The PERK-eIF2α signaling pathway is involved in TCDD-induced ER stress in PC12 cells

Studies have shown that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces apoptotic cell death in neuronal cells. However, whether this is the result of endoplasmic reticulum (ER) stress-mediated apoptosis remains unknown. In this study, we determined whether ER stress plays a role in the TCDD-induced apoptosis of pheochromocytoma (PC12) cells and primary neurons. PC12 cells were exposed to different TCDD concentrations (1, 10, 100, 200, or 500 nM) for varying lengths of time (1, 3, 6, 12, or 24 h). TCDD concentrations much higher than 10 nM (100, 200, or 500 nM) markedly increased glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP) levels, which are hallmarks of ER stress. We also evaluated the effects of TCDD on ER morphology in PC12 cells and primary neurons that were treated with different TCDD concentrations (1, 10, 50, or 200 nM) for 24 h. Ultrastructural ER alterations were observed with transmission electron microscopy in PC12 cells and primary neurons treated with high concentrations of TCDD. Furthermore, TCDD-induced ER stress significantly promoted the activation of the PKR-like ER kinase (PERK), a sensor for the unfolded protein response (UPR), and its downstream target eukaryotic translation initiation factor 2 cc (eIF2 alpha.); in contrast, TCDD did not appear to affect inositol-requiring enzyme 1 (IRE1) and activating transcription factor 6 (ATF6), two other UPR sensors. Importantly, TCDD significantly inhibited elF2 alpha phosphorylation and triggered apoptosis in PC12 cells after 6-24 h of treatment. Salubrinal, which activates the PERK-elF2 alpha pathway, significantly enhanced eIF2 alpha phosphorylation in PC12 cells and attenuated the TCDD-induced cell death. In contrast, knocking down eIF2 alpha using small interfering RNA markedly enhanced TCDD-induced cell death. Together, these results indicate that the PERK-eIF2 alpha pathway plays an important role in TCDD-induced ER stress and apoptosis in PC12 cells. (C) 2014 Elsevier Inc. All rights reserved.