Elsevier

Journal of Theoretical Biology

Volume 455, 14 October 2018, Pages 269-280
Journal of Theoretical Biology

The dynamics of integration, viral suppression and cell-cell transmission in the development of occult Hepatitis B virus infection

https://doi.org/10.1016/j.jtbi.2018.06.020Get rights and content

Highlights

  • •

    Viral suppression is a major but not the only contributor in the development of OBI.

  • •

    Viral suppression of at least 99% from peak production values enables the development of OBI.

  • •

    Integrated DNA are mostly absent or silent during OBI.

  • •

    OBI seropositive cases are more likely to progress into liver cirrhosis compared to OBI seronegative cases.

  • •

    Cell-to-cell transmission is the primary mechanism of infection transmission during OBI.

Abstract

Background

Out of several phases of HBV infection, the least understood phase is occult hepatitis B virus infection. The paucity of data due to non-availability of biological tissues and the prerequisite of ultra-sensitive assays for the detection of occult hepatitis B virus infection prompted us to utilize mathematical modeling in determining mechanisms that lead to occult hepatitis B virus infection and characteristics of HBV infection during occult hepatitis B virus infection.

Methods

We proposed two mathematical models (M1 and M2), considering two different phenomenon for episomal maintenance and accumulation of covalently closed circular DNA (cccDNA) in infected hepatocytes: (i) M1 – recirculation of the relaxed circular DNA/double-stranded linear DNA from cytoplasm to the nucleus, and (ii) M2 – reinfection of infected hepatocytes with virions. We further incorporated the dynamics of integrated Hepatitis B virus DNA (iHBV) to investigate its role in the development of occult hepatitis B virus infection.

Results

The analysis showed that the main mechanism for the spread of infection during occult hepatitis B virus infection is cell-to-cell transmission and not cell-free virus transmission. A significant viral suppression (of at least 99% from its peak production values) was essential but not sufficient in the development of occult hepatitis B virus infection under M1; however under M2, the viral suppression was neither sufficient nor essential as the inhibition of the production of HBsAg without viral suppression can also explain the development of occult hepatitis B virus infection. Our analysis also revealed that occult hepatitis B virus infection seropositive cases are more likely to progress into liver cirrhosis compared to occult hepatitis B virus infection seronegative cases. The iHBV was found to be mostly silent (by either being absent or non-productive for HBsAg) during occult hepatitis B virus infection.

Conclusion

The viral suppression is neither essential nor sufficient to explain the development of occult hepatitis B virus infection on its own. Not only the viral suppression but the inhibition -of the production and the export of HBsAg from cccDNA and iHBV also plays an important role in the development of occult hepatitis B virus infection. This is the first study, which incorporates the dynamics of iHBV and shows that HBV primarily spreads via cell-cell transmission during occult hepatitis B virus infection.

Introduction

Occult Hepatitis B virus (HBV) infection is defined as the long-lasting persistence of viral genetic material in the liver tissue of individuals with negativity for Hepatitis B surface antigen (HBsAg) with or without detectable HBV DNA in the serum (Pollicino and Raimondo, 2014, Raimondo et al., 2007). In the last decade, the understanding of occult hepatitis B virus infection has improved significantly largely due to concentrated efforts and technological advancements (Gutierrez-Garcia et al., 2011, Honarkar et al., 2004, Kwak and Kim, 2014, Martinez et al., 2015, Nishikawa and Osaki, 2013, Pollicino and Raimondo, 2014, Raimondo et al., 2007, Raimondo et al., 2013, Raimondo et al., 2008, Samal et al., 2012, Shire and Roberts, 2011, Squadrito et al., 2014, Zobeiri, 2013). Resultantly, occult hepatitis B virus infection in humans is further characterized into two categories as (i) occult hepatitis B virus infection seronegative (negative for anti-HBc and anti-HBs, termed as OBIAb-ve), and (ii) occult hepatitis B virus infection seropositive (positive for anti-HBc and anti-HBs, termed as OBIAb + ve). Additionally, “anti-HBc alone” where an individual is positive for anti-HBc, but negative for both HBsAg and anti-HBs is suspected (but not confirmed as it could also imply acute or resolved infection) as an atypical possible scenario of occult hepatitis B virus infection (Shire and Roberts, 2011, Wu et al., 2017a). In the ever-evolving natural history of HBV, occult hepatitis B virus infection is gaining recognition as one of the possible phases of chronic HBV infection (CHB) (Chemin and Trepo, 2005, McMahon, 2010, Pollicino and Raimondo, 2014). Although we have a good understanding of HBV life cycle and several phases in its natural progression (Ganem and Prince, 2004, McMahon, 2009), we still do not fully comprehended occult hepatitis B virus infection due to the lack of detectable serum biomarkers in this phase, in addition to the fact that there is an absolute need of invasive liver biopsy to confirm occult hepatitis B virus infection cases. For example, the reasons for the absence of HBsAg in the serum despite the presence of HBV genome/genetic material in the liver at low levels are not fully understood yet (Morales-Romero et al., 2014). occult hepatitis B virus infection is not only a challenge in clinical settings but also a serious issue in epidemiological settings as it stays below the radar and thus, the true HBV prevalence stays underestimated (Morales-Romero et al., 2014). This issue is further attenuated by the fact that infections with such low levels of HBV DNA during occult hepatitis B virus infection can still act as a source of infection transmission through blood transfusion and transplantation (Pollicino and Raimondo, 2014). Despite no signs of liver disease/damage in most of occult hepatitis B virus infection cases (Morales-Romero et al., 2014), occult hepatitis B virus infection still silently culminates in hepatocellular carcinoma, with or without cirrhosis but not in such high proportions as that of CHB (Pollicino and Raimondo, 2014).

