Figure 4. NF-α1 and E300Q-NF-α1 (non-enzymatic mutant of NF-α1) promotes the differentiation of neural stem/progenitors into astrocytes.

Neurospheres dissociated into single cells were grown for 5 days in the presence of 1% FBS (differentiation media) and treated with or without NF-α1 or E300Q-NF-α1 on day 0. At the end of day 5 single immunocytochemistry (ICC) staining for astrocytes (anti-GFAP), neurons (anti-β-III tubulin), and oligodendrocytes (CNPase), as well double immunostaining for astrocyte (anti-GFAP) and nestin (anti-Nestin) was carried out. (A) Representative ICC pictures (20X), of control (left) & NF-α1 treated neural stem/progenitor cells (right). Scale bar = 100 µm. (B) & (C) Bar graph shows the percentage of each cell phenotype population. Either NF-α1 or E300Q-NF-α1 treatment on neural stem/progenitor cells significantly increased the number of astrocytes (***p<0.0001) (open bars), without significantly altering the percentage of the neuron (shaded bars) and oligodendrocyte (solid bars) populations. (D) Representative ICC pictures of control (untreated) (left) & NF-α1 treated cells double immunostained for Nestin and GFAP (right). (E) Bar graph shows the percentage of differentiated phenotype populations. NF-α1 treatment on neural stem/progenitors significantly increases differentiation into GFAP⁺ alone, concomitant with a reduction in the Nestin⁺/GFAP⁺ cells population (***p<0.0001) without significantly altering the percentage of Nestin⁺ population. N=3, the values represent the mean ± SEM, one way ANOVA (Tukey’s multiple comparison test).