Valsartan lowers brain beta-amyloid protein levels and improves spatial learning in a mouse model of Alzheimer disease

Abstract

Recent epidemiological evidence suggests that some antihypertensive medications may reduce the risk for Alzheimer disease (AD). We screened 55 clinically prescribed antihypertensive medications for AD-modifying activity using primary cortico-hippocampal neuron cultures generated from the Tg2576 AD mouse model. These agents represent all drug classes used for hypertension pharmacotherapy. We identified 7 candidate antihypertensive agents that significantly reduced AD-type beta-amyloid protein (Abeta) accumulation. Through in vitro studies, we found that only 1 of the candidate drugs, valsartan, was capable of attenuating oligomerization of Abeta peptides into high-molecular-weight (HMW) oligomeric peptides, known to be involved in cognitive deterioration. We found that preventive treatment of Tg2576 mice with valsartan significantly reduced AD-type neuropathology and the content of soluble HMW extracellular oligomeric Abeta peptides in the brain. Most importantly, valsartan administration also attenuated the development of Abeta-mediated cognitive deterioration, even when delivered at a dose about 2-fold lower than that used for hypertension treatment in humans. These preclinical studies suggest that certain antihypertensive drugs may have AD-modifying activity and may protect against progressive Abeta-related memory deficits in subjects with AD or in those at high risk of developing AD.

Figure 1. Valsartan prevents oligomerization of Aβ_1–42 peptide into a HMW Aβ species in vitro.

(A) Western blot analysis of Aβ_1–42 oligomers in the presence of losartan, valsartan, carvedilol, hydralazine, propranolol, nicardipine, or amiloride. Bands at 3.5 kDa represent the monomeric Aβ form, whereas the smear between 55 and 130 kDa represents the oligomeric form of Aβ. (B) Valsartan decreases the accumulation of HMW Aβ_1–42 species. Inset: Nonaggregated Aβ_1–42 control (lane 1). Representative Western immunoblot of Aβ_1–42 in the absence (lane 2) or presence of 10 μM valsartan (lane 3). (C) Quantitative dot blot analysis of valsartan inhibition of Aβ_1–42 oligomerization. The same samples used in B were subjected to dot blot analysis using oligomer-specific antibody A11. Inset: Representative dot blot image. Results are expressed as percent of negative control (negative control presents nonaggregated Aβ), and values represent mean ± SEM. *P < 0.05.

Figure 2. Chronic valsartan treatment is highly tolerable in Tg2576 mice.

Valsartan was provided to female Tg2576 mice from 6 to 11.5 months of age at 10 mg/kg/d or 40 mg/kg/d. (A and B) Body weight and fluid consumption were monitored weekly. (C) Postprandial glucose tolerance response was examined after 5 months of valsartan treatment. (D) Tg2576 blood pressure measurements in response to approximately 5 months of valsartan treatments. (E) Baseline measurements of systolic and diastolic blood pressure and mean arterial blood pressure (MAP) in adult female Tg2576 mice and strain-, age-, sex-matched WT mice. The blood pressure determination for each animal was calculated as the mean of 10 individual measurements. Values represents group mean values ± SEM; n = 7–9 mice per group.

Figure 3. Chronic valsartan treatment of Tg2576 mice resulted in dose-dependent attenuations of AD-type spatial memory deterioration in Tg2576 mice, which is coincidental with significant reductions in HMW soluble Aβ species and AD-type neuropathology in the brains of Tg2576 mice.

(A) The influence of Aβ-related spatial memory in response to valsartan treatment at 10 and 40 mg/kg/d versus the untreated control Tg2576 mice was assessed using an MWM test in approximately 11-month-old female Tg2576 mice. Latency score represents time taken to escape to the platform from the water. (B) Assessments of soluble, extracellular HMW Aβ peptide contents in the brain using an antibody specific for HMW oligomeric Aβ peptides in a dot blot analysis. Inset: Representative dot blot analysis of HMW soluble Aβ contents. (C) Assessment of total PBS-soluble Aβ peptide using ELISA assay. (D) Assessment of Aβ_1–42 and Aβ_1–40 peptide concentrations in the cerebral cortex and hippocampus of valsartan-treated (10 or 40 mg/kg/d) or control mice. (E) Stereological assessment of cerebral cortex and hippocampal Aβ plaque burden in valsartan-treated or control mice expressed as thioflavin-S–positive volume as a percentage of regional volume. Inset: Representative photograph of thioflavin-S–positive Aβ plaque neuropathology in neocortex (CTX) and hippocampal formation (H) in untreated control (left panel) and valsartan-treated (40 mg/kg/d) Tg2576 mice (right panel). Arrowheads point toward thioflavin-S positive amyloid plaques. Original magnification, ×125. Values represent group mean ± SEM; n = 7–9 mice per group. In B and C, *P < 0.001; In D and E, *P < 0.05, **P < 0.01; 1-way ANOVA followed by Newman-Keuls post-hoc analysis.

Figure 4. Valsartan treatments prevented cognitive impairment and attenuate AD-type neuropathology, in part, by promoting membrane-bound insulin degradation enzyme activity.

(A) APP content in the cortex of valsartan-treated or untreated control Tg2576 mice. Inset: Representative immunoreactive APP (C8 antibody) and β-actin signals. (B) Assessments of cellular α-, β-, and γ-secretase activities in the cerebral cortex of Tg2576 mice in response to valsartan treatment. (C) Assessment of Aβ_1–42 and Aβ_1–40 peptide content in peripheral blood (serum). (D) Assessments of CM-associated (left panel) and cytosolic (right panel) IDE activity in the cerebral cortex of Tg2576 mice in response to valsartan treatment. Inset: Representative immunoblot signals of membrane and cytosolic IDE protein content from the same samples. (E) Assessments of neprilysin content by Western blotting using a commercial rabbit anti-mouse neprilysin antibody. Inset: Representative neprilysin and actin protein signals from the same blot. (F) Assessment of ECE activity using an Endothelin-1 ELISA System. Values represent group mean ± SEM; n = 7–9 mice per group. *P < 0.05; 1-way ANOVA, followed by Newman-Keuls post-hoc analysis.