Screening And Interaction Of Leading Compounds With Beta Amyloid

Alzheimer disease (AD) is the most common form of dementia. The two major pathological hallmarks of AD are extra cellular amyloid plaques, which are formed mainly from the amyloid-β (Aβ) peptide, and intracellular neurofibrillary tangles (NFTs), which contain hyperphosphorylated tau. According to the amyloid cascade hypothesis, which emphasizes a central role for AP in the pathogenesis of AD, Aβ has become a major therapeutic target with increasing pathological, biochemical and genetic evidences that supports a role for this molecule in the disease process. On the basis of this hypothesis, various anti-Aβ strategies including lowering the production of the peptide, preventing the formation of Aβ aggregates, and increasing the rate of Aβ clearance from the brain have been pursued.The Aβ peptide, which comprises40-42amino acids, is generated following proteolytic cleavage of the amyloid precursor protein (APP). Naturally-derived AP monomers are soluble stable, but appear neurotoxic effect once they become insoluble aggregation state. It has been shown that Aβ1-40and Aβ1-42possess different biochemical properties, and Aβ1-42is considered to be the major etiologic agent in the pathogenesis of AD due its more aggressive aggregation or oligomerization properties identified through NMR and other technique. It is necessary for investigating the different Aβ properties and exploring drugs targeting at Aβ to set up optional experimental conditions including temperature.In the present study, using isothermal titration calorimetry with little modification, circular dichroism spectra, and atomic force microscope, we first investigated the aggregation process of Aβ1-42and Aβ1-40at different temperatures including4,25, and37℃without stirring, then picked out leading compounds with potential anti-Aβ aggregation ability by use of surface plasmon resonance to, and finally corroborated the ability of representative compound resveratrol (RES) through fluorescence of thioflavin T, atomic force microscope, and1H-NMR. Our results are as follows:Neither Aβ1-40nor Aβ1-42aggregated on the condition of incubating at4℃for24h without stirring. However, the results from isothermal titration calorimetry showed the thermodynamics change of both Aβ1-40and Aβ1-42during the incubation process at25or37℃for24h, which was supported by the result from circular dichroism spectra in which both Aβ1-40and Ap1-42showed secondary structural change, and was consistent with the data from atomic force microscope with particle diameter changes. These results demonstrated that both Aβ1-40and Aβ1-42could self-aggregate at25or37℃, but not4℃.Using surface plasmon resonance technique, we identified that the compounds Congo red and ThS could bind not only monomer Aβ1-40and Aβ1-42, but also fibril Aβ1-40and Aβ1-42, which was consistent with the results from other group. These results demonstrated that surface plasmon resonance is one of the effective techniques on preliminary screening of compounds with potential potential anti-Aβ aggregation ability.Using surface plasmon resonance technique, we picked out some compounds that could bind to monomer or fibril Aβ1-40and Aβ1-42, with weaker binding ability than that of Congo red. Furthermore, we found different binding ability to monomer or fibril Aβ1-40and Aβ1-42of the same compound in our study, and we need design further studies to find out whether the mechanism of which being relative with the different secondary structures between monomer and fibril Aβ.Resveratrol could bind to both monomer and fibril Apl-40or Aβ1-42with the concentration ranging from1.56to100μM, and could weaken the ThT intensity, change the1H-NMR spectrum and particle diameter while incubating with monomer or fibril Aβ1-40and Aβ1-42at concentration of50μM. These results demonstrated that resveratrol could perturb and even inhibit the self-aggregation process of Aβ. The identity of the senile plaques stained with resveratrol was confirmed by staining serial sections in hippocampus tissue from AD brains compared with thioflavine S.Lipopolysaccharide (250μg/kg) injected intraperitoneally once could not induce spatial learning and memory impairment in ICR mice, and there was no effect of RES on spatial learning and memory of ICR mice by intragastric administration with dose of10mg/kg for21days. However, ICR mice are competent enough for MWM with gender differences that females perform better in both the acquisition phase and the test phase than males.Taking together, our results indicated that fibril Aβ1-40and fibril Aβ1-42could aggregate at25℃for24h without stirring, and that SPR could be taken as an effective method to screen out compounds with potential ability of anti-aggregation of Aβ. Furthermore, our results demonstrated that RES could inhibit the self-aggregation of Aβ and bind to senile plaques in vitro. Additionally, our results showed that ICR mice could used in MWM task with consideration of the gender differences that females perform better in both the acquisition phase and the test phase than males.