Activation of ERK1/2 by protein kinase C-α in response to hydrogen peroxide-induced cell death in human gingival fibroblasts

Hydrogen peroxide (H2O2) increases protein tyrosine phosphorylation of numerous proteins in human gingival fibroblasts (HGFs). Two main proteins, with an apparent molecular weight of 44 and 42 kDa, were phosphorylated after hydrogen peroxide stimulation of the human gingival fibroblasts. Further analysis identified these two proteins as ERK1/2. Maximum phosphorylation was detected at 10 min post-H2O2 treatment. Pretreatment with an MEK inhibitor, PD98059, inhibited H2O2-stimulated ERK1/2 phosphorylation in a dose-dependent manner. Treatment with H2O2 also induced phosphorylation of protein kinase C-α (PKCα). Staurosporine, a PKC inhibitor, blocked ERK1/2 phosphorylation induced by H2O2. In addition, H2O2-induced cell death was prevented by PD98059, SB203580, and calphostin C, which are MEK, p38 and PKC inhibitors, respectively. These results suggest that H2O2 leads to the phosphorylation and activation of ERK1/2 in a PKC-dependent manner. These findings demonstrate that the MAPK signaling pathway plays an active role in mediating the H2O2-induced decrease in HGF cell viability and ATP depletion.