Fig. 5

Apoptosis-inducing effect of XAG on HCC cells was abrogated by autophagy mediated by triggering ERS signaling pathway. a Bel 7402 and SMMC 7721 cells were co-cultured with 10 and 20 μM of XAG or vehicle. Western blotting analysis detected the expression of ERS-related proteins, including CHOP, GRP78, ATF-6, p-eIF2α, IRE1α, and cleaved caspase-12. GAPDH was used as control group. **p<0.01. b After cells were pre-treated with or without 2.5 mM of TUDCA (as ERS inhibitor), then co-cultured with 20 μM of XAG for 48 h. The expression levels of LC3B-II, p62/SQSTM1, Beclin 1, Atg5 were analyzed by Western blotting, and GAPDH was used as control group. **p<0.01. (c) After cells were transfected with shRNA targeting CHOP, then were treated with or without 20 μM of XAG for 48 h. Expression of autophagy-associated proteins (LC3B-II, p62/SQSTM1, Beclin-1, and Atg5) was detected by Western blotting, and GAPDH was used as control group. **p<0.01. d After cells were pre-treated with or without 2.5 mM of TUDCA (an ERS inhibitor), then were co-cultured with 20 μM of XAG for 48 h. Cell apoptotic ratio in each treatment group was quantified by flow cytometry. **p<0.01. e After cells were pre-treated with or without 2.5 mM of TUDCA, then were co-cultured with 20 μM of XAG for 48 h. The expression of Bax, Bcl-2, and cleaved caspase-3 was analyzed by Western blotting, and GAPDH was used as control group. **p<0.01. f After cells were transfected with shRNA targeting CHOP, then were treated with or without 20 μM of XAG for 48 h. Cell apoptotic ratio was quantified by flow cytometry. **p<0.01. g After cells were transfected with shRNA targeting CHOP, then were treated with or without 20 μM of XAG for 48 h. The expression levels of Bax, Bcl-2, and cleaved caspase-3 were analyzed by Western blotting, and GAPDH was used as control group. **p<0.01