DCA stimulation of EGFR-MAPK signaling in Caco-2 cells is mediated by PA.

(A) Caco-2 cells grown to a monolayer were pre-serum-starved before incubation with 1μM DCA for various time points to ensure that the total serum starvation time was 3 hours without / with DCA. Whole cell lysates were collected and blotted using antibodies against pMEK, total MEK, pERK, total ERK, pAkt or total Akt, as well as the loading control actin. (B) Caco-2 cells grown to a monolayer were pre-serum-starved for 2 hours and 55 minutes before incubation with various concentrations (1–30μM) DCA for 5 minutes. Whole cell lysates were collected and blotted using antibodies against pMEK, total MEK, pERK, total ERK, pAkt or total Akt, as well as the loading control actin. (C) Caco-2 cells were pre-serum-starved for 2 hours and 30 minutes before incubation with 0.75μM FIPI for 25 minutes and subsequent co-incubation with FIPI and 1μM DCA for 5 minutes. Whole cell lysates were collected for Western blotting against pMEK, total MEK, pERK, total ERK, pAkt or total Akt, as well as the loading control actin. In all signaling experiments, 3 individual experiments were performed separately and quantitation is shown as mean±SEM. Statistical significance was evaluated using one-way ANOVA, with * indicating p<0.05.