Hepatitis B virus (HBV) is a blood-borne pathogen responsible for chronic, cirrhosis, and liver . The mechanism of HBV entry into hepatocytes remains to be investigated. Recently, sodium taurocholate cotransporting polypeptide (NTCP) was discovered as a major HBV receptor based on an in vitro system using NTCP-reconstituted HepG2 cells. However, this system relies on the compound polyethylene glycol (4% PEG), which is not physiologically relevant to human . High concentration of heparin has been commonly used as an inhibitor control for in vitro in the field. Surprisingly, we found that heparin at physiological concentration can enhance HBV in a PreS1-peptide sensitive, NTCP-dependent manner in both HepaRG and HepG2-NTCP-AS cells. O-sulfation of heparin is more important for the enhancement than N-sulfation. This system based on the HepG2-NTCP-AS cells can support in vitro with HBV genotypes B and C, as well as using serum samples from HBeAg positive and negative carriers. In summary, our study provides a PEG-free system closely resembling human natural . In addition, it points to a future research direction for heparin and heparin-binding host factor(s) in the , which are potentially involved in viral entry. To our knowledge, this is the first soluble and host factor which can enhance HBV in vitro .
HBV is an enveloped and partially double-stranded DNA virus which established 1. These have a high risk to develop liver cirrhosis and hepatocellular 2. Studies on the HBV life cycle have been hampered by the lack of an efficient and user-friendly in vitro system. Primary human hepatocytes (PHHs) and HepaRG cells had served as valuable tools for studying the early event of viral entry, albeit the viral receptor remained elusive3,4,5. One major concern of primary human hepatocytes is its expensive cost from the commercial source. In addition, it is not a robust system because the qualities of hepatocytes tend to vary from lot to lot. In contrast, the HepaRG cell system is relatively more reliable and consistent. However, the efficiency is not high, and it involves tedious work to grow and maintain well-differentiated HepaRG cells5. Recently, sodium taurocholate cotransporting polypeptide (NTCP) was identified as a functional receptor for HBV and HDV6. This NTCP-reconstituted HepG2 system has emerged as a powerful resource in the field of HBV research7,8,9.in around 240 million carriers worldwide. Current treatment for HBV cannot effectively eradicate the from hepatitis B patients
One common way to achieve a higher efficiency in HBV 4. Similarly, PEG was able to promote HBV in HepaRG and NTCP-reconstituted HepG2 hepatocytes5,6. On the other hand, the use of PEG is not without any reservations. For example, PEG is known to induce membrane fusion, such as its use in the generation of hybridoma10,11. Therefore, PEG might provoke non-specific fusion between the virions and the host cell membrane. Most importantly, PEG is a non-biological chemical not found in human body. It remains a concern whether the current PEG-containing protocol could faithfully mimic the viral entry event in vivo.is to include 4–5% of polyethylene glycol (PEG) during the of viral . For example, in the presence of PEG, the efficiency of HBV on PHH was elevated up to 20 times, mainly due to the enhanced adsorption between virus and hepatocytes
HBV 12. Soluble heparins, which share a similar structure with HSPGs, could therefore inhibit HBV entry by interrupting the interaction between HBV and HSPGs on target cells12. As a highly sulfated glucosaminoglycan, heparin is mainly secreted by basophils and mast cells13. The normal physiological concentration of heparin in human plasma ranges from 1 to 5 μg/ml14,15,16. The physiological roles of heparin and HSPGs are highly diverse, including anticoagulation, signaling, development, and anti-metastatic17,18. To date, more than 400 heparin-binding proteins have been reported in literature19.is supposed to be initiated by HBV binding to the heparan sulfate proteoglycans (HSPGs) on the hepatocyte surface