Fig 2.

Hes1 influence on MEL cell differentiation. MEL cells were stably transfected with vectors expressing shRNA directed against Hes1 mRNA (shHes1 [sh327 or sh329]) or with a nonspecific scrambled shRNA (ShScr). (A) Western blot analysis of HES1 and GAPDH (control) expression in whole-cell extracts of uninduced cells. The antibodies used for the immunodetections as well as specific bands and their corresponding molecular masses are presented. (B) Relative expression of Hes1 and Hprt (negative control) as a function of DMSO induction measured by qRT-PCR. Standard deviations are indicated. Relative expression was calculated according to Pfaffl (43) using Actb as the internal control. −, uninduced cells; +, cells induced with DMSO for 3 days. (C and D) Percentage of diaminofluorene (DAF)-positive cells (C) or benzidine-positive cells (D), with standard deviations, as a function of time in DMSO. Asterisks (*, P < 0.01; **, P < 0.005) indicate significant differences between the value for sh327 or sh329 and that for shScr according to Student's t test. (E and F) Relative expression of Hba-a1, Alas2, Epor, and Trfr2 as a function of DMSO induction as described for panel B. Untr, untreated cells. Asterisks (*, P < 0.05; **, P < 0.001) indicate significant differences between the values for sh329 and shScr according to Student's t test.