Abstract
Suberoylanilide hydroxamic acid (SAHA) is a histone deacetylase inhibitor that has demonstrated clinical activity against various solid tumors. The aim of the present study was to explore the effects of SAHA on the apoptosis of HepG2 liver cancer cells, as well as the potential mechanisms involved in histone acetylation and endoplasmic reticulum (ER) stress. HepG2 cells were treated with various doses of SAHA (0, 1, 6 and 12 µM), and apoptosis was measured by flow cytometry. The levels of ER stress-associated molecules, including 78 kDa glucose-regulated protein (GRP78), PRKR-like endoplasmic reticulum kinase (PERK), phosphorylated (p)-PERK, activating transcription factor 4 (ATF4) and C/EBP-homologous protein (CHOP), were quantitated by western blot analysis and reverse transcription-quantitative PCR assay. The expression levels of acetylated histone H4 (acH4, acH4 lysine (K)5 and acH4K12) were detected by western blot analysis. The effects of SAHA on the acetylation of H4 in the promoter regions of GRP78, ATF4 and CHOP were evaluated by chromatin immunoprecipitation assays. Following treatment with higher doses of SAHA (6 and 12 µM) for 48 h, the proliferation of HepG2 cells was significantly suppressed. SAHA induced dose-dependent apoptosis and increased both protein and mRNA expression levels of GRP78, ATF4 and CHOP in HepG2 cells. The protein expression of PERK was markedly decreased by treatment with SAHA, whereas the p-PERK expression level was notably increased, which resulted in increased p-PERK/PERK ratio. Furthermore, the acetylation levels of H4 in the promoter regions of GRP78, ATF4 and CHOP were significantly increased in HepG2 cells exposed to 6 µM SAHA for 36 h. Thus, SAHA induces apoptosis in HepG2 cells by activating the ER stress-mediated apoptotic signaling pathway, at least partially by enhancing the acetylation of histone H4 on the promoter regions of ER-stress associated genes, including GRP78, ATF4 and CHOP.