Figure 2.
Rescue of lethal phenotype in GFAPTg;Cryab−/− mice by astrocyte-specific restoration of Cryab expression. (A) The Tg141 transgene, consisting of the human GFAP promoter ligated to the hamster Cryab gene, was constructed to over-express Cryab in astrocytes. (B) Several founder lines were generated (Tg141 1–4, 6, 9) with varying levels of Cryab expression, as shown by western analysis. (C) Survival proportions show lethality in 54% of GFAPTg;Cryab−/− mice (n = 13) compared with ∼10% of GFAPTg;Cryab−/−;CryabTg (n = 9) and GFAPTg;Cryab+/+ (n = 10) littermates (P = 0.0281, log-rank Mantel–Cox test). (D) Rosenthal fibers under light microscopy are eosinophilic (red) and appear to outline astrocytic processes in the GFAPTg;Cryab−/−;CryabTg mice (50 µm scale bar). (E) Fibers in GFAPTg;Cryab−/−;CryabTg mice have the same ultrastructural qualities as those in GFAPTg mice (1 µm scale bar). (F, G) Immunofluorescence shows the absence of Cryab in null mice (F) compared with wild-type (G, 50 µm scale bar) with confocal microscopy using equivalent gain. (H, J, L) Expression of the CryabTg (green) in Cryab−/− animals is astrocyte-specific (GFAP, red). (L) The inset shows an enlarged view of an astrocyte from the same field expressing Cryab (yellow). (I, K, M) No Cryab (green) staining is apparent in neurons (NeuN, red). (F–M) Olfactory bulb (granular layer) shown at same magnification.