Fig. 4

The cooperation between PSMD10 and GRP78 facilitates ER stress-mediated apoptosis induced by homocysteine. A The interaction between PSMD10 and GRP78 was confirmed by co-immunoprecipitation (Co-IP) in hepatocytes treated with Hcy. Upper panel: Co-IP using anti-GRP78 antibody; Lower panel: Co-IP using anti-PSMD10 antibody. B Co-immunofluorescent staining of GRP78 (green) and PSMD10 (red) in hepatocytes which was treated with 100 μmol/L Hcy (scale bars = 50 μm, 40×). Nuclei were stained with DAPI (blue). C The expression of p-PERK, PERK, p-eIF2α, eIF2α, IRE1α, p-IRE1α and CHOP in hepatocytes with or without Hcy treatment was detected by western blot, after the cells were transfected with si-NC or si-GRP78. D The expression of apoptosis-related markers was analyzed by western blot in hepatocytes after transfection with si-NC or si-GRP78 under Hcy treatment. E The apoptosis ratio of hepatocytes was measured by flow cytometry, after the cells were transfected with si-NC or si-GRP78 and treated with Hcy. F, G The protein levels of Bax, Bcl-2, cleaved caspase-3 and cleaved caspase-12 were assessed by western blot in the hepatocytes co-transduced with adenoviruses encoding PSMD10 (ad-PSMD10) and si-GRP78. All data are expressed as mean ± SD. *P < 0.05, **P < 0.01 versus si-NC group. ^#P < 0.05, ^##P < 0.01 versus si-NC + Hcy group or ad-PSMD10 + si-NC + Hcy group