FIG. 2.

hMCT8 mRNA and protein in cerebrum of mice injected with AAV9-ShMCT8 and AAV9-LhMCT8. (A) The MCT8 cDNA contained in the AAV9 transcribed in a dose-dependent manner. Note that the ICV injection induced 10–40 times more mRNA than IV injection did using doses one order of magnitude lower than those used for IV injections. Doses were as follows: for IV administration, low = 8 × 10¹⁰, middle = 2 × 10¹¹, and high = 4 × 10¹¹ vp per mouse; for ICV administration, low = 5 × 10⁹ and high = 3 × 10¹⁰ vp/mouse. Results are given as mean ± standard error of the mean (SEM). Number (#) of animals per group is indicated. (B) Western blot analysis after SDS-PAGE of cerebrum homogenates developed with an antibody against hMCT8. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is used as a loading control. In agreement with the mRNA data, vectors injected ICV induced more protein than those injected IV did (lanes 5 and 6 compared with lanes 3 and 4). The expected molecular weight of hMCT8 is 60 kDa. ShMCT8 produced a monomer as well as a dimer (lanes 3, 5, and 6). In contrast, LhMCH8 produced predominantly a monomer (lanes 7 and 8). (C) Quantitative analysis of the MCT8 in the bands shown in (B), corrected for the GAPDH loading control. Note that the amount of MCT8 in the brain of the Wt mouse given EV was not calculated because of the unknown level of cross-reactivity.