Fast Differentiation of HepaRG Cells Allowing Hepatitis B and Delta Virus Infections

HepaRG cells are liver bipotent progenitors acquiring hepatocytes features when differentiated in the presence of dimethylsulfoxide (DMSO). Differentiated HepaRG (dHepaRG) are considered the best surrogate model to primary human hepatocytes (PHH) and are susceptible to several hepatotropic viruses, including Hepatitis B Virus (HBV) and Hepatitis Delta Virus (HDV) infection. Despite these advantages, HepaRG cells are not widely used for the study of these two viruses because of their long differentiation process and their rather low and variable infection rates. Here, we tested the use of a cocktail of five chemicals (5C) combined or not with DMSO to accelerate the cells' differentiation process. We found that NTCP-mediated HDV entry and replication are similar in HepaRG cells cultivated for only 1 week with 5C and DMSO or differentiated with the regular 4-week protocol. However, even though the NTCP-mediated HBV entry process seemed similar, cccDNA and subsequent HBV replication markers were lower in HepaRG cells cultivated for 1 week with 5C and DMSO compared to the regular differentiation protocol. In conclusion, we set up a new procedure allowing fast differentiation and efficient HDV-infection of HepaRG cells and identified differential culture conditions that may allow to decipher the mechanism behind the establishment of the HBV minichromosome.