| Research Background:With the advent of aging around the world,aging-related illness are paid more and more attention.Alzheimer's disease(AD)is a neurodegenerative disease that is closely related to aging.It has a high incidence after 65 years old and it is mainly characterized by the progressive deterioration of cognitive function.The main pathological features are neuronal loss,senile plaques and neurofibrillary tangles.The pathogenesis of this disease has not yet been fully elucidated.Researchers have proposed the beta-amyloid cascade hypothesis,tau protein hypothesis and angiogenic hypothesis and so on.Among them beta-amyloid cascade hypothesis is that the deposition of senile plaques containing beta-amyloid protein,is a key pathological product,which can cause brain damage in patients and can cause a series of neuron lesions.The study showed that beta-amyloid can bind to multiple receptors on the surface of the cell membrane and cause abnormal activation of intracellular signaling pathways,and it can also insert into the membrane to form beta-amyloid type ion channels,destroy the original ionic homeostasis,damage mitochondrial function,increase reactive oxygen species,lipid peroxidation,and other cell injury,and accelerate neuron apoptosis.These lesions,in return,can further promote the production and deposition of beta-amyloid protein in the body,cause a cascade damage effect,and aggravate learning and memory impairment.Beta-amyloid production is closely related to the aging,and the brain aging animal model induced by D-galactose is commonly used for aging study at home and abroad.D-galactose intraperitoneal injection for a short period of time can cause,energy metabolism disorders,decrease immune function,increas beta-amyloid protein content in the brain,decrease the brain-derived neurotrophic factor(BDNF)expression,and cause the learning and memory impairment similar to Alzheimer's disease.Delaying senility and alleviating beta-amyloid protein production,deposition and toxicity are the hot issue in the durg research for AD.Polygala Radix is a traditional Chinese herba which can restore normal coordination between heart and kidney,tranquilize the mind,and eliminate phlegm and resuscitation.Resent years,oligosaccharide esters derived from Polygala Radix are paid attention by pharmacologists.Modern pharmacological studies have showed that oligosaccharide esters of Polygala Radix have many pharmacological effects such as antioxidation,anti-inflammation,antidepressionand neuroprotection.Previous studies in our lab found that oligosaccharide esters of Polygala Radix can improve the learning and memory impairment of mice induced by scopolamine and enhance the synaptic plasticity in the hippocampus of APP/PS1 transgenic mice,indicating that it has the potential possibilities for AD treatment.Based on the above research background,in this study,we investigated the effect of oligosaccharide esters of Polygala Radix on the A?25-35-induced SH-SY5Y cell damage and D-galactose-induced learning and memroy impairment and the relative mechanisms for the further studying its anti-dementia and neuroprotective effects.Part 1 Effects of oligosaccharide esters of Polygala Radix on SH-SY5Y cell injury induced by A?25-35 and related mechanisms.Objective:This study is to investigate the effects of oligosaccharide esters of Polygala Radix on SH-SY5Y cells injury induced by A?25-35 and relative mechanisms Methods:Cultured SH-SY5Y cells were pretreated with 50?g·mL-1,100?g·mL-1,200 ?g·mL-1,and 400 ?g·mL-1 of oligosaccharide esters of Polygala Radix for 2 h,then was incubated with 25?mol·L-1 beta-amyloid peptide 25-35 for 24h.Cell viability was measured by MTT assay;Intracellular superoxide dehydrogenase activity and malondialdehyde content,and Lactate dehydrogenase level in extracellular were measured by biochemical assay;andcell apoptosis ratio was detected by flow cytometry.the expression of Bcl-2,Bax,p-CREB/CREB,p-AKT/AKT and BDNF were determined by Western Blotting method.Results:The results showed that oligosaccharide esters of Polygala Radix could significantly increase the survival rate of SH-SY5Y cells damaged by beta-amyloid peptide 25-35,reduce the apoptosis ratio,reduce the release of LDH,increase the activity of intracellular SOD,and reduce MDA.Content,increase expression of anti-apoptotic protein Bcl-2,decrease expression of apoptotic protein Bax,and promote the phosphorylation levels of AKT,CREB and BDNF in the cells..Conclusion:Oligosaccharide esters of Polygala Radix can against the injury of SH-SY5Y cells induced by beta-amyloid peptide 25-35,which is reated with its anti-oxidation,up-regulation of AKT/CREB/BDNF and anti-apoptosis.Part 2 Effects of Oligosaccharide esters of Polygala Radix on the impairment of learning and memory induced by D-galactose in mice and its related mechanisms.Objective:This study is to investigate the effects of oligosaccharide esters of Polygala Radix on D-galactose induced cognitive deficts in mice and its relative mechanisms.Methods:72 male 8-week-old Kunming mice were randomly divided into normal control group,D-galactose group,Donepezil hydrochloride group(1.67mg·kg-1·d-1)and oligosaccharide esters of Polygala Radix groups(28.67 mg·kg-1·d-1,57.33 mg·kg-1·d-1 and 114.66 mg·kg-1·d-1.Except for the control mice,all the mice in the other group were continuous intraperitoneally injectedwith D-galactose120 mg·kg-1·d-1 and intragastric administered the dr?gs for 60 days.Normal control mice were given equal amount of vehicle.The autonomous activities of mice were detected by open field test;learning and memory ability was determined by step-down test and Morris water maze test;the superoxide dismutase activity and malondialdehyde content in serum were tested by biochemical assays;the expression of BDNF in cerebral cortex was observed by immunohistochemical analysis;the changes of p-CREB/CREB,p-AKT/AKT and BDNF in mouse cerebral cortex were detected by Western blotting method.Results:The results of open-field experiment showed that there are no significant change between the several groups.Midium dosage of With the prolongation of training time,oligosaccharide esters of Polygala Radix(57.33 mg·kg-1·d-1)could significantly shorten the latency to find the platform,decrease the latency of the first platform-passing,and increase the number of platform-passing in probe trial in mice induced by D-galactose.Donepezil hydrochloride and oligosaccharide esters of Polygala Radix(28.67 mg·kg-1·d-1,114.66 mg·kg-1·d-1)have no significant influence on the latency of first platform-passing and the number of platform-passing compared with D-galactose-induced aging mice.The step-down experiment results showed that the oligosaccharide esters of Polygala Radix(57.33 mg·kg-1·d-1,114.66 mg·kg-1·d-1)and Donepezil hydrochloride can significantly reduce the number of errors in D-galactose-induced aging mice.Oligosaccharide esters of Polygala Radix(57..33 mg·kg-1·d-1)can significantly shorten the latency of first-step-down in mice induced by D-galactose.Oligosaccharide esters of Polygala Radix(28.67 mg·kg-1·d-1,57.33 mg·kg-1·d-1,114.66 mg·kg-1·d-1)significantly increased the activity of SOD and reduced the MDA content in the ser?M of mice induced by D-galactose.Donepezil hydrochloride has no significant influence on SOD activity but significantly reduce MDA content in serum of D-galactose-induced aging mice.Oligosaccharide esters of Polygala Radix also significantly increased the expression of BDNF,the phosphorylation of CREB and AKT in the cortex of mice induced by D-galactose.However,there was no significant difference in the expression of beta amyloid protein in mouse cerebral cortex between groups.Conclusion:Oligosaccharide esters of Polygala Radix can significantly improve the learning and memory ability of D-galactose-induced aging mice.This improvement maybe relate to its anti-oxidation,up-regulation of AKT/CREB/BDNF signal pathway. |
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