Figure 4.

High-titer AAV induces immune cell infiltration

(A) Low-magnification Zeiss fluorescence microscope imaging of GFP (green) co-stained with CD45 (red) at 30 dpi of AAV injection. Note that many infiltrated leukocytes were present in high-titer AAV treated mice. Scale bar, 200 μm. (B) Confocal images of CD45 immunostaining in 10⁸ GC, 10⁹ GC, and 10¹⁰ GC groups. Scale bar, 20 μm. (C and D) Quantifications of CD45 fluorescence intensity (C) and CD45⁺ cell density (D) in 10⁸ GC, 10⁹ GC, and 10¹⁰ GC groups. (E) High-magnification confocal image of CD45 (red) co-stained with IBA1 (green) at 30 days after high-titer AAV injection. Arrowhead indicates a typical infiltrated lymphocyte that is lacking Iba1. Scale bar, 20 μm. (F) Pie chart showing that 93.5% of CD45 positive cells were Iba1 negative. (G) Experimental design. (H) Low-magnification confocal images of CD45 signal at different time points. Scale bar, 50 μm. (I) Quantifications of CD45⁺ cell density at different time points. (J) High-magnification confocal images of CD45 (red) co-stained with CD8 (green, upper row), and CD45 (red) co-stained with CD4 (green, bottom row) at different time points. Arrowheads indicate some infiltrated lymphocytes co-labeled with CD8 (top) and CD4 (bottom) T cell antigens. Scale bars, 20 μm. (K and L) Quantifications of CD8⁺ and CD4⁺ cell density at different time points. (M and N) Quantifications of the percentage of CD8 and CD4 positive cells. Values are shown as mean ± SD. n = 5 mice per group. One-way ANOVA analysis with Tukey test. Significance reported as ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.