{'fraction_non_alphanumeric': 0.04154929577464789, 'fraction_numerical': 0.022183098591549297, 'mean_word_length': 5.136069114470843, 'pattern_counts': {'":': 0, '<': 1, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
P64 Hepatitis B virus upregulates hepatocyte expression of PD-L1 to evade hepatoxic adaptive immune responses

Introduction Hepatitis B virus employs a variety of strategies aimed at overwhelming, evading or neutralising the host immune response to infection resulting in chronicity. We have previously shown that the Programmed Cell Death Pathway (PD-1/PD-L1) is an inhibitory T-cell pathway implicated in the homeostasis of immune responses and the balance between cytolytic and non-cytolytic CD8+ T-cell effector functions. Aim The aim of this study was to investigate the impact of hepatitis B virus (HBV) infection on hepatocytic PD-L1 expression. Method A human hepatoma cell line that constitutively expresses HBV-DNA (HepG2215), its parent cell line (HepG2) were cultured. A human hepatoma cell line (Huh7) was transfected with a plasmid containing an HBV head-to-tail dimer using Fugene 6 reagent. We also cultured a further HepG2 cell line (AD38) that produces full infectious virus under the control of a tetracycline (Tet)-responsive promoter. HBV-DNA and PD-L1 were quantitated longitudinally. Intracellular and secreted HBV-DNA was quantified with qRT-PCR. PD-1/PD-L1 expression was assessed by FACS and qRT-PCR. Co-cultures between virus-specific CD8+ T-cell lines and hepatocytes producing HBV were also established and analysis of T cell functions performed. Results The hepatoma cell lines which constitutively produce HBV virions (HepG2215) had significantly higher basal levels of PD-L1 expression compared with their parent cell line (HepG2) (p=0.01). A significant increase in intracellular and secreted HBV-DNA levels confirmed successful transfection of Hepatitis B virus. Following transfection there was a significant increase in PD-L1 levels (p=0.01) on infected hepatocytes, which was not observed following transfection with an empty vector. A significant correlation was observed between PD-L1 expression and both intracellular HBV-DNA (r=0.98, p=0.01) and secreted HBV-DNA (r=0.908, p=0.046) following transfection. Following activation of HBV-DNA expression in the AD38 cell line (-Tet), PD-L1 expression increased. Moreover, subsequent fluctuations in HBV-DNA in the absence/presence of Tet was temporally associated with the expression of PD-L1 (r=0.83, p<0.001). Hyperexpression of PD-L1 on hepatocytes was associated with a predominance of non-cytolytic T cell functions. Conclusion These results demonstrate that HBV-DNA drives PD-L1 expression on infected hepatocytes. As we have previously demonstrated, upregulation of PD-L1 impairs adaptive immune responses to HBV infection, and this novel function of HBV may reflect an important strategy by which hepatitis B virus extends the life-span of target hepatocytes and escapes an effective immune response contributing to the development of chronicity.