Verbenalin Reduces Amyloid-Beta Peptide Generation in Cellular and Animal Models of Alzheimer's Disease

Abstract

Verbenalin, among the major constituents of Verbena officinalis, has been reported to exhibit sleep-promoting and antioxidant activities. This study demonstrates the effects of verbenalin on amyloid-beta (Aβ) peptide generation in Swedish mutant amyloid precursor protein (APP)-overexpressing Neuro2a cells (SweAPP/N2a) and in Alzheimer's disease (AD) animal models. We further performed molecular biological analyses of these in vitro and in vivo models of AD. The effects of verbenalin were assessed based on the expression of factors related to Aβ peptide production using Western blotting, enzyme-linked immunosorbent assay, and immunohistochemistry (IHC). The intracellular expression and release of APP protein were both decreased by verbenalin treatment in SweAPP/N2a cells. Thus, the production of Aβ peptides was decreased. Compared to those in AD transgenic (Tg) mice, IHC revealed that verbenalin-treated animals showed decreased Aβ and tau expression levels in the hippocampus. In addition, verbenalin restored the expression of brain-derived neurotrophic factor (BDNF) in the hippocampus of AD animal models. These findings suggest that verbenalin may decrease Aβ formation both in vitro and in vivo. Verbenalin may also help improve the pathological hallmarks of AD.

Keywords: Alzheimer’s disease (AD); amyloid beta; brain-derived neurotrophic factor; tau; verbenalin.

Figure 1

Effects of verbenalin on APP expression in SweAPP/N2a cells. (A) SweAPP/N2a cells were treated with the indicated verbenalin concentration for 24 h. APP protein levels in cell lysate and supernatant were analyzed by Western blotting. β-actin was used as loading control. (B) Data were quantified by densitometric analysis. Results are represented as the mean ± SEM. * p < 0.05, ** p < 0.01, and *** p < 0.001 by one-way analysis of variance (ANOVA).

Figure 2

Effects of verbenalin on Aβ peptides generation in SweAPP/N2a cells. (A) Swe APP/N2a cells were treated with the indicated verbenalin concentration for 24 h. The levels of Aβ 42 (A) and Aβ 40 (B) were determined using ELISA. Data were quantified by densitometric analysis. Results are represented as the mean ± SEM. * p < 0.05 by Student’s t-test.

Figure 3

Effects of verbenalin on Aβ- and tau-positive fibers in hippocampus. Aβ (A) and tau (B) expression is examined using IHC. N: normal group (C57BL/6J mice), C (B6Cg-Tg mice): control group, D: donepezil group, VM: verbenalin medium concentration group (100 mg/kg), VH: verbenalin high concentration group (200 mg/kg), CA1: cornu ammonis area 1, CA2: cornu ammonis area 2, CA3: cornu ammonis area 3, PoDG: polymorphic layer of the dentate gyrus, and DG: dentate gyrus.

Figure 4

Effects of verbenalin on BDNF-positive fibers in hippocampus. BDNF expression is examined using IHC. N: normal group (C57BL/6J mice), C (B6Cg-Tg mice): control group, D: donepezil group, VM: verbenalin medium concentration group (100 mg/kg), VH: verbenalin high concentration group (200 mg/kg), CA1: cornu ammonis area 1, CA2: cornu ammonis area 2, CA3: cornu ammonis area 3, PoDG: polymorphic layer of the dentate gyrus, and DG: dentate gyrus.