FIG 3.

AAV transduction in Ext1^f/f control and Ext1^HEP mutant mice. Shown is an analysis of liver transduction 8 weeks after intravenous injection of 1 × 10¹¹ VP/mouse of an AAV2 vector encoding green fluorescent protein (AAV-GFP) in Ext1^f/f control or Ext1^HEP mutant mice. (a) GFP immunohistochemistry in livers from Ext1^f/f control or Ext1^HEP mutant mice. GFP is visible as brown staining in individual hepatocytes (red arrows). The graph depicts quantification of GFP-positive cells in liver sections. (b) GFP content in liver extracts from Ext1^f/f control (Ext1^f/f Cre−) or Ext1^HEP mutant (Ext1^f/f Cre+) mice. Liver extracts were assayed for GFP and GAPDH by immunoblotting. “+” indicates the presence of the AlbCre transgene, leading to the targeted deletion of the Ext1 gene to create Ext1^HEP mutant mice; “−” indicates the absence of the AlbCre transgene, resulting in Ext1^f/f mice with an intact Ext1 gene. (c) Total liver RNA was analyzed for GFP mRNA expression by quantitative real-time RT-PCR and normalized to the mouse liver GAPDH mRNA of the same samples. (d) AAV vector genomes in livers from AAV-injected Ext1^f/f control or Ext1^HEP mutant mice were quantified after DNA extraction by quantitative real-time PCR, using primers within the CMV promoter of the transgene expression cassette, and normalized to liver GAPDH DNA values of the same sample. Values are means ± SD. Differences in number of GFP-expressing cells per field, GFP mRNA expression, and vector genomes between control and mutant mice were compared by Student's t test. *, P < 0.05; n = 3. Scale bar represents 50 μm.