Figure 2.

Expression of CYP27A1 from AAV vectors

(A) Schematic representation of vector genomes. ITR, inverted terminal repeat; pA, polyadenylation signal. (B) The AAV8-EAAT-CYP27A1 and AAV8-C27P-CYP27A1 vectors (abbreviated as EAAT and C27P, respectively) were administered intravenously to 7-week-old CTX mice at the indicated doses; 2 weeks later, mice were sacrificed for quantification of human and mouse CYP27A1 mRNA by qRT-PCR. WT littermates were included as a reference. Data are represented as relative mRNA content, using the housekeeping gene 36b4 as a reference and multiplied by a factor of 1,000 for easier visualization (WT and CTX controls, n = 10; treated CTX, n = 5). Data include pooled male and female mice because no difference was observed between the sexes. (C) One portion of liver samples was used to detect the CYP27A1 protein by western blot. Image shows a representative blot together with quantification of all samples (WT and CTX controls, n = 10; treated CTX, n = 5). Note that the antibody is able to detect both mouse and human protein using this technique. (D) Immunohistochemistry for detection of CYP27A1 (representative images) and quantification of positive hepatocytes. Bars represent means ± SEM for each group. ∗p < 0.05; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. Kruskal-Wallis with Dunn’s post test.