Figure 4.

Simultaneous Liver and Muscle Expression of hSGCA-SIINFEKL Controls the Anti-transgene Immune Response

(A) Five-week-old C57BL/6J mice intramuscularly injected (TA) with 1 × 10¹⁰ vg/mouse of AAV1-SPc5.12-hSGCA-SIINFEKL vector and intravenously injected with 1 × 10¹¹ vg/mouse of AAV9-hAAT-hSGCA-SIINFEKL (Muscle-Liver group). Two groups of mice received the two vectors separately either by intramuscular (Muscle group) or intravenous (Liver group) injection. PBS-injected mice were used as controls (Control group). One month after treatment, mice were sacrificed, and tissues were collected. (B) IFN-γ-ELISPOT performed on splenocytes stimulated with SIINFEKL peptide. (C) Anti-hSGCA IgG titers measured by ELISA using recombinant hSGCA protein. (D) Vector genome copy number (VGCN) per diploid genome measured in TA. (E and F) CD8 (E) and IFN-γ (F) mRNA measured in TA. (G) Anti-hSGCA (green), CD8 (red), and DAPI (blue) immunostaining performed in TA muscle (scale bar: 50 μm). White arrows indicate CD8-positive cells. (H) Vector genome copy number (VGCN) per diploid genome measured in liver. (I and J) CD8 (I) and IFN-γ (J) mRNA measured in liver. Data were expressed as mean ± SD. Statistical analyses were performed by ANOVA in all panels except for (B) and (C), where a Kruskal-Wallis test was used (*p < 0.05, as indicated; n = 3 per group).