Figure 2.

Delivery of ApoB¹¹:ASO α-syn reduces the accumulation of α-syn in human neuronal cells in vitro. (A) Human neuroblastoma cells, SH-SY5Y, were plated in 12-well dishes, differentiated and infected with LV-α-syn or LV-Ctrl for 72 hours in differentiation media. Cells were then treated with 100pmol α-syn ASO (2’OMe) or ASO scrambled RNA conjugated to ApoB¹¹ peptide. (B) Representative immunohistochemistry images of coverslips stained with an antibody against α-syn which recognizes total human α-syn (mouse anti- α-syn), counterstained with DAPI. Secondary blotting to detect α-syn was done using anti-mouse AF-488. Scale bars, 100μm in low-power images and 20μm in high-power images (C) Image analysis of SH-SY5Y α-syn stained cells represented as fluorescence intensity, corrected for background intensity (ImageJ). N=3 (3 different wells, one representative image per well). (D) Representative immunoblots from SH-SY5Y cells treated as described above and probed for α-syn and Actin. Secondary staining with anti-mouse-AF488 (green) for α-syn and anti-rabbit-AF594 (red) for actin. Densitometric analysis of α-syn immunoreactive band from immunoblot at 20kDa as a ratio to Actin (42 kDa). Blots for the LV-Asyn and the LV-Ctrl group are shown separately. N=2 (2 of the wells were used for each group). One-way ANOVA with post hoc Dunnett’s multiple comparisons test. ** p < 0.005, ****p<0.0001.