Figure 3.

Mice requiring early euthanasia display liver pathology and Fe-S cluster deficiency

(A) Vector genome copy (VGC) of AAV9-CAG-FXN in liver samples, as measured by ddPCR in WT untreated (n = 15), MCK-Fxn saline (n = 12), and MCK-Fxn treated at 1 × 10¹³ vg/kg (n = 13), 3 × 10¹³ vg/kg (n = 10), or 1 × 10¹⁴ vg/kg (n = 12). (B) mRNA expression of endogenous mouse Fxn and human FXN transgene in liver extracts, as measured by ddPCR in WT untreated (n = 15), MCK-Fxn saline (n = 12), and MCK-Fxn treated at 1 × 10¹³ vg/kg (n = 13), 3 × 10¹³ vg/kg (n = 9), or 1 × 10¹⁴ vg/kg (n = 12). (C) Detection of Fxn transgene mRNA expression by in situ hybridization (ISH, upper panels) and human FXN protein by immunohistochemistry (IHC, lower panels) in liver sections. Scale bars, 200 μm (ISH), 300 μm (IHC). (D) Measurements of liver-specific glutamate dehydrogenase (GLDH) from sera taken at time of necropsy: 57 days post-injection (dpi) for WT untreated (n = 15) and MCK-Fxn treated at 1 × 10¹³ vg/kg (n = 12), 14 dpi for MCK-Fxn saline group (n = 10), 26–57 dpi for MCK-Fxn treated at 3 × 10¹³ vg/kg (n = 11) and 20–26 dpi for MCK-Fxn treated at 1 × 10¹⁴ vg/kg (n = 13). All data are mean ± SEM. ∗∗∗∗p < 0.0001. (E) Western blot analysis of mitochondrial and Fe-S client proteins from three mouse liver extracts from each group. Vinculin (VINC) was used as a loading control.