Figure 2.

AAV/GAN Vectors Restore Normal IF Morphology in GAN Patient Fibroblasts

Cultures of fibroblasts from three patients (GAN-T, GAN-S, and GAN-M) were infected with either vehicle, AAV2/JeT-GFP, AAV2/JeT-FLAG-GAN, or AAV2/CMV-GAN for 72 hr. Cells were then stained for vimentin (red), GFP (green), and DAPI to visualize the nuclei (blue). (A–C) Representative images of GAN-S fibroblasts. Arrows indicate inclusions. (A) Enlarged view of normal (left) and GAN-specific (right) distribution of vimentin in vehicle-treated GAN fibroblasts to illustrate how inclusion-negative and inclusion-positive cells were scored. (B) GFP staining of GFP-treated patient cells shows the relative transduction efficiency of AAV2 in both normal and inclusion-bearing cells. (C) GAN fibroblasts treated with AAV2/FLAG-JeT-GAN show a reduction in the number of cells containing vimentin inclusions. (D) Quantification of the percent of cells containing inclusions for each patient culture. Values are means ± SEM for each group. One-way ANOVA with Bonferonni’s post-hoc analysis of each treatment condition compared to control cells (vehicle and GFP-treated) (**p < 0.01). The number of trials performed per condition are indicated at the base of each bar.