Alveolar epithelial cells undergo stretching during breathing and mechanical ventilation. Stretch can modify cell viscoelastic properties, which may compromise the balance of forces in the alveolar epithelium. We studied the viscoelasticity of alveolar epithelial cells (A549) subjected to equibiaxial distention with a novel experimental approach. Cells were cultured on flexible substrates and subjected to stepwise deformations of up to 17% with a device built on an inverted microscope. Simultaneously, cell storage (G') and loss (G'') moduli were measured (0.1-100 Hz) with optical magnetic twisting cytometry. G' and G'' increased with strain up to 64 and 30%, respectively, resulting in a decrease in G''/G' (15%). This stretch-induced response was inhibited by disruption of the actin cytoskeleton with latrunculin A. G' increased with frequency following a power law with exponent alpha = 0.197. G'' increased proportionally to G' but exhibited a more marked frequency dependence at high frequencies. Stretching (14%) caused a fall in alpha (13%). At high stretching amplitudes, actual cell strain (14.4%) was lower than the applied substrate strain (17.3%), which could indicate a partial cell detachment. These data suggest that cytoskeletal prestress modulates the elastic and frictional properties of alveolar epithelial cells in a coupled manner, according to soft glassy rheology. Stretch-induced cell stiffening could compromise the balance of forces at the cell-cell and cell-matrix adhesions.