Different accumulation of activated extracellular signal-regulated kinases (ERK 1/2) and role in cell-cycle alterations by epidermal growth factor, hydrogen peroxide, or asbestos in pulmonary epithelial cells

The extracellular signal-regulated kinase (ERK) pathway is induced by cytokines and oxidative stress. In this study we examined the patterns of localization of phosphorylated ERK proteins in relationship to subsequent phenotypic: responses by the mitogenic agent epidermal growth factor (EGF) (5 ng/ mi); hydrogen peroxide (H2O2) (100 to 300 muM), an inducer of apoptosis; and crocidolite asbestos (5 mug/cm(2) dish) in a nontransformed murine alveolar type II epithelial cell line (C10), Laser scanning cytometry and flow cytometry were used to determine: (1) whether expression of phosphorylated ERKs was cell cycle-related; and (2) whether cell-cycle alterations by agents could be modified after addition of the mitogen-activated protein kinase/ERK kinase (MEK) 1 inhibitor PD98059, In contrast to other stimuli which induced transient increases in phosphorylated ERKs, asbestos caused fiber-associated localization of phosphorylated ERKs that were elevated from 1 to 24 h (P less than or equal to 0.05), and striking apoptosis followed by increased numbers of cells in the S phase at 72 h. In both control and asbestos-exposed cells, the percentage of phosphorylated ERK-positive cells was greatest in cells in the C-2/M and 5 phases of the cell cycle. All stimuli caused increased proportions of cells in C-2/M at 24 h that were inhibited by PD98059 (30 muM). Increases in G(2)/M cells by H2O2 and asbestos also were decreased at 48 h by the MEK-1 inhibitor. In addition, PD98059 abrogated elevations in S-phase cells by EGF and H2O2 at 24 h and by asbestos at 72 h. Our results suggest that ERKs mediate cell-cycle alterations during the development of epithelial cell apoptosis or proliferation by diverse ERK stimuli.