Figure 1.
miR-342-5p Was a Downstream miRNA of the Notch Pathway in NSCs
(A) Screening of Notch downstream miRNAs using microarray hybridization. Primary neurospheres were cultured with cells derived from the ganglion eminences (GE) of NesCre-Rbp-jf/f (cKO) and NesCre-Rbp-j+/f embryos (Ctrl) (E11.5). Differentially expressed miRNAs were compared by using microarray hybridization and clustered. RNA samples were derived from three different pairs of littermates.
(B) The expression of Evl mRNA, miR-342-3p, miR-342-5p, Hes1 mRNA, and Hes5 mRNA in Rbp-j cKO and control neurospheres was determined by qRT-PCR. Neurospheres were derived from four different pairs of littermates.
(C) Primary neurospheres were cultured by using cells derived from GE of wild-type embryos (E15.5). Cells were treated with 75 μM GSI for 12 hr, and the expression of miR-342-3p and miR-342-5p was examined by qRT-PCR. DMSO was used as a control. RNA samples were extracted from four pairs of GSI- and DMSO-treated neurospheres.
(D) The expression of miR-342-5p, miR-342-3p, Hes1 mRNA, and Hes5 mRNA in the GE and striatum during neural development in mice was determined by qRT-PCR. The brain tissues at the specific time point came from four mice).
(E) The expression of miR-342-5p in the telencephalon of E14.5 embryo was determined by in situ hybridization using a locked nucleic acid probe. The brain sections were derived from three different wild-type embryos. (a–d) Coronal sections of E14.5 telencephalon. (c′ and c″) The cortex and GE regions of the telencephalon section in (c) are shown in magnification, respectively. VZ, ventricular zone; SVZ, subventricular zone; IZ, intermediate zone; CP, cortical plate.
(F) The expression of miR-342-5p and miR-342-3p in neurospheres of the first-generation (1°) and third-generation (3°) was determined by qRT-PCR. RNA samples were extracted from four pairs of passaged neurospheres.
Bars, means ± SD. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ns, not significant.