Effects of steroid hormones on cell migration and the actin cytoskeleton
in A549 cells. A, effects of steroid hormones on cell migration.
A549 cells were cultured with the indicated steroid hormones for 48 h and
treated with mitomycin C for 2 h preceding the assay. Migration toward FBS in
the presence of the steroids was measured in a Transwell migration chamber
(mean ± S.D., **, p < 0.01 compared with
vehicle; n.s., not significant). B, effect of steroid
hormones on the actin cytoskeleton. A549 cells were treated with 1
μm of each steroid for 48 h and stained with phalloidin
(red in merged image) and anti-vinculin antibody (green in
merged image). Bar, 50 μm. C, effect of DEX on Rho
activity. The Rho activity was determined by RBD-beads pulldown assay as
described under “Experimental Procedures.” The amounts of total
and GTP-RhoA protein were detected by Western blot with anti-RhoA antibody.
The amounts of Rho downstream effectors, ROCK1, ROCK2, mDia1, cofilin 1,
phospho(Ser-3)-cofilin 1, MLC2, phospho(Ser-19)-MLC2, and profilin were also
analyzed by Western blot analysis using their corresponding antibodies.