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Flavonoids lower Alzheimer's A? production via an NF?B dependent mechanism.

ABSTRACT: Alzheimer's disease (AD) is characterized by the brain accumulation of A? peptides and by the presence of neurofibrillary tangles. A? is believed to play an important role in AD and it has been shown that certain flavonoids can affect A? production. Recently, it was suggested that the A? lowering properties of flavonoids are mediated by a direct inhibition the ?-secretase (BACE-1) activity, the rate limiting enzyme responsible for the production of A? peptides. Westernblots and ELISAs were employed to monitor the impact of flavonoids on amyloid precursor protein processing and A? production. A cell free chemoluminescent assay using human recombinant BACE-1 was used to assess the effect of flavonoids on BACE-1 activity. The effect of flavonoids on NF?B activation was determined by using a stable NF?B luciferase reporter cell line. Molecular docking simulations were performed to predict the binding of flavonoids to the BACE-1 catalytic site. Real time quantitative PCR was used to determine the effect of flavonoids on BACE-1 transcription. We show in a cell free assay that flavonoids are only weak inhibitors of BACE-1 activity. Docking simulation studies with different BACE-1 structures also suggest that flavonoids are poor BACE-1 inhibitors as they appear to adopt various docking poses in the active site pocket and have weak docking scores that differ as a function of the BACE-1 structures studied. Moreover, a weak correlation was observed between the effect of flavonoids on A? production in vitro and their ability to lower BACE-1 activity suggesting that the A? lowering properties of flavonoids in whole cells are not mediated via direct inhibition of BACE-1 activity. We found however a strong correlation between the inhibition of NF?B activation by flavonoids and their A? lowering properties suggesting that flavonoids inhibit A? production in whole cells via NF?B related mechanisms. As NF?B has been shown to regulate BACE-1 expression, we show that NF?B lowering flavonoids inhibit BACE-1 transcription in human neuronal SH-SY5Y cells. Altogether, our data suggest that flavonoids inhibit A? and sAPP? production by regulating BACE-1 expression and not by directly inhibiting BACE-1 activity.

SUBMITTER: Paris D

PROVIDER: S-EPMC3124791 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Flavonoids lower Alzheimer's Aβ production via an NFκB dependent mechanism.

Paris Daniel D Mathura Venkat V Ait-Ghezala Ghania G Beaulieu-Abdelahad David D Patel Nikunj N Bachmeier Corbin C Mullan Michael M

Bioinformation 20110606 6

Alzheimer's disease (AD) is characterized by the brain accumulation of Aβ peptides and by the presence of neurofibrillary tangles. Aβ is believed to play an important role in AD and it has been shown that certain flavonoids can affect Aβ production. Recently, it was suggested that the Aβ lowering properties of flavonoids are mediated by a direct inhibition the β-secretase (BACE-1) activity, the rate limiting enzyme responsible for the production of Aβ peptides. Westernblots and ELISAs were emplo ...[more]

PMID: 21738321

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Project description:Several mutations conferring protection against Alzheimer's disease (AD) have been described, none as profound as the A673T mutation, where carriers are four times less likely to get AD compared to noncarriers. This mutation results in reduced amyloid beta (Aβ) protein production in vitro and lower lifetime Aβ concentration in carriers. Better understanding of the protective mechanisms of the mutation may provide important insights into AD pathophysiology and identify productive therapeutic intervention strategies for disease modification. Aβ(1-42) protein forms oligomers that bind saturably to a single receptor site on neuronal synapses, initiating the downstream toxicities observed in AD. Decreased formation, toxicity, or stability of soluble Aβ oligomers, or reduction of synaptic binding of these oligomers, may combine with overall lower Aβ concentration to underlie A673T's disease protecting mechanism. To investigate these possibilities, we compared the formation rate of soluble oligomers made from Icelandic A673T mutant and wild type (wt) Aβ(1-42) synthetic protein, the amount and intensity of oligomer bound to mature primary rat hippocampal/cortical neuronal synapses, and the potency of bound oligomers to impact trafficking rate in neurons in vitro using a physiologically relevant oligomer preparation method. At equal protein concentrations, mutant protein forms approximately 50% or fewer oligomers of high molecular weight (>50 kDa) compared to wt protein. Mutant oligomers are twice as potent at altering the cellular vesicle trafficking rate as wt at equivalent concentrations, however, mutant oligomers have a >4-fold lower binding affinity to synaptic receptors (Kd = 1,950 vs. 442 nM). The net effect of these differences is a lower overall toxicity at a given concentration. This study demonstrates for the first time that mutant A673T Aβ oligomers prepared with this method have fundamentally different assembly characteristics and biological impact from wt protein and indicates that its disease protecting mechanism may result primarily from the mutant protein's much lower binding affinity to synaptic receptors. This suggests that therapeutics that effectively reduce oligomer binding to synapses in the brain may be beneficial in AD.

Project description:RationaleAirway hyperresponsiveness (AHR) is classically found in asthma, and persistent AHR is associated with poor asthma control. Although airway smooth muscle (ASM) cells play a critical pathophysiologic role in AHR, the paracrine contributions of surrounding cells such as fibroblasts to the contractile phenotype of ASM cells have not been examined fully. This study addresses the hypothesis that nicotine promotes a contractile ASM cell phenotype by stimulating fibroblasts to increase nerve growth factor (NGF) secretion into the environment.MethodsPrimary lung fibroblasts isolated from wild type and α7 nicotinic acetylcholine receptor (α7 nAChR) deficient mice were treated with nicotine (50 µg/ml) in vitro for 72 hours. NGF levels were measured in culture media and in bronchoalveolar lavage (BAL) fluid from asthmatic, smoking and non-smoking subjects by ELISA. The role of the NFκB pathway in nicotine-induced NGF expression was investigated by measuring NFκB nuclear translocation, transcriptional activity, chromatin immunoprecipitation assays, and si-p65 NFκB knockdown. The ability of nicotine to stimulate a fibroblast-mediated, contractile ASM cell phenotype was confirmed by examining expression of contractile proteins in ASM cells cultured with fibroblast-conditioned media or BAL fluid.ResultsNGF levels were elevated in the bronchoalveolar lavage fluid of nicotine-exposed mice, current smokers, and asthmatic children. Nicotine increased NGF secretion in lung fibroblasts in vitro in a dose-dependent manner and stimulated NFκB nuclear translocation, p65 binding to the NGF promoter, and NFκB transcriptional activity. These responses were attenuated in α7 nAChR deficient fibroblasts and in wild type fibroblasts following NFκB inhibition. Nicotine-treated, fibroblast-conditioned media increased expression of contractile proteins in ASM cells.ConclusionNicotine stimulates NGF release by lung fibroblasts through α7 nAChR and NFκB dependent pathways. These novel findings suggest that the nicotine-α7 nAChR-NFκB- NGF axis may provide novel therapeutic targets to attenuate tobacco smoke-induced AHR.

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Flavonoids lower Alzheimer's A? production via an NF?B dependent mechanism.

Publications

Flavonoids lower Alzheimer's Aβ production via an NFκB dependent mechanism.

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