Most importantly, mechanisms that can lead to occult hepatitis B virus infection are just speculative and require intense research (Morales-Romero et al., 2014). The list of such mechanisms include: (i) viral DNA integration, (ii) “a” determinant mutations leading to inefficient production of small-Hepatitis B surface antigen (S-HBsAg), (iii) the time elapsed since acute infection, (iv) a significant suppression of HBV replication (viral suppression), (v) immune factors such as non-cytolytic innate immune responses, (vi) genetic and epigenetic factors such as DNA methylation, which in large resembles like a significant viral suppression, (vii) coinfection with other viruses such as Hepatitis C virus (HCV) and human immunodeficiency virus (HIV), which also resembles like viral suppression, (viii) mutations in transcription-controlling regions of the polymerase, which resembles like points (ii) and (iv) together, and lastly, (ix) long half-life of hepatocytes and covalently closed-circular DNA (cccDNA) (Allweiss and Dandri, 2017, Hollinger and Sood, 2010, Huang et al., 2017, Kaur et al., 2010, Lucifora and Protzer, 2016, Morales-Romero et al., 2014, Pollicino and Raimondo, 2014, Pollicino et al., 2009).

In this article, we investigated the contribution of any combination of above-mentioned mechanism(s) in the transformation of acute HBV infection into occult hepatitis B virus infection. Investigations were conducted by employing mathematical forms of the HBV life cycle and experimental data sets, under one assumption that assays are ultra-sensitive and robust enough to detect occult hepatitis B virus infection without any misdiagnosis including no cases of false-positivity and false-negativity. Just like other viruses (Ciupe et al., 2007, Goyal and Murray, 2016, Goyal et al., 2017, Graw and Perelson, 2016, Ribeiro et al., 2010, Whalley et al., 2001), mathematical modeling can also play a vital role in the investigation of occult hepatitis B virus infection because (i) designing and conducting in vivo occult hepatitis B virus infection experiments is a time and resource consuming process as occult hepatitis B virus infection is a later phase of HBV infection which can take years to establish and even then it may not actually occur, and (ii) results from in vitro and in vivo experiments often varies from the human data (Allweiss and Dandri, 2017), and (iii) ultrasensitive molecular assays are unavailable, which could detect HBV at each and every step of the replication cycle. To the best of our knowledge, this is the first study that investigates occult hepatitis B virus infection from a modelling viewpoint.

Section snippets

Materials and methods

In this manuscript, we use the terminology: (i) infected hepatocytes: hepatocytes that have cccDNA, (ii) non-infected hepatocytes: hepatocytes with no cccDNA, and (iii) no productive integrated viral DNA: this implies that either there is no integrated DNA or if there is integrated DNA then there is no HBsAg production from them.

Significant viral suppression and no productive integrated viral DNA in non-infected hepatocytes is necessary to explain occult hepatitis B virus infection when the recirculation of nucleocapsids is the reason for cccDNA accumulation (Model 1)

From experimental observations, we have 0.17≥I*I#≥10−3 andV#V*≥105, and using them in condition (2) yields R#R*(1−e−λS#)(1−e−λS*)≥102, which is essential for PAHB transformation into occult hepatitis B virus infection. Therefore, mechanism(s) that can lead PAHB to occult hepatitis B virus infection are summarized as following (Fig. 2),

  • (i)

    By “a” determinant mutations: There is a need for a significant inhibition of the production of HBsAg from integrated viral DNA in infected hepatocytes and cccDNA

Discussion

The two hallmarks of occult Hepatitis B virus infection are serological negativity and the presence of miniscule HBV DNA loads (< 200 IU/ml) in the liver (Morales-Romero et al., 2014, Pollicino and Raimondo, 2014). It has been reported that out of all occult hepatitis B virus infection cases, there are about 78% positive for anti-HBc and anti-HBs (or, seropositive cases or OBIAb + ve) while the rest being negative for anti-HBc and anti-HBs (or, seronegative cases or OBIAb-ve) (Ocana et al., 2011

Acknowledgements

This work was supported by National Institutes of Health (Grant numbers R01-AI116868, R01-AI028433 and R01-OD011095). Portions of this work were performed under the auspices of the U.S. Department of Energy under contract DE-AC52-06NA25396. The funding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.

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