwide despite the availability of a preventive vaccine. Medication to repress viral ...replication is available but a cure is rarely achieved. The narrow species and tissue tropism of the virus and the lack of reliable in vitro models and laboratory animals susceptible to HBV infection, have limited research progress in the past. As a result, several aspects of the HBV life cycle as well as the network of virus host interactions occurring during the infection are not yet understood. Only recently, the identification of the functional cellular receptor enabling HBV entry has opened new possibilities to establish innovative infection systems. Regarding the in vivo models of HBV infection, the classical reference was the chimpanzee. However, because of the strongly restricted use of great apes for HBV research, major efforts have focused on the development of mouse models of HBV replication and infection such as the generation of humanized mice. This review summarizes the animal and cell culture based models currently available for the study of HBV biology. We will discuss the benefits and caveats of each model and present a selection of the most important findings that have been retrieved from the respective systems.
Multiorgan and Renal Tropism of SARS-CoV-2 Puelles, Victor G; Lütgehetmann, Marc; Lindenmeyer, Maja T ...
The New England journal of medicine,
08/2020, Volume:
383, Issue:
6
Journal Article
Abstract Background & Aims Hepatitis E virus (HEV) is a major cause of acute hepatitis as well as chronic infection in immunocompromised individuals; however, in vivo infection models are limited. ...The aim of this study was to establish a small animal model to improve our understanding of HEV replication mechanisms and permit the development of effective therapeutics. Methods UPA/SCID/beige mice repopulated with primary human hepatocytes were used for infection experiments with HEV genotype (GT) 1 and 3. Virological parameters were determined at the serological and intrahepatic level by real time PCR, immunohistochemistry and RNA in situ hybridization. Results Establishment of HEV infection was achieved after intravenous injection of stool-derived virions and following co-housing with HEV-infected animals but not via inoculation of serum-derived HEV. GT 1 infection resulted in a rapid rise of viremia and high stable titres in serum, liver, bile and faeces of infected mice for more than 25 weeks. In contrast, viremia in GT 3 infected mice developed more slowly and displayed lower titres in all analysed tissues as compared to GT 1. HEV-infected human hepatocytes could be visualized using HEV ORF2 and ORF3 specific antibodies and HEV RNA in situ hybridization probes. Finally, six-week administration of ribavirin led to a strong reduction of viral replication in the serum and liver of GT 1 infected mice. Conclusion We established an efficient model of HEV infection to test the efficacy of antiviral agents and to exploit mechanisms of HEV replication and interaction with human hepatocytes in vivo.
No specific drugs are currently available against hepatitis delta virus (HDV), a defective virus leading to the most severe form of chronic viral hepatitis in man. The lack of convenient HDV ...infection models has hampered the development of effective therapeutics. In this study, naïve and hepatitis B virus (HBV) chronically infected humanized uPA/SCID mice were employed to establish a small animal model of HBV/HDV coinfection and superinfection. For preclinical antiviral drug evaluation, the GMP version of the myristoylated preS‐peptide (Myrcludex‐B), a lipopeptide derived from the pre‐S1 domain of the HBV envelope, was applied to prevent de novo HBV/HDV coinfection in vivo. Virological parameters were determined at serological and intrahepatic level both by real‐time polymerase chain reaction (PCR) and by immunohistochemistry. Establishment of HDV infection was highly efficient in both HBV‐infected and naïve chimeric mice with HDV titers rising up to 1 × 10E9 copies/mL. Notably, HDV superinfection led to a median 0.6log reduction of HBV viremia, which although not statistically significant suggests that HDV may hinder HBV replication. In the setting of HBV/HDV simultaneous infection, a majority of human hepatocytes stained HDAg‐positive long before HBV spreading was completed, confirming that HDV can replicate intrahepatically also in the absence of HBV infection. Furthermore, the increase of HBV viremia and intrahepatic cccDNA loads was significantly slower than in HBV mono‐infected mice. Treatment with the HBV entry inhibitor Myrcludex‐B, efficiently hindered the establishment of HDV infection in vivo. Conclusion: We established an efficient model of HBV/HDV infection to exploit mechanisms of viral interference in human hepatocytes and to test the efficacy of an HDV‐entry inhibitor in vivo. (HEPATOLOGY 2011)
BACKGROUND AND AIMS
To date, conflicting data exist as to whether hepatitis B virus (HBV) has the ability to induce innate immune responses. Here, we investigated cellular changes after the first ...contact between HBV and primary human hepatocytes (PHH) in vitro and in vivo.
