Regulation Of HBV C Gene On NLRP3 Inflammasome Signaling Pathwayand Its Mechanism

Background and aims: Hepatitis B virus(HBV)has four open reading frames(ORFs)of which ORF C is consists of the pre Core and Core genes encodes the Hepatitis B core antigen(HBcAg)and Hepatitis B e antigen(HBeAg).Studies have shown that HBeAg significantly inhibits the NLRP3 inflammasome activation and IL-1? production.However,the role of HBcAg and ORF C(in this paper,ORF C = HBcAg + HBeAg)were remain unclear.Our study aims to assess whether HBcAg and ORF C proteins can affect the NLRP3 inflammasome pathway.Methods: HepG2 cells were stimulated with 0,50,100,500,1000,2000 ng/ml bacterial lipopolysaccharide(LPS),respectively,aims to find the optimal concentration of LPS-stimulated HepG2 cells.And then,HepG2 cells were stimulated with the optimal concentration of LPS for 0,1,6,12,18,24,or 30 hours,in order to find the optimal action time point for LPS to stimulate HepG2 cells.Vectors carrying the ORF C gene or the core gene were separately constructed and transfected into HepG2 cells.And then the cells were stimulated with an optimal concentration of LPS for theoptimal action time point.Western blot,quantitative reverse transcription polymerase chain reaction(PCR),enzyme-linked immunosorbent assay(ELISA)and immunofluorescence were used to detect the activity of NLRP3 inflammatory corpuscles and the expression levels of interleukin-1? and interleukin-18.Results: The expression of NLRP3 and interleukin-1? peaked when HepG2 cells were stimulated with 1 ug/ml LPS for 18–24 h.HBcAg,but not ORF C,promoted LPS-induced NLRP3 inflammasome activation and IL-1?,IL-18 production by phosphorylate nuclear transcription factor ?B.Conclusion: HBcAg can promote LPS-induced activation of NLRP3 inflammasome and expression of IL-1? and IL-18.These findings provide a novel mechanism on how the Hepatitis B Virus causes liver inflammation and may provide insights into the search for new therapeutic strategies.