〔Abstract〕 Objective To investigate the effects and mechanism of knockdown mesencephalic astrocyte-derived neurotrophic factor(MANF) on rifampicin(RFP) induced HepG2 cell injury. Methods A MANF-knockdown stable cell line(MANF Y25) and a control cell line(MANF Y07) were constructed by lentivirus transfection. After HepG2 cells were treated with 100 μmol/L RFP for 24 h, the experiment was divided into MANF Y07+DMSO group, MANF Y07+RFP group, MANF Y25+DMSO group and MANF Y25+RFP group. Western blot and qRT-PCR were used to detect the protein and gene expression levels of MANF and unfolded protein response(UPR)-related genes such as glucose-regulated protein 78(GRP78), protein kinase R-like ER kinase(PERK), eukaryotic translation initiation factor 2α( e IF2α),activating transcription factor 4( ATF4),C/EBP-homologous protein( CHOP),tribbles homolog 3( TRIB3),inositol-requiring enzyme 1( IRE1),spliced X-box binding protein1-S( XBP1-S),non-spliced X-box binding protein 1-U( XBP1-U) and activating transcription factor 6( ATF6); Annexin V-PE/7-AAD double staining was used to detect the apoptosis rate of cells in each group; The changes in the cell proliferation ability were determined by the cell counting kit-8 assay; The relative contents of alanine aminotransferase( ALT),aspartate aminotransferase( AST),alkaline phosphatase( AKP),total bilirubin( TBIL) and indirect bilirubin( IBIL) in the supernatant of cell culture were detected by kits. Results At the protein level,RFP induced the protein expression of MANF,GRP78,p-e IF2α,ATF4 and ATF6; at the gene level,RFP induced the gene expression of MANF and UPR-related genes GRP78,PERK,e IF2α,ATF4,CHOP and TRIB3 in Hep G2 cells. After MANF knockdown,the protein expression levels of GRP78,p-PERK,p-e IF2α,ATF4,ATF6 and the genes expression levels of UPR-related genes mentioned above were further up-regulated( P<0. 05). Moreover,it was also found that after MANF knockdown,the rate of apoptosis increased( P<0. 01),the cell proliferation ability decreased( P<0. 01),and the levels of cell injury markers ALT,AST,AKP,TBIL and IBIL in the supernatant of cell culture increased( P<0. 05). Conclusion RFP activates the UPR,which is further enhanced by MANF knockdown,and cell injury is aggravated,indicating that MANF may play a protective role in RFP-induced Hep G2 cell injury by regulating the UPR.