Fig. 1

Sema 3A overexpression induces axonal retraction and increased dendritic arborization in neurons after 4 days in culture. A IF analysis with Ank G (green) and β-III Tubulin (red) markers of neurons transfected with: Sema 3A-GFP (Sema 3A); Sema 3A siRNA (siSema 3A); Npn 1 siRNA (siNpn 1); and Sema 3A-GFP + siNpn 1 (Sema 3A + siNpn 1). As controls, we used neurons transfected with GFP empty vector (GFP) or non-transfected (Ctrl). Scale bar: 10 µm. 40 × objective. Skeleton mask is reported in B. C Axonal length was assessed by Neuron J plug-in. Data are the mean ± SEM of three independent experiments in triplicate (approximately 100 neurons). One-way ANOVA followed by Tukey’s test for multiple comparisons. *P < 0.05; **P < 0.01; ****P < 0.0001. D IF analysis with MAP-2 marker (red). Scale bar: 50 µm. 40 × objective. E Sholl analysis was performed by Neuroanatomy plug-in of ImageJ software on MAP-2-stained neurons. Data represent the number of dendritic branches at a given distance from the soma (1 µm-radius interval) and are the mean ± SEM of three independent experiments in triplicate. Two-way ANOVA followed by Tukey’s test for multiple comparisons. *P < 0.05 vs Ctrl and.^#P < 0.05 vs Sema 3A + siNpn 1. See also Additional File 1: Table S2 for single value analysis. F Dendritic branches were clustered according to Strahler orders’ classification using Neuroanatomy plug-in of ImageJ software. At least 10 neurons were analysed in each slide. Experiments were replicated three times in triplicates. Two-way ANOVA followed by Tukey’s test for multiple comparison. **P < 0.01; ***P < 0.001; ****P < 0.0001