Fig. 2

BTZ effect on P-gp protein expression and transport activity in brain capillaries isolated from hAPP mice. A Representative Western blot for P-gp showing bands for brain capillary membranes isolated from wild-type (WT), hAPP, hAPP treated with 0.25 mg/kg BTZ, and hAPP treated with a combination of BTZ/CSA (0.25/25 mg/kg). β-Actin was used as protein loading control. Tissue was pooled from all mice per treatment group (WT: n = 20 mice; hAPP: n = 18 mice; hAPP-BTZ: n = 20 mice, hAPP-BTZ/CSA: n = 19 mice). B Densitometric analysis of three western blots for P-gp levels. Blotting and probing were performed on three separate gels, as described in the main text. Data were analyzed by mixed-level generalized linear models to account for individual blot effects. Columns represent model-estimated marginal means (WT = 1), excluding random effects ± SEM for 3 blots. C Representative confocal images of isolated brain capillaries from WT, hAPP, hAPP-BTZ, and hAPP-BTZ/CSA mice showing luminal accumulation of the P-gp-specific substrate [N-ε (4-nitrobenzofurazan-7-yl)-D-Lys8]-cyclosporine A (NBD-CSA) after a 1-h incubation (steady state; 2 μM NBD-CSA). D Data obtained by image analysis using ImageJ. Specific NBD-CSA fluorescence is the difference between total luminal NBD-CSA fluorescence and NBD-CSA fluorescence in the presence of the P-gp-specific inhibitor PSC833 (5 μM). Columns represent mean ± SEM for 10 capillaries from one preparation (pooled tissue: WT: n = 20 mice; hAPP: n = 18 mice; hAPP-BTZ: n = 20 mice, hAPP-BTZ/CSA: n = 19 mice), as arbitrary fluorescence units (scale 0–255). The ω² is an unbiased estimator of the coefficient of determination (R²) for an effect. Symbols indicate statistical categories at p ≤ 0.05 derived from pairwise comparisons adjusted for false discovery rate. Treatments sharing a symbol did not differ at p ≤ 0.05