Fig. 9. (A) Neuronal differentiation of NSC (10 days) in serum-free differentiation media supplemented with B27 that contains insulin (5 μg/ml). (B and C) Neuronal pathway of differentiation of 1205Lu and OM431 melanoma cells (10 days) in the differentiatione media with the B27 supplement.
Confocal images were developed after immunostaining using rabbit polyclonal Ab to SOX2, a pluriopotency marker (green) and monoclonal antibody to Nestin, an early neuroprogenitor marker; polyclonal Ab to Doublecortin, a neuronal marker (green) and monoclonal Ab to Nestin (red); polyclonal Ab to phospho-ERK (green) and PI, polyclonal Ab to STAT3 (green) and PI (red). Bar = 50 μm. Images from independent experiments: (I), (Ia) and (Ib); (IIIa), (IIIb) and (IIIc). (D) 1205Lu cells, which were stably transfected with dominant-negative Rac1N17, after 10 days in differentiation media. (E and F) Relative length of Nestin-containing neurites before (day 0) and 10 days after differentiation was determined using 5 random areas of images. Error bars represent mean ± SD (p < 0.05, Student’s t test). Stars indicate significant difference in relative neurite length for normal 1205Lu cells at days 0 and 10; double star indicates significant difference between control and Rac1N17-transfected 1205Lu cells at day 10.