Study On Anti-AD Activity And Mechanisms Of Terminalia Chebula Retz.

The dried fruit of Terminalia chebula Retz.,（T.Chebula）has a long history in medicine in China,recored in varieties of ancient medicical books.In the Tibetan medicine system,possessing significant effect in the treatment of several symptoms in folk.Recently,studies had revealed that T.Chebula has significant activity in anti-tumor,anti-inflammatory and analgesic and so on.In our previous study,T.Chebula showed a strong antioxidant activity in a screen work amind compare 40 tibetan medicines antioxidant capacity.The occurrence and development of Alzheimer’s disease is closely related to the homeostasis of oxidative stress.Therefore,our lab used transgenic C.elegans model to carry out a large number of studies on the anti-AD activity of T.Chebula..The results showed that T.Chebula inhibiting Aβaggregation by regulating the daf-16 nuclear translocation and the expression of DAF-16 and Hsp16.2.AD transgenic C.elegans model produces Aβpathogenesis by integrating the human-derived Aβ_1-42 gene into nematode tissue.In C.elegans,because of APL-1,the homolog of APP,lacks the Aβsequence and does not possess the activity ofβ-secretase,a key enzyme in Aβshearing,which leading frequently in drug screening and limit in-depth study in Aβproduction and degradation.Therefore,we investigated the effect of T.Chebula on the amelioration of exogenous Aβ-induced oxidative damage in SH-SY5Y cells,and the effect onα/β/γ-secretase activity and of Aβdegradation in APPswe transgenic SH-SY5Y cells.The study comprehensively elucidated the anti-AD activity and mechanism of T.Chebula,it also evaluated the effective components of T.Chebula that exerted anti-AD activity.The main research contents and results of the work are as follows.The study found that T.Chebula water extract（TWE）can protect the oxidative damage of human neuroblastoma cells SH-SY5Y induced by AβO.After 24 h of AβO exposure,the cells showed Ca²⁺abnormally increased（62.8%higher than control）,blocked ATP synthesis（33.2%lower than control）,up-regulated cellular ROS levels,and depolarized mitochondrial membrane potential,while inducing Cyt C/Caspase-3-dependent apoptosis.After administration of 40μg/m L TWE,the level of Ca²⁺was decreased 30.4%,the level of ATP was increased 33.0%,the levels of ROS and MMP were improved,and the level of cell apoptosis was decreased 8.23%,which effectively protected abnormal Ca²⁺metabolism,and mitochondrial metabolic disorders induced by AβO.Studies showed that the level of Aβ_1-40 and Aβ_1-42 were decreased in a time-dependent manner after TWE treatment in transgenic cells.Specifically,after 24 h and48 h of TWE at 80μg/m L,Aβ_1-40 decreased 39.6%and 42.8%,and Aβ_1-42 decreased36.0%and 50.7%,respectively.After 80μg/m L TWE acted on SH-SY5Y/APPswe cells for 24 h and 48 h,the levels of extracellular Aβ_1-40 reduced 39.6%and 42.8%,Aβ_1-42decreased by 36.0%and 50.7%,respectively.For APP cleavage-protease,ADAM10expression increased 1.37-fold,BACE1 expression decreased 2.1-fold,and 1.24-fold reduce of PS1 expression.The level of s APPαincreased 4.93-fold and s APPβdecreased1.21-fold.The results indicating that TWE inhibited Aβgeneration by block the amyloid metabolic pathway and promoted the non-amyloid metabolic pathway of APP protein.In addition,TWE upregulated autophagic activity in SH-SY5Y/APPswe cells,and the blockade of autophagy by the autophagy inhibitor CQ resulted in a 1.27-fold increase in extracellular Aβ_1-42 levels,a 1.41-fold increase in ADAM10 levels,a 1.23-fold increase in BACE1 levels,a 1.72-fold increase in s APPαlevels,and a 1.17-fold increase in s APPβlevels,indicating TWE-induced autophagy involved in inhibiting extracellular Aβ_1-42 accumulation.Screening of components in TWE found that chebulinic acid（CN）may be an important substance basis for its anti-AD effect.The results showed that Aβ_1-40 was down-regulated 14.8%,and Aβ_1-42 decreased 42.7%after 10μM CN treatment in SH-SY5Y/APPswe cells.During 10μM CN treatment,ADAM10 expression was elevated1.23-fold,BACE1 expression was decreased 2.3-fold,PS1 expression was downregulated 1.66-fold,s APPαlevel was increased 2.61-fold,s APPβlevel was decreased 4.35-fold,and BACE1 enzyme activity was decreased 1.15-fold,indicating that CN inhibited BACE1 activity and shift APP metabolic pathway to non-amyloid metabolic pathway.In addition,CN improved the oxidative damage induced by AβO.The results indicated that the level of Ca²⁺was down-regulated 32.8%,ATP was up-regulated 27.0%,ROS and MMP were improved,and the apoptotic cells ratio decreased6.5%after 20μM CN treatment.It has been shown that CN could significantly protect AβO-induced abnormal Ca²⁺metabolism,as well as mitochondrial metabolic disorders.In summary,the work deeply discussed the anti-AD activity of T.chebulae,and its component CN.The results showed T.Chebula and CN could effectively protect the abnormal Ca²⁺metabolism and mitochondrial dysfunction induced by AβO,and inhibit Aβlevel by blocking generation process and increasing degradation pathway.It provides data for the development of anti-AD drugs and the in-depth exploration of the modern medicinal value of ethnic medicines.