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. 2015 Apr 22;35(16):6429–6443. doi: 10.1523/JNEUROSCI.3966-14.2015

Figure 1.

Figure 1.

Increased oxidative stress in PICK1−/− mouse brain. A, Total proteins extracted from 4-month-old WT and PICK1−/− (KO) mouse cortex (ctx), hippocampus (hip), and cerebellum (cb) were probed with antibodies to nitrotyrosine (nitro), 4-HNE (HNE), PICK1, and β-actin (actin). B, Levels of nitrotyrosine and 4-HNE intensity were normalized to the corresponding β-actin level, and percentage changes relative to corresponding WT are shown in the histograms: nitro, 100 ± 10% (WT, ctx), 270 ± 26% (KO, ctx), 100 ± 6% (WT, hip), 201 ± 12% (KO, hip), 100 ± 9% (WT, cb), 248 ± 25% (KO, cb); HNE, 100 ± 8% (WT, ctx), 295 ± 25% (KO, ctx), 100 ± 8% (WT, hip), 186 ± 22% (KO, hip), 100 ± 9% (WT, cb), 249 ± 27% (KO, cb). n = 4 pairs. C, Immunoreactivity for nitrotyrosine in the CA3, forebrain (FB), and CA1 from 4-month-old WT and KO mice. Dashed lines show pyramidal cell layers. The higher magnification of areas in the white boxes shows the different staining intensity in CA3 and FB areas between WT and KO mice, but no difference was found in CA1 pyramidal layers. Scale bars, 50 μm. D, Expression of nitrotyrosine and 4-HNE in WT and KO cortices at 1, 2, 4, and 12 months. Signal intensity ratios (nitrotyrosine/actin) were as follows: WT, 1.4 ± 0.5% (1 month), 5.5 ± 1.4% (2 months), and 25.4 ± 7.1% (12 months); KO, 1.5 ± 0.5% (1 month), 8.8 ± 2.5% (2 months), and 76.8 ± 13.3% (12 months). Signal intensity ratios (4-HNE/actin) were as follows: WT, 1.2 ± 0.4% (1 month), 7.8 ± 1.8% (2 months), and 21.7 ± 6.6% (12 months); KO, 1.2 ± 0.4% (1 month), 11.9 ± 3.4% (2 months), and 79.4 ± 16.2% (12 months). n = 4 pairs. Note that levels of both nitrotyrosine and 4-HNE in WT mice were significantly increased with age. E, Concentrations of nitrotyrosine in serum samples from controls (ctrl) and PICK1-mutant patient (mut). “single” indicates each ELISA test performed in one subject. All tests from the same subject were averaged and labeled as “average.” Averaged concentrations of nitrotyrosine were 154 ± 5 ng/ml (mut, n = 4), 53 ± 1 ng/ml (ctrl1, n = 2), 34 ± 1 ng/ml (ctrl2, n = 2), 26 ± 1 ng/ml (ctrl3, n = 2), and 35 ± 1 ng/ml (ctrl4, n = 2). *p < 0.05, **p < 0.01.