PMC:1283364 / 27512-30499 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/1283364","sourcedb":"PMC","sourceid":"1283364","source_url":"http://www.ncbi.nlm.nih.gov/pmc/1283364","text":"\nFigure 2 Aβ Levels Are Dramatically Reduced by Transgene Suppression\nCortical homogenates from young, predeposit tTA/APP mice used for Western blot in Figure 1 (line 107) were fractionated by sequential multi-step extraction with PBS, 2% SDS, and 70% FA followed by human-specific Aβ ELISA to measure transgene-derived peptide in each fraction. Aβ40 is shown in white, Aβ42 in black.\n(A) PBS-soluble Aβ levels are substantially reduced by both acute and chronic dox treatment (ANOVA, effect of treatment group F 4,24 = 137.10 and 386.01, p \u003c 0.001, for Aβ40 and Aβ42, respectively). Aβ levels in treated animals are indistinguishable from nontransgenic (NTg) animals (p \u003e 0.3, Tukey post-hoc test).\n(B) In the young animals tested here prior to the formation of visible amyloid deposits, most Aβ is extracted into the SDS fraction (84% and 76% of all transgene-derived Aβ40 and Aβ42, respectively). As in the PBS-soluble fraction, Aβ levels in the SDS fraction are significantly lowered by dox treatment compared to untreated animals (ANOVA effect of group F 4,24 = 197.57 and 163.48, p \u003c 0.001, for Aβ40 and Aβ42, respectively). Acutely treated animals retained a small (although significant) amount of residual peptide (p \u003c 0.001 compared to nontransgeinc, Tukey post-hoc test), whereas Aβ levels in mice born and raised on dox were reduced to levels indistinguishable from nontransgenic (p \u003e 0.8, Tukey post-hoc test).\n(C) The FA-soluble fraction already contains a small but significant pool of aggregated Aβ42 in untreated animals by 4 wk of age (p \u003c 0.05 versus nontransgenic; Tukey post-hoc test applied to significant effect of group ANOVA F 4,24 = 17.11, p \u003c 0.001). By 6 wk of age, the amount of Aβ in the FA fraction is increased significantly preceding the appearance of visible deposits 2 wk later. The FA pool is the only peptide fraction not lowered by acute dox treatment (4 wk untreated = 4 wk + 2 wk dox, p \u003e 0.9, Tukey post-hoc test), consistent with poor turnover of aggregated Aβ species.\n(D) Measurements of total Aβ, including both endogenous and transgene-derived peptides, show that animals born and raised on dox harbor Aβ levels identical to nontransgenic animals (p \u003e 0.9, Tukey post-hoc test, effect of group ANOVA F 4,24 = 39.13 and 35.29, p \u003c 0.001, for Aβ40 and Aβ42, respectively). Whereas chronic transgene suppression fully prevents synthesis of both peptides, acute dox treatment fully suppresses Aβ40 levels (p \u003e 0.8, Tukey post-hoc test), but leaves a small amount of nonsuppressed Aβ42. The residual Aβ42 observed in acutely treated young animals derives from uncleared aggregates extracted in the SDS and FA fractions. *, p \u003c 0.05; **, p \u003c 0.005; ***, p \u003c 0.001 versus 4-wk-old untreated mice, Tukey post-hoc applied to significant effect of group ANOVA. The ELISA data also confirmed that incorporation of the Swedish and Indiana mutations led to high levels of Aβ42, which we predicted would induce rapid plaque formation in untreated 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