Fig. 1: Identification and characterization of trametinib (SNR1611), a small molecule that induces neuronal differentiation of adult NSCs from 5XFAD mice.
a Schematic diagram of drug screening using the ATRIVIEW® platform. Changes in the morphology and Tuj1 expression level of adult NSCs from Tg2576 AD model mouse brains upon treatment with FDA-approved drugs were monitored. b–e MEK/ERK signaling was activated in GFAP-positive cells in the hippocampal DG (b, c) and SVZ (d, e) of 5XFAD mice. Immunofluorescence staining images (b, d) and quantification of pERK intensities (c, e) in the hippocampal DG or SVZ of 7.5-month-old 5XFAD mice. Trametinib was administered to 5-month-old 5XFAD mice for 2.5 months. n = 3 mice per group; sagittal sections from each mouse brain. Scale bars, 25 μm. Normalized to the WT-vehicle group. f, g Adult NSCs from 5XFAD mice were cultured to form neurospheres in a medium containing 20 ng/ml EGF and 20 ng/ml bFGF. Gross images were obtained on the 4th day after seeding (f), and neurosphere size was quantified (g). Scale bars, 50 μm. h–l Adult NSCs from 5XFAD mice were seeded on plates and treated with 100 nM trametinib in medium without growth factors for 48 h. Cell lysates were subjected to immunoblot analyses of TUJ1, GFAP, pERKs and ERKs (h) and quantified by band intensities (i). The levels of Cdkn2b and Ngn2 mRNA were analyzed using qRT‒PCR. Each sample was normalized to the expression of Gapdh (j). Cell lysates were subjected to immunoblot analyses of P15 and Neurod1 (k), and band intensities were quantified (l). m The proposed mechanism by which MEK1/2 inhibition induces cell cycle arrest and neuronal differentiation. Data are representative of three independent experiments, and values are expressed as the mean ± SEM. P values were obtained by one-way ANOVA (c, e) and Student’s t-test (i–l). *p < 0.05, **p < 0.005 and ***p < 0.001.
