Preliminary Study Of S Antigen Mutation In The Pathogenesis Of Occult Hepatitis B Infection Through The UPR-ERAD Pathway Of Endoplasmic Reticulum Stress

Background: The mutation of hepatitis B virus surface antigen can lead to the occurrence of occult hepatitis B infection,especially the substitution of amino acids(AA)in the main hydrophilic region(MHR)of hepatitis B virus small surface proteins(SHBs).It can change the antigenicity of surface antigen and its expression level,but its specific mechanism is still unclear.The purpose of this study is to explore the potential effects of SHBs mutations on serum HBsAg levels and its mechanisms.Methods: Huh7 and Hep G2 cells were transfected with hepatitis B SHBs mutations(G145R,Q129 R,T126A,G119 R,D144A)and wild-type SHB plasmids,and electrochemiluminescence was used to detect the level of HBsAg in cell culture supernatants 72 hours after transfection.The expression level of HBsAg,endoplasmic reticulum molecular chaperone protein GRP78 and endoplasmic reticulum stress-related proteins(p-PERK,p-e IF2α,p-IRE1α,XBP1,ATF6)and endoplasmic reticulum-related degradation proteins(EDEMs)expression levels in the cell lysate was detected by western blotting,immunofluorescence was used to observe the intracellular localization of HBsAg.Results: Compared with the wild-type plasmid transfection group,the SHBs mutant plasmid transfection group had lower levels of HBsAg in the cell supernatant and cell lysate.Compared with HBsAg in cell lysate/cell supernatant of the SHBs mutation plasmid transfection group and the wild-type plasmid transfection group,there was a certain intracellular accumulation.Immunofluorescence localization also showed that HBsAg accumulated in the endoplasmic reticulum.And the levels of endoplasmic reticulum molecular chaperone protein(GRP78),endoplasmic reticulum stress-related proteins(p-PERK,p-e IF2α,p-IRE1α,XBP1,ATF6)and endoplasmic reticulum-related protein degradation proteins(EDEMs)was significantly increased.Conclusion: The occult hepatitis B is related to the hepatitis B SHBs mutation,and the mutant protein accumulate in the endoplasmic reticulum and cause ER stress.Endoplasmic reticulum stress can reduce the transcription and translation of surface antigens by activating the endoplasmic reticulum unfolded protein response.At the same time,ER stress can activate endoplasmic reticulum associated protein degradation pathway(ERAD)and reduce the assembly and secretion of HBsAg in ER.