FIG. 3.
Increased expression of methylmalonyl-CoA mutase in the liver of mice treated with a single intrahepatic injection of AAV8-TBG-mMut (2 × 1011 GC). (A) Liver Mut mRNA expression was determined by quantitative PCR. Treated Mut–/– mice had significantly higher mRNA expression: 13% at 90 days of life (n = 3; p < 0.01) and 46% at 270 days of life (n = 3; *p < 0.01), compared with untreated Mut+/− mice (n = 3). Untreated Mut–/– mice had less than 0.5% of the Mut mRNA expression of Mut+/− mice. (B) The expression of Serpina7 (TBG) was assayed by quantitative PCR in treated Mut–/– mice. The mice at 270 days (n = 3) had, on average, 2.4-fold increased Serpina7 RNA levels compared with younger animals (n = 3). (C) Immunoblotting showed that one of two treated Mut–/– mice studied at 90 days had hepatic Mut protein levels similar to that of Mut+/− mice. No Mut protein was detected in untreated Mut–/– mice. (D) rAAV-mediated transgene expression in the liver. Shown is GFP expression and localization in liver sections of mice killed 60 days after injection of rAAV8-TBG-GFP at 2 × 1011 GC (A–C) compared with control mice (D–F). GFP expression (A and D) and nuclear DNA (B and E) images were merged (C and F), using LSM Image Browser software (Carl Zeiss). The white arrow indicates an area where the nuclear contour is preserved with green signal present. Color images available online at www.liebertonline.com/hum.