Figure 2.

Generation and Analyses of FH-Expressing Transgenic Mice to Investigate the Role of Cytosolic FH/Fumarate in Renal Cyst Development

(A) The FH and FH^cyt transgenes were cloned into the CB92 vector and targeted to the Rosa26 locus, using phage-mediated recombination (Chen et al., 2011).

(B and C) Localization of FH and FH^cyt was confirmed in ES cells by immunocytochemistry. Colocalization (yellow) of FH (green) and mitochondria (red) is evident (B), whereas FH^cyt is absent from the mitochondria (C). Nuclei (blue) are visualized with DAPI.

(D) Kidneys harvested from 30-week-old control, transgene-only (FH, FH^cyt), and genetically “rescued” mice (FH1KO+FH and FH1KO+FH^cyt) appeared macroscopically normal compared to FH1KO kidneys, which appeared enlarged and cystic.

(E) Hematoxylin and eosin (H&E) staining of kidneys harvested from mice at 13, 20, and 30 weeks reveal that transgenic expression of either FH or FH^cyt is sufficient to ameliorate cyst development.

(F–H) Numbers and frequency of macrocysts (>0.5 mm) and microcysts (>0.1 mm) were determined in each group (n = 6). Error bars represent the SEM.

Scale bars, 5 μm (B and C), 10 mm (D), and 100 μm (E). Error bars indicate SEM. See also Figure S1.