Study On The Effect And Mechanism Of Yulangsan Polysccharide In Injured PC12 Cell Induced By Beta Amyloid And Dexamethasone

Objectives:This study evaluates the intervention effect of Yulangsan polysaccharide (YLSPS) against vitro Alzheimer’s disease model (PC 12 cells); observed the vitality damage effect of PC 12 cells that induced by combined amyloid peptide 25-35(A(325-35) and dexamethasone(Dexamethasone, DXM); to explore its possible mechanisms and provide experimental reference for further development of Yulangsan polysaccharide in treatment of nervous system-related diseases.Methods:1.Research the vitality damage effect of PC 12 cells that induced by combined AP25-35 and DXM; depending on different intervention factors, the experimental groups were divided into four groups, (1) control group (without any drug-induced);(2) AP25-35 drug-induced group; (3) DXM-induced group; (4) combined drug-induced group (Aβ25-35 combined with DXM). Neuronal PC12 cells were cultured in vitro, cell density of 85% were cultured and given drug-induced according to above, and for 24 hours, were detected from the following aspects:(1)observed cell morphology of each group change in inverted phase contrast microscope; (2) using CCK-8 assay to detect the change in cell proliferation;(3) changes of cell membrane damage in each group were detected by LDH level; (4)by using flow cytometry to AnnexinV-HTC/PI double staining, detected early and late apoptotic cells change rate.2.Research the possible mechanisms of intervention effects of Yulangsan polysaccharide on neuronal PC 12 cells intended disease model that induced by combination drugs; culture the cells to 80% density, and according to the above group, given drug-induced and drug-intervention. Give each inducing drugs and intervention drugs (Yulangsan Polysaccharides) effect of before and after 24 hours, and then using quantitative PCR, immunohistochemistry and Western Blot method to detect the expression of APP, Bax, caspase-3 and Bcl-2 in mRNA expression and protein levels expression.Results:1. These two drugs can induce toxic effects on PC 12 cells, within a certain range of concentrations, toxic effect directly dependent on dose and time. On this basis, we select Aβ25.35 10μM (24h) and DXM (24h) as the next test model conditions. Injury model that established by the above, the results of LDH in cells medium and CCK-8 were basically the same, and Yulangsan Polysaccharide were dose-dependently decrease of LDH leakage of the cells; from the results of flow cytometry showed that YLSPS can reduce model apoptotic death.2. Real-time PCR results showed that compared with the model group, Yulangsan polysaccharide high-dose group can reduce APP, Bax, caspase-3 mRNA expression levels (P<0.05), and improve the Bcl-2 mRNA expression (P<0.05); Yulangsan polysaccharide low dose group can reduce Bax, caspase-3 mRNA levels (P<0.05).3. Immunohistochemistry results showed that, compared with the model group group, Yulangsan polysaccharide high dose groups can decrease varying degrees of APP, Bax and caspase-3,and increase the expression of Bcl-2 protein (P<0.05);western blot results showed that Yulangsan polysaccharide each dose group can reduce Bax protein expression (P<0.05) and increase Bcl-2 protein expression (P<0.05).Conclusions:1. PC 12 cells that induced by Aβ and DXM PC 12, can inhibit the cell proliferation, presents apoptosis changes.2. Yulangsan polysaccharide may antagonize the apoptosis of PC 12 cells that induced by Aβ25-35 and DXM, reversing the expression of these genes, which play a role in anti-apoptosis.