Background and Aim:  The aim of the present study was to examine the role of mitogen‐activated protein (MAP) kinase pathway on gastric surface epithelium using an established cell culture model in which differentiation is promoted in GSM06 cells by air–liquid interface.

Methods:  A double‐dish culture system of mouse gastric surface mucous cell line GSM06 in Ham's F12 medium supplemented with 10% fetal calf serum and 50 μg/mL gentamicin at 37°C in a humidified atmosphere of 5% CO₂ in air was used for an air–liquid interface. Culture cells were examined on histology, cell proliferation was evaluated by bromodeoxy‐uridine (BrdU) uptake, and western blot analysis of extracellular signal‐regulated kinase (ERK)1/2 and phosphate ERK1/2. On day 3, U0126, an inhibitor of MAP kinase kinase (MEK), was added to medium of incubated cells.

Results:  GSM06 cells were differentiated with an air–liquid interface for 3 weeks. Compared to immersion control culture, phosphorylated ERK 1/2 expression increased significantly. This increase was completely suppressed with U0126, and tall columnar cells developed by air–liquid interface in GSM06 were not observed in U0126‐treated cells. Increase in BrdU uptake with air–liquid interface was suppressed by U0126.

Conclusion:  These results suggested that MAP kinase signaling, activated by air–liquid interface, was, at least in part, related to cell differentiation in GSM06 cells induced by air–liquid interface.