Fig. 3: Down-regulation of PLCβ1 reduces AraC-induced NT2 cell differentiation.

Sample Western blot (A) and densitometric analysis (B) of PLCβ1, NF200 and βIII-tubulin expression in NT2 cells transfected either with scrambled off-target siRNA or siRNA targeting the PLCB1 transcript (PLCβ1 siRNA) and treated with AraC for 72 h. Equal amounts of total protein (30 μg) were loaded on the same gel and run in parallel. The y-axis of the graph shows the mean optical density (OD) as a percentage of that in NT2 cells (100%). One-way ANOVA followed by post hoc Tukey’s test (*p < 0.05; **p < 0.01; ***p < 0.001). Data are mean ± SEM (n = 4). C–D Immunofluorescence visualizing PLCβ1 (red) and βIII-tubulin (green) combined with Hoechst’s chromatin staining (blue) in NT2 cells after transfection with either scrambled siRNA (C) or PLCβ1 siRNA (D) and treated with AraC for 72 h. Scale bar = 100 µm (applies to C, D). E–N Immunofluorescence studies of primary rat astrocytes under control conditions or treated with si(RNA)PLCβ1a where NF-H refers to the heavy subunit of neurofilament. More images can be found in Fig. S4G. Scale bar = 50 μm.