DCX down regulation is time dependent and irreversible. WB representative image (A): lysates (50 μg/lane) of SK-N-SH control and RA treated cells at different time intervals (2, 4, 6, 6 plus 6 of withdrawal and 12 days) were analyzed by immunoblotting using an anti-DCX specific antibody; membranes were reprobed with anti-tubulin as a loading control. Quantification of DCX protein amount by scanning densitometry (B): data are means ± standard error, N = 8, and are expressed as a percentage of the control (100%). After 2 or 4 days of RA treatment DCX levels are significantly decreased, but to a lesser extent than after a 6 days exposure to RA. Lengthening SK-N-SH exposure to RA up to 12 days does not modify the magnitude of the DCX reduction observed after 6 days of treatment. Moreover, in cells treated with RA for 6 days and then grown for 6 additional days in medium without RA (6 days+6), DCX protein amount remains at low levels; single star = p < 0.05; double star = p < 0.01; C, control, RA, samples treated with RA 10 μM.