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[Preprint]. 2023 Feb 15:rs.3.rs-2551245. [Version 1] doi: 10.21203/rs.3.rs-2551245/v1

Figure 8. Impact of HBMEC-derived EVs on synaptic protein expression in NPCs.

Figure 8

Non-transfected HBMEC were treated with HIV and/or Aβ and EVs were isolated as in Figure 3. Then, human NPCs were exposed to HBMEC-derived EVs for 24 h, with selected cultures additionally treated with 2 μM PAI039 (P) as in Figure 7. (A) synaptophysin (green; arrow heads) and PSD95 (red; arrows) immunoreactivity as imaged by confocal microscopy. DAPI staining (blue) visualizes the NPC nuclei. The combined z-stack images with maximum intensity projection are representative from three experiments. Scale bar: 10 μm. Individual panels are shown in Supplementary Figure 2. (B) Total intensity of synaptophysin (left graph) and PSD95 (right graph) immunoreactivity as quantified from the confocal images. Values are mean ± SEM, n=7. One-, two- and three-way ANOVA with Šídák’s multiple comparisons tests. (C) Example of a brightfield confocal image with random identical rectangular areas on NPC projections. (D) Synaptophysin (green) and PSD95 (red)immunoreactivity on representative confocal images of NPC projections (1 control-EVs, 2 control-EVs+PAI039, 3 Aβ-EVs, 4 Aβ-EVs+PAI039, 5 HIV-EVs, 6 HIV-EVs+PAI039, 7 HIV+Aβ-EVs, 8 HIV+Aβ-EVs+PAI039). Scale bar: 2 μm. (E) Intensity of synaptophysin and PSD95 immunoreactivity as quantified from random projection areas. Values are mean ± SEM, n=160–180. One-, two- and three-way ANOVA with Šídák’s and Tukey’s multiple comparisons tests. *Statistically significant at p<0.05, ****p<0.0001.