Fig 2: haFGF14-154 inhibits AEP activity via miR-206-3p/BDNF pathway in the brain of APP/PS1 mice. (A-C) miR-206-3p level in the PFC and hippocampal DG region of APP/PS1 mice with intranasal administration of 600 µg/kg haFGF14-154-loaded cationic liposomes (L+H600) or 0.9% saline (vehicle), measured by fluorescence in situ hybridization (FISH), n = 4/5. (D-F) The expression of BDNF and AEP (full-length and cleaved AEP) in the PFC of APP/PS1 mice with intranasal administration of L+H600 or vehicle, n = 6. The plasma levels of miR-206-3p (G) and BDNF (H) in patients with Alzheimer's disease (AD), PD and Schizophrienia detected by qPCR. P values are calculated by age-adjusted ANCOVA model with Tukey's post-hoc test. n = 25/16, 18/18, 15/13 and 12/8 for normal control and patients with AD, PD and Schizophrienia in the experiments of qPCR and ELISA, respectively. Data are mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001. P values are calculated by one-way ANOVA with Tukey's post-hoc test and student's t-test.

Fig 3: Overexpression of the synapsin I fragmentation induces synaptic dysfunction. (a) Schematic diagram of synapsin I domain and its cleavage by AEP. The binding sites of synapsin ? to SVs and F-actin are shown. (b) Fluorescent micrographs showing the distribution of full-length synapsin ? and synapsin ? C83 fragment. Scale bar, 20 µm. (c) Immunostaining showing the colocalization of full-length and synapsin ? C83 fragment with synaptophysin. Scale bar, 20 µm. (d, e) FM4-64 uptake assay. The uptake of FM4-64 is decreased in neurons expressing synapsin ? C83 fragment when compared with neurons expressing full-length synapsin ?. Scale bar, 20 µm. (f) FM4-64 releasing assay. The release of FM4-64 was decreased in neurons expressing synapsin ? C83 fragment when compared with neurons expressing full-length synapsin ?. Scale bar, 20 µm. (g) Dil staining of dendritic spines. The density of dendritic spines in neurons expressing synapsin ? C83 is lower than that in neurons expressing full-length synapsin ?. Scale bar, 10 µm. Bar graphs are the quantification of the dendritic densities (n = 10). (h) Representative traces of mEACs recorded from isolated hippocampal neurons expression EGFP-vector, EGFP-synapsin I, and EGFP-synapsin ? C83. (i) Cumulative plots and mean values (inset) of mEACs amplitude in isolated hippocampal neurons expression vector, synapsin ?, and synapsin ? C83. (j) Cumulative plots of mEACs inter-event interval and mean values (inset) of mEACs frequency in isolated hippocampal neurons in various conditions. Kolmogorov-Smirnov test in cumulative plots (n = 4–6 per group). Data are mean ± SEM; one-way ANOVA; * p < 0.005, ** p < 0.001, *** p < 0.001, **** p < 0.0001

Fig 4: MO decreases production and increases clearance of Aß in APP/PS1 mice. (A) The expression level of BACE1, PS1, AEP, IDE, NEP, and LRP1 from hippocampal lysates of WT mice and APP/PS1 mice with or without MO treatment was evaluated by Western blotting. (B–G) The quantification of the Western blot bands from panel A. ß-actin serves as the loading control. The data are presented as Mean ± SEM, (n = 3). * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 vs. WT or vs. APP/PS1 mice groups.

Fig 5: Netrin-1 reduction in human PD brains, associated with AEP activation and UNC5C cleavage. A) Netrin-1 expression profiling by array of PD substantia nigra from dataset GDS3129 (reference series: GSE8397). Means + s.e.m. are shown (18 Control, 29 PD cases; unpaired t-test). B) Western blot (WB) showing that netrin-1 reduction accompanied by AEP and caspase-3 activation in substantia nigra (SN) samples from subjects with PD. Actin, loading control; MW, molecular weight; ?, full length; ?, cleaved. The data of WB quantitative analysis are presented as means ± s.e.m.(n = 3; 2-tailed t-test). C) AEP activity assay of the human SN samples from subjects with PD and age-matched healthy controls. RFU, relative fluorescence units. Means + s.e.m. are shown (n = 5; 2-tailed t-test). D) Immunohistochemistry of UNC5C N467 fragment and TH in SN sections from people with PD and age-matched healthy controls. Scale bar, 100 µm. Means + s.e.m. are shown (n = 5; 2-tailed t-test). E–G) Immunofluorescence co-localization analysis was used to detect the expression of netrin-1, TH, N467, and total AEP. Images are representative of nine sections from three subjects with PD and age-matched healthy controls. Scale bar, 30 µm. Bar graph shows data as mean ± s.e.m.; 2-tailed t-test.