Hepatitis B virus particles activate toll-like receptor 2 signaling initial upon infection of primary human hepatocytes

Background & aims:

To date conflicting data exist, whether hepatitis B virus (HBV) has the ability to induce innate immune responses. Here, we investigated cellular changes after the first-contact between primary human hepatocytes (PHH) and HBV in vitro.

Methods:

PHH were isolated after perfusion and digestion of liver tissue obtained after tumor resections. PHH were treated with cell culture-derived HBV particles (HepG2.117) or mock control preparation and cultured for 1 – 10 days. Infection was monitored by the release of HBsAg and HBeAg, transcription of viral genes, and immunocytochemistry staining. The gene expression profiles in PHH were analyzed by gene chip array and the relevant changes were verified by quantitative RT-PCR and western blot analysis. TLR ligands, HBV entry inhibitor and neutralizing antibodies were used to characterize the initial immune response in PHH exposed to HBV.

Results:

First of all, PHH could be efficiently infected with cell culture-derived HBV particles, indicated by the secretion of viral antigens and expression of viral mRNA transcripts. Indeed, the exposure of PHH to HBV particles resulted in a gene expression profile, similar to that induced by TLR2 ligand Pam3Cys, but different from those induced by the viral sensors TLR3 or TLR7 – 9, and led to the induction of inflammatory and chemoattractant cytokines but not interferons. Treatment with both, HBV particles and Pam3Cys led to phosphorylation of ERK1, JNK and p38 mitogen-activated protein kinases and NFκB. Finally, HBV-induced gene expression could be neutralized by TLR2-specific antibodies. The blockade of HBV-mediated TLR2 activation enhanced viral transcription but did not affect HBsAg or HBeAg secretion. Of note, pre-treatment with an HBV entry inhibitor attenuated the TLR2-mediated response to HBV, suggesting an entry-related mechanism.

Conclusions:

Primary human hepatocytes are able to sense HBV particles via TLR2 leading to an activation of anti-HBV immune responses. These findings challenge the previously described stealth properties of HBV.