Figure 6.

miRNA-mediated detargeting acts independent of Treg immunosuppression and reduces tissue clearance by downregulation of OVA-specific CTL response. (A–C) Cells from TA muscle, lymph nodes and spleen were harvested from rAAV1 injected C57BL/6 mice four weeks after treatment and stained for Treg markers (CD4+, CD25+, FOXP3+). The frequencies of Tregs were quantified by flow cytometry and displayed as box and whisker plots (n = 5). (D) C57BL/6 male mice, six weeks old, were i.m. injected with rAAV1.OVA vectors with or without miRBS (1 × 10¹¹ GCs/mouse, n = 5) and sacrificed two weeks after injection to harvest muscles. Tissue sections were stained for H&E (upper panels, original magnification: 20x), CD8 and granzyme B (lower panels: DAPI, blue; red, granzyme B, red; and CD8, green; original magnification: 40x). Scale bars = 200 μm (H&E images), 50 μm (fluorescence images). (E) Quantification of H&E images for nuclear infiltrates in whole tissue section at original magnification of ×20. (F, G) Quantification of CD8 (F) and granzyme B (G) images in four fields at original magnification of 40x. p values were estimated by one-way ANOVA with Tukey’s post hoc test. *p < 0.05, **p < 0.01, ***p < 0.001.