APPROACH AND RESULTS
The exposure of PHH to HBV particles resulted in nuclear translocation of NFκB, followed by the expression and secretion of inflammatory cytokines (IL interleukin 1B, IL6, and TNF tumor necrosis factor). Ultraviolet irradiation of viral particles suppressed HBV infectivity but not the induction of cytokines in PHH, suggesting that the inoculum contains the immune‐inducing agent. Purified HBV particles on the whole, which were prepared from HBV DNA‐positive and protein‐rich fractions after heparin column separation, still had immune‐inducing capacity in PHH. The HBV‐induced gene expression profile was similar to that induced by toll‐like receptor 2 (TLR2) ligand Pam3Cys, but different from those induced by the viral sensors TLR3 or TLR7‐9. Treatment of PHH with both HBV particles and Pam3Cys led to phosphorylation of ERK (extracellular signal–regulated kinase), JNK, and p38 mitogen‐activated protein kinases as well as NFκB (nuclear factor kappa B). Finally, HBV‐induced gene expression could be neutralized by TLR2‐specific antibodies. Of note, pretreatment with an HBV entry inhibitor attenuated the TLR2‐mediated response to HBV, suggesting a receptor binding‐related mechanism. In liver‐humanized uPA/severe combined immunodeficient (SCID)/beige mice challenged with HBV in vivo, immune induction could only marginally be seen.
CONCLUSIONS
PHHs are able to sense HBV particles through TLR2, leading to an activation of anti‐HBV immune responses in vitro. These findings challenge the previously described stealth properties of HBV.
Chronic hepatitis B virus (HBV) infection has been associated with alterations in lipid metabolism. Moreover, the Na+‐taurocholate cotransporting polypeptide (NTCP), responsible for bile acid (BA) ...uptake into hepatocytes, was identified as the functional cellular receptor mediating HBV entry. The aim of the study was to determine whether HBV alters the liver metabolic profile by employing HBV‐infected and uninfected human liver chimeric mice. Humanized urokinase plasminogen activator/severe combined immunodeficiency mice were used to establish chronic HBV infection. Gene expression profiles were determined by real‐time polymerase chain reaction using primers specifically recognizing transcripts of either human or murine origin. Liver biopsy samples obtained from HBV‐chronic individuals were used to validate changes determined in mice. Besides modest changes in lipid metabolism, HBV‐infected mice displayed a significant enhancement of human cholesterol 7α‐hydroxylase (human hCYP7A1; median 12‐fold induction; P < 0.0001), the rate‐limiting enzyme promoting the conversion of cholesterol to BAs, and of genes involved in transcriptional regulation, biosynthesis, and uptake of cholesterol (human sterol‐regulatory element‐binding protein 2, human 3‐hydroxy‐3‐methylglutaryl‐coenzyme A reductase, and human low‐density lipoprotein receptor), compared to uninfected controls. Significant hCYP7A1 induction and reduction of human small heterodimer partner, the corepressor of hCYP7A1 transcription, was also confirmed in liver biopsies from HBV‐infected patients. Notably, administration of Myrcludex‐B, an entry inhibitor derived from the pre‐S1 domain of the HBV envelope, provoked a comparable murine CYP7A1 induction in uninfected mice, thus designating the pre‐S1 domain as the viral component triggering such metabolic alterations. Conclusion: Binding of HBV to NTCP limits its function, thus promoting compensatory BA synthesis and cholesterol provision. The intimate link determined between HBV and liver metabolism underlines the importance to exploit further metabolic pathways, as well as possible NTCP‐related viral‐drug interactions. (Hepatology 2014;60:1483–1493)
