Fig. 7
Doxycycline (Dox) did not affect signal transduction by transient LPA stimulation, but attenuated the long term effect of LPA on cell growth. a. 4T1 cells were serum starved with DMEM/0.1% BSA and stimulated with 1 μM LPA for 5, 10 and 20 min. Dox at 10 μg/ml was included in serum starvation and stimulation. LPA-induced stimulations of Akt and ERK phosphorylations were not affected by Dox. b. MDA-MB-231 cells were serum starved with DMEM/0.1% BSA and stimulated with 10 μM LPA. Dox at 10 μg/ml was included in serum starvation. LPA-induced Ca2+-mobilization was not affected by Dox. c. A three-dimensional culture system was established by layering 400 μl of 4T1 cell suspension (6000 cells in DMEM with 10% FBSC and 2% Matrigel™) over 150 μl Matrigel™ in an 8-well chamber with daily replacement with fresh medium containing LPA or drugs. Dox decreased the stimulation of 4T1 cell colony formation by 5 μM LPA after incubation for 9 days. Scale bar = 1000 μm. *p < 0.05 relative to cells without Dox and LPA treatment, # p < 0.05 relative to cells treated with LPA but not Dox. Results are means ± SEM from three independent experiments. Results were analyzed by ANOVA with an SNK posthoc test