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Monitoring Activation of the Antiviral Pattern Recognition Receptors RIG-I And PKR By Limited Protease Digestion and Native PAGE
Published on: July 29, 2014
Expression and functionality of Toll- and RIG-like receptors in HepaRG cells
Souphalone Luangsay1, Malika Ait-Goughoulte1, Maud Michelet2
1INSERM U1052, CNRS UMR_5286, Cancer Research Centre of Lyon (CRCL), 69008 Lyon, France; University of Lyon, Université Claude Bernard (UCBL), UMR_S1052, 69008 Lyon, France; Pharma Research & Early Development (pRED), Roche Innovation Center Basel, F. Hoffmann-La Roche AG, 4070 Basel, Switzerland.
Background & Aims:
HepaRG cells are considered as the best surrogate model to primary human hepatocyte (PHH) culture to investigate host-pathogen interactions. Yet their innate immune functions remain unknown. In this study, we explored the expression and functionality of Toll-like (TLR) and retinoic acid-inducible gene-1 (RIG-I)-like receptors (RLR) in these cells.
Methods:
Gene and protein expression levels of TLR-1 to 9 and RLR in HepaRG were mainly compared to PHH, by RT-qPCR, FACS, and Western blotting. Their functionality was assessed, by measuring the induction of toll/rig-like themselves and several target innate gene expressions, as well as the secretion of IL-6, IP-10, and type I interferon (IFN), upon agonist stimulation. Their functionality was also shown by measuring the antiviral activity of some TLR/RLR agonists against hepatitis B virus (HBV) infection.
Results:
The basal gene and protein expression profile of TLR/RLR in HepaRG cells was similar to PHH. Most receptors, except for TLR-7 and 9, were expressed as proteins and functionally active as shown by the induction of some innate genes, as well as by the secretion of IL-6 and IP-10, upon agonist stimulation. The highest levels of IL-6 and IP-10 secretion were obtained by TLR-2 and TLR-3 agonist stimulation respectively. The highest preventive anti-HBV activity was obtained following TLR-2, TLR-4 or RIG-I/MDA-5 stimulations, which correlated with their high capacity to produce both cytokines.
Conclusions:
Our results indicate that HepaRG cells express a similar pattern of functional TLR/RLR as compared to PHH, thus qualifying HepaRG cells as a surrogate model to study pathogen interactions within a hepatocyte innate system.
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