Supplementary Figure 3: The loss of miR-137 in the nervous system leads to the synaptic overgrowth but has a negligible effect on cell viability.

a, A cartoon schematic indicating the position of brain section for IHC staining. PSD-95 and synaptophysin IHC staining were carried out on 40-μm thick floating sections containing cortical tissue from four pairs of littermate cKO mice, that is, Mir137^loxP/+, Mir137^loxP/+;Nestin-Cre and Mir137^loxP/loxP;Nestin-Cre mice, at the age of postnatal day 18 (P18). b, Relative fluorescence intensity of PSD-95 (F_2,9 = 6.733, P = 0.0163) and synaptophysin (F_2,9 = 7.607, P = 0.0116) elevated upon the loss of miR-137 in the cortex (n = 4 mice per group). Image analyses and quantification were performed using ImageJ software. Data represent means ± s.e.m; One-way ANOVA with Tukey’s post hoc test; n.s., nonsignificant; *, P < 0.05; **, P < 0.01. c, Partial loss of miR-137 resulted in a significant increase in PSD-95 (t = 5.331, P = 0.0060) and Synaptophysin (t = 3.566, P = 0.0235) protein levels (relative to Actin) in the hippocampus (n = 3 mice). Full-length blots are presented in Supplementary Fig. 15. Data are means ± s.e.m; Unpaired Two-Tailed t-test; *, P < 0.05; **, P < 0.01.d, Immunohistochemistry (IHC) staining of Caspase-3 and DAPI in Mir137^loxP/+ and Mir137^loxP/+;Nestin-Cre mice hippocampal tissues. Caspase-3 and DAPI IHC staining was carried out on 40-μm thick floating sections containing hippocampal tissue from four pairs of littermate Mir137^loxP/+ and Mir137^loxP/+;Nestin-Cre mice (n = 10 slices from 4 mice per group) at the age of postnatal day 0 (P0), 18 (P18) and 60 (P60). Relative fluorescence intensity of Caspase-3 and DAPI did not change upon the loss of miR-137 in the hippocampus (t_P0 = 0.1055, P = 0.9171; t_P18 = 0.1561, P = 0.8777; t_P60 = 0.5895, P = 0.5629). Image analyses and quantification were performed using ImageJ software. Data are means ± s.e.m; Unpaired Two-Tailed t test; n.s., nonsignificant.