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. 2012 Aug 29;135(9):2661–2675. doi: 10.1093/brain/aws209

Figure 2.

Figure 2

Expression of lysosomal enzymes in brain of Gnptabc.3082insC mice. (A) Brain homogenates from 4-month-old wild-type and Gnptabc.3082insC mice were analysed by western blotting using antibodies against mannose 6-phosphate residues (M6P), cathepsin D (CtsD), cathepsin B (CtsB), Npc2 and Lamp1. β-Tubulin and Gapdh were used as loading controls. p = precursor; m = mature form. (B) Immunodetection of M6P residues on lysosomal proteins demonstrate lack of M6P in cortical brain sections of 1-month-old Gnptabc.3082insC mice. Scale bars = 50 µm (left panel), 10 µm (right panel). (C) The relative enzyme activities of the lysosomal hydrolases β-hexosaminidase (β-hex), β-galactosidase (β-gal), α-mannosidase (α-man), α-l-fucosidase (α-fuc), arylsulphatase A (asa), cathepsin B (CtsB) and cathepsin D (CtsD) were measured in homogenates of perfused wild-type and Gnptabc.3082insC whole-brain tissue of 4-month-old mice. The specific activities of the wild-type were set to 1 (mean ± SD, n = 3, *P < 0.05). (D) The relative messenger RNA level of the lysosomal hydrolases β-hexosaminidase (Hexa), β-galactosidase (Glb1), α-mannosidase (Man2b1), α-l-fucosidase (Fuca1) and arylsulphatase A (Arsa), cathepsin D (Ctsd), Npc2, Lamp1 and the phosphotransferase subunits encoding genes Gnptab and Gnptg were determined by real-time PCR and normalized to β-actin messenger RNA expression (mean ± SD, n = 3, *P < 0.05).