Figure 3.

Ad-RAR-β mediated RAR-β over-expression improves neuronal differentiation of rat MSCs. Cells were divided into 4 groups. Cells in control group were untreated, cells in ATRA/MNM induction group were induced with 1 µmol/l ATRA for 24 h, followed by MNM incubation for another 24 h. Before neuronal induction, cells in Ad-null/ATRA/MNM induction group and Ad-RAR-β/ATRA/MNM induction group were infected by adenovirus Ad-null and Ad-RAR-β for 48 h, respectively. A – Cell morphology, a – Control, b – ATRA/MNM treatment, c – Ad-null/ATRA/MNM treatment, d – Ad-RAR-β/ATRA/MNM treatment. B – Expression of nestin, NSE, MAP-2, and Tau was analyzed with GAPDH normalization (n = 3, ^* p < 0.05, Ad-RAR-β/ATRA/MNM vs. Ad-null/ATRA/MNM). Real-time PCR results were confirmed in at least three batches of independent experiments. C – Rat MSCs were treated with ATRA for 24 h and collected at the indicated time points with MNM culture, then lysed and subjected to SDS-PAGE and western blot using NSE and Tuj1 antibody. Equal loading of the samples was confirmed by β-actin expression. D – Immunofluorescence staining of neural specific markers. Cells were fixed and probed with antibodies against nestin, NSE, and Tuj1 followed by staining with DyLight 488 labeled secondary antibodies and staining of nuclei with DAPI. Scale bar = 200 µm