Nox2-derived radicals contribute to neurovascular and behavioral dysfunction in mice overexpressing the amyloid precursor protein

Abstract

Alterations in cerebrovascular regulation related to vascular oxidative stress have been implicated in the mechanisms of Alzheimer's disease (AD), but their role in the amyloid deposition and cognitive impairment associated with AD remains unclear. We used mice overexpressing the Swedish mutation of the amyloid precursor protein (Tg2576) as a model of AD to examine the role of reactive oxygen species produced by NADPH oxidase in the cerebrovascular alterations, amyloid deposition, and behavioral deficits observed in these mice. We found that 12- to 15-month-old Tg2576 mice lacking the catalytic subunit Nox2 of NADPH oxidase do not develop oxidative stress, cerebrovascular dysfunction, or behavioral deficits. These improvements occurred without reductions in brain amyloid-beta peptide (Abeta) levels or amyloid plaques. The findings unveil a previously unrecognized role of Nox2-derived radicals in the behavioral deficits of Tg2576 mice and provide a link between the neurovascular dysfunction and cognitive decline associated with amyloid pathology.

Fig. 1.

Nox2 inactivation rescues the cerebrovascular dysfunction in Tg2576 mice. CBF responses to whisker stimulation (A) and topical neocortical application of acetylcholine (10 μM) (B), bradykinin (50 μM) (C), or adenosine (400 μM) (D) in young and aged WT, Nox2^−/−, Tg2576, or Tg2576/Nox2^−/− mice. *, P < 0.05 from young WT; #, P < 0.05 from young Tg2576; analysis of variance and Tukey's test; n = 5 per group.

Fig. 2.

Nox2 inactivation attenuates neuronal and vascular oxidative stress in 12–15-month-old Tg2576 mice. (A) Wild-type mice. Neurons immunolabeled with NeuN (a) are colocalized with the ROS (HE) signal (b), as shown by the merged image (c). Endothelial cells immunolabeled with CD31 (d) are also colocalized with the ROS signal (e and f). (Scale bar: 50 μm.) (B) Tg2576 mice. NeuN-positive neurons (a) are colocalized with the ROS signal (b and c). CD31-positive endothelial cells (d) are also colocalized with the ROS signal (e and f). (Scale bar: 50 μm.) (C) The increased ROS signal in Tg2576 is attenuated by the ROS scavenger MnTBAP and a NADPH peptide inhibitor (gp91-ds) but not by a scrambled peptide (sgp91ds) or in Tg2576/Nox2^−/− crosses. *, P < 0.05 from WT; analysis of variance and Tukey's test; n = 5 per group.

Fig. 3.

Short-term application of ROS scavengers counteracts the cerebrovascular dysfunction in 12- to 15-month-old Tg2576 mice. Cerebrovascular responses are rescued by a 30-min neocortical application of MnTBAP (A–C) or gp91ds-tat (gp91ds) (E–G) but not by a scrambled peptide (sgp91) (E–G). Responses to adenosine are not attenuated in Tg2576 mice (D and H). *, P < 0.05 from vehicle; analysis of variance and Tukey's test; n = 5 per group. V, vehicle.

Fig. 4.

Brain Aβ and amyloid burden are not reduced in 12- to 15-month-old Tg2576 mice lacking Nox2. (A and B) SDS-soluble (A) and -insoluble (B) Aβ are not different in Tg2576 and Tg2576/Nox2^−/− mice (P > 0.05; analysis of variance and Tukey's test; n = 12 per group). (C and D) The amyloid load is comparable between Tg2576 (C) and Tg2576/Nox2^−/− mice (D). (E and F) The number of plaques (E) and plaque load (F) do not differ between Tg2576 and Tg2576/Nox2^−/− mice (P > 0.05; n = 6 per group).

Fig. 5.

Nox2 inactivation rescues the behavioral deficits in Tg2576 mice tested in a Y maze. The reduction in novel arm entry (A), time spent in the novel arm (B), and the increase in total arm visits (C) and defecation (D) observed in Tg2576 mice are not seen in Tg2576/Nox2^−/− mice. *, P < 0.05, χ² test in A and analysis of variance and the Newman–Keul test in B–D; n = 10–15 per group.