Chronic infection with hepatitis B virus (HBV) is caused by the persistence of closed circular DNA (cccDNA) in the nucleus of infected hepatocytes. Despite available therapeutic anti-HBV agents, ...eliminating the cccDNA remains challenging. Thus, quantifying and understanding the dynamics of cccDNA are essential for developing effective treatment strategies and new drugs. However, such study requires repeated liver biopsy to measure the intrahepatic cccDNA, which is basically not accepted because liver biopsy is potentially morbid and not common during hepatitis B treatment. We here aimed to develop a noninvasive method for quantifying cccDNA in the liver using surrogate markers in peripheral blood. We constructed a multiscale mathematical model that explicitly incorporates both intracellular and intercellular HBV infection processes. The model, based on age-structured partial differential equations, integrates experimental data from in vitro and in vivo investigations. By applying this model, we roughly predicted the amount and dynamics of intrahepatic cccDNA within a certain range using specific viral markers in serum samples, including HBV DNA, HBsAg, HBeAg, and HBcrAg. Our study represents a significant step towards advancing the understanding of chronic HBV infection. The noninvasive quantification of cccDNA using our proposed method holds promise for improving clinical analyses and treatment strategies. By comprehensively describing the interactions of all components involved in HBV infection, our multiscale mathematical model provides a valuable framework for further research and the development of targeted interventions.
Co-infection with hepatitis B (HBV) and D virus (HDV) is associated with the most severe course of liver disease. Interferon represents the only treatment currently approved. However, knowledge about ...the impact of interferons on HDV in human hepatocytes is scant. Aim was to assess the effect of pegylated interferon alpha (peg-IFNα) and lambda (peg-IFNλ), compared to the HBV-polymerase inhibitor entecavir (ETV) on all HDV infection markers using human liver chimeric mice and novel HDV strand-specific qRT-PCR and RNA in situ hybridization assays, which enable intrahepatic detection of HDV RNA species. Peg-IFNα and peg-IFNλ reduced HDV viremia (1.4 log and 1.2 log, respectively) and serum HBsAg levels (0.9-log and 0.4-log, respectively). Intrahepatic quantification of genomic and antigenomic HDV RNAs revealed a median ratio of 22:1 in untreated mice, resembling levels determined in HBV/HDV infected patients. Both IFNs greatly reduced intrahepatic levels of genomic and antigenomic HDV RNA, increasing the amounts of HDAg- and antigenomic RNA-negative hepatocytes. ETV-mediated suppression of HBV replication (2.1-log) did not significantly affect HBsAg levels, HDV productivity and/or release. In humanized mice lacking adaptive immunity, IFNs but not ETV suppressed HDV. Viremia decrease reflected the intrahepatic reduction of all HDV markers, including the antigenomic template, suggesting that intracellular HDV clearance is achievable.
Chronic hepatitis B virus (HBV) infection continues to be a major health burden worldwide; it can cause various degrees of liver damage and is strongly associated with the development of liver ...cirrhosis and hepatocellular carcinoma. The molecular mechanisms determining HBV persistence are not fully understood, but these appear to be multifactorial and the unique replication strategy employed by HBV enables its maintenance in infected hepatocytes. Both the stability of the HBV genome, which forms a stable minichromosome, the covalently closed circular DNA (cccDNA) in the hepatocyte nucleus, and the inability of the immune system to resolve chronic HBV infection are believed to be key mechanisms of HBV chronicity. Since a true cure of HBV requires clearance of intranuclear cccDNA from infected hepatocytes, understanding the mechanisms involved in cccDNA biogenesis, regulation and stability is mandatory to achieve HBV eradication. This review will summarize the state of knowledge on these mechanisms including the impact of current treatments on the cccDNA stability and activity. We will focus on events challenging cccDNA persistence in dividing hepatocytes.
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