Role of ROS-activated ERK1/2 pathway in chemical hypoxia-induced injury in PC12 cells

Aim To explore the role of extracellular signal-regulated kinase 1/2(ERK1/2) in the chemical hypoxia-induced injury in PC12 cells.Methods PC12 cells were treated with cobalt chloride(CoCl2),a well-known chemical hypoxia mimetic agent,to establish a chemical hypoxia-induced cellular injury model.Cell viability was detected by cell counter kit(CCK-8).Apoptotic cells were analysed by Flow cytometry(FCM).Mitochondrial membrane potential(MMP) was examined by rhodamine 123(RH123) staining and photoflutograph.The expression levels of caspase-3,ERK1/2 and p38 mitogen-activated protein kinase(MAPK) were tested by Western blot assay.Results Exposure of PC12 cells to 600 μmol·L-1 CoCl2 for 60 min markedly enhanced the expression of phosphorylated(p)ERK1/2.Pretreatment of PC12 cells with 500 μmol·L-1N-acetyl-L-cystein(NAC),a reactive oxygen species(ROS) scavenger,for 1h before exposure to CoCl2 inhibited the upregulation of p-ERK1/2 expression induced by CoCl2 treatment.Pretreatment of PC12 cells with 10 μmol·L-1U0126(an inhibitor of ERK1/2) for 2h prior to exposure to 600 μmol·L-1CoCl2 protected against the CoCl2-induced injuries,leading to decreases in the number of apoptotic cells,expression of cleaved caspase-3 as well as MMP loss.Pretreatment with 10 μmol·L-1U0126 also attenuated the CoCl2-induced upregulation of p-p38MAPK expression.Furthermore,preteatment with 20 μmol·L-1 SB203580,an inhibitor of p38MAPK,blocked the CoCl2-induced upregulation of p-ERK1/2 expression.Conclusions ROS-activated ERK1/2 pathway mediates the CoCl2-induced injury and there is a synergetic interaction between ERK1/2 pathway and p38MAPK in CoCl2 treated PC12 cells.