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HepG2 and Huh7 cells expressing HBV DNA are frequently can be utilized to understand the effects of natural HBV
used in the study of HBV biology. While HepG2 cells and infection. HepG2 cells expressing hNTCP (HepG2-hNTCP),
Huh7 cells support HBV replication, similar to nearly all theoretically, provide a convenient in vitro system for HBV
existing human liver cell lines, they cannot be directly infection. The exogenous expression of hNTCP in HepG2
infected with HBV, partly due to the low expression levels cells does render them susceptible to HBV infection;
of hNTCP, the functional cellular receptor for HBV. In however, low levels of infection, typically around 10%, and
[42]
order to analyze differences between cells with and a requirement for large viral inoculums limit their use.
without replicating HBV, HepG2.2.15 cells have sometimes Infection-based studies are hampered even further by the
been compared to HepG2 cells. HepG2.215 cells were low levels of virus released by HBV-infected cells, believed
originally derived from HepG2 cells and stably express to be around 1 virion per day, making it difficult to produce
HBV from two integrated head-to-tail dimers of the HBV the large quantities of virus needed for these types of
genome. [196] Results obtained by comparing HepG2.215 studies. Despite these issues, HepG2-hNTCP cells provide
cells to the parental HepG2 cells, however, need to be a valuable in vitro model system for elucidating the effects
interpreted with caution; because of the continuous of natural HBV infection, investigating the complete HBV
passaging of HepG2.215 cells since their development in life cycle including the early steps of an HBV infection, and
1987, dissimilarities beyond the expression of HBV may identifying novel therapeutic options. [42,103,172,198,202-204]
exist between HepG2.2.15 cells and the parental HepG2
cells. Due to these dissimilarities, phenotypic differences As the natural target of an HBV infection, primary human
that are observed between HepG2.215 and HepG2 cells hepatocytes would be the ideal in vitro system for
might not be an exclusive consequence of replicating HBV. studies of HBV. Unfortunately, cultured primary human
hepatocytes lose susceptibility to HBV infection within
Together, the use of exogenously delivered HBV DNA into days of isolation and culture, potentially because hNTCP
established cell lines, such as HepG2 and Huh7, and the expression rapidly decreases over time in culture. [42,198,205]
use of cell lines stably expressing HBV, such as the HepG2- Interestingly, Rice and colleagues recently reported that
derived cell lines HepG2.2.15 and HepAD38, [196,197] make induced pluripotent stem cell-derived hepatocytes(iPSC-
up the majority of the studies that have been conducted derived iHeps) can support HBV infection, opening
to understand HBV biology. While these cell culture potential new avenues to study HBV biology and virus-
models have proven extremely valuable to study HBV host interaction and to test antiviral candidates. However,
DNA replication, viral assembly, and virion secretion, they a long induction process involving differentiation of the
have limitations that prevent them from recapitulating all iPSCs is required prior to HBV infection, and viral markers
the aspects of an authentic human HBV infection. [198] of infection can only be detected more than a week after
inoculation. [198,205,206]
Some recent developments have lead to increased
optimism for the development of an effective in vitro Although studies in immortalized or transformed cells
system to study the complete HBV life cycle. For example, have served as powerful models for studying various
fusion of primary human hepatocytes with hypoxanthine- aspects of HBV replication and the functions of HBV-
guanine phosphoribosyltransferase null [HGPRT(-)] encoded proteins, these studies have also demonstrated
HepG2 cells led to the establishment of the immortal cell that HBV-mediated activities, particularly those
line, HepCHLine-4/-7, that may provide a model system associated with HBV proteins such as HBx, may vary in
for HBV infection. This cell line supports HBV replication different cellular contexts. [92,207] Alternatively, studies in
and is susceptible to HBV infection when incubated with cultured primary hepatocytes have begun to clarify HBV
serum from HBV-positive patients. [199,200] However, an replication strategies and the function of HBV proteins
uncertain genetic stability during maintenance hampers in a more relevant context. Recently, cultured primary
[92]
the use of this system. [198] In addition, the HepaRG cell rat hepatocytes have been used to study HBV replication
line, a human hepatoma cell line, can also be directly and functions of the HBx protein; [83-86,208] HBx activities
infected with HBV and supports HBV replication. While in cultured primary rat hepatocytes were similar to
this cell line is often used in studies of HBV infection, its HBx activities in cultured primary human hepatocytes,
use is limited by a low HBV infection efficiency of only supporting the use of cultured primary rat hepatocytes
about 10-20% and the need to induce differentiation as a model system for studying the impact of HBV on
prior to infection, which involves the maintenance of hepatocyte physiology. [86,185,186]
cells in 2% DMSO for 2 weeks before the induction of
differentiation. [198,201] HBV NATURAL HISTORY

The recent discovery of hNTCP as the functional HBV HBV infection can lead to high viral titers in the blood
receptor has important implications for basic HBV of HBV-infected individuals, with levels of HBV virions
research and antiviral development. In particular, reaching as high as 10 particles/mL. [209] Because of
10
identification of hNTCP has opened new avenues for the these high titers of HBV in blood, the main mechanism
establishment of novel cell culture model systems that for the transmission of the virus is through the blood.
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