Natural Medicines Against Aβ42in Alzheimer's Disease And Their Molecular Mechanism

Alzheimer's disease (AD) is neurodegenerative diseases among older peoplewhich leads to memory loss and cognitive dysfunction and is difficult to heal afterthe onset. AD is characterized pathologically by the deposition β-amyloid protein(Aβ) which further formation of senile plaques (SP), and intracellularneurofibrillary tangles caused by hyperphosphorylation of Tau protein and neuronsnecrosis. Abundant evidences suggest that oligomeric forms of β-amyloid proteinare the main neurotoxins causing AD. So how to safely and effectively prevent oreven remove the toxic Aβ42peptide is considered to be an effective method of theprevention and treatment of AD. With Aβ42oligomers as the target of AD, thethesis aimed at finding the natural medicines which could inhibit the neurotoxicityof Aβ42oligomers and could really through the blood-brain barrier (BBB) to takeeffect on AD, and investigating the preliminary mechanism to the Aβ42inhibition.We first screened the Chinese medicine for the treatment of AD, includingCistanche, Epimedium, Acorus gramineus, and Poria, Astragalus, fleece-flowerroot, Rhodiola, Polygala, Gynostemma, Tianma, Angelica, Sophora, rehmannia,cnidium fruit, treating diabetes, Ligustrum lucidum, a total of16kinds of Chineseherbal medicines as the natural medicine which has the potential treatment ofAlzheimer's disease. Through the high-throughput screening to test their affect onthe anti-Aβ42oligomers neurotoxicity, we identified10compounds that mayinhibit the cytotoxicity of Aβ42, then according to the extract concentration ofactive ingredient, the effect which reported in the literature, and consideration ofthe blood-brain barrier, two compounds: ferulic acid (sample A) and matrine(sample B) were finally choosed as the potential target compounds with thefunction of anti-Aβ42cytotoxicity. Using the general model SH-SY5Y cell for AD,the two compounds were further detected their inhibitory effects on the Aβ42aggregation at the cellular level and molecular level and the inhibitory mechanismof the two compounds were also investigated. The inhibitory effects of sample A and B on Aβ42-induced cytotoxicity werefirst examined in vitro and the results showed that sample A showed the protectiveeffects on Aβ42monomer and oligomer-induced cytotoxicity in SH-SY5Y cells,and sample B showed the same protective effects both in PC12cells andSH-SY5Y cells. These results showed that both sample A and B have theneuroprotective effect against Aβ42. On the otrher hand, the influence of sample Aand B on the secondary structures of Aβ42aggregates was measured by CDspectroscopy, the result suggested that sample A could inhibit the β-sheetformation in Aβ42monomer, or could accelerate the β-sheet formation in Aβ42oligomer, whereas sample B could inhibit the β-sheet formation both in Aβ42monomers and oligomers. The subsequent results of Transmission electronmicroscope (TEM) observation and Th-T fluorescence staining also supported theabove results from the CD spectroscopy. These results indicated that sample Acould prevent Aβ42monomer from aggregating or fibrillating, or accelerate theAβ42aggregations from oligomer to fibril, while sample B could always inhibitthe aggregation of Aβ42. The specificity of sample A and B to Aβ42monomer,oligomer or fibril were evaluated in vitro by binding assay. The results showedthat sample A and B generally displayed the significant binding affinities to all thethree Aβ42species and the specificity of sample A to monomer and fibril isstronger than oligomer, but sample B does to oligomer and fibril than monomer.In view of the lesion area of AD, if the drugs can pass effectively through theblood-brain barrier (BBB) is an important character that must be considered. Weexamined the efficiencies of sample A and B to pass through the membrane ofSH-SH5Y cell, the results indicated that sample A could pass through the cellmembrane of SH-SH5Y cell, but no data were obtained in sample B experiment.In the other study, sample B was found in the brain of mice that were gavagedwith sample B, suggested sample B could pass through the BBB. Based on theMTT assay of the serum and brain samples of the gavaged mice, we confirmedthat sample B could still inhibit the Aβ42oligomer-induced cytotoxicity after themetabolism.The above results confirmed that sample A and B have interaction with Aβ42,and sample A and B could changed the Aβ42aggregation state, then ultimately inhibited Aβ42-induced neurotoxicity by this interaction. However, what is themolecular mechanism of this interaction? Docking analysis was used to simulatethe interaction of these two compounds and Aβ42monomer molecular. Thesimulation images showed that the sample A and B are able to approach to andinteract with Aβ42based on the different mechanism. Sample A could interactwith Aβ42mainly by the hydrogen bonds. Sample B mainly relied on theelectrostatic interactions with Aβ42.In summary, two natural compounds, ferulic acid (sample A) and matrine(B sample), obtained through the high-throughout screening, could alter (inhibitor promote) the aggregation state of Aβ42, leading to decreasing the contents ofAβ42oligomer, the most neurotoxic species of Aβ42. The results of animalexperiments confirmed that the sample B could pass through the BBB into thebrain and still inhibits the Aβ42oligomer-induced cytotoxicity. Preliminarydocking simulation images showed that sample A and B interact with Aβ42by thedifferent mechanisms. All these results provided a theoretical guidance for theapplication of natural medicines and drug design in AD therapy.