The Effects Of Raspberry Decoction On Apoptosis Of BEnd.3 Cells Induced By H₂O₂ And Related Clinical Application

Objective:In the clinical and experimental studies of the treatment of Alzheimer's disease(AD),the "Bu shen yi jing fang" has achieved remarkable results.Basic Studies in Earlier Stage,observing the effect of Raspberry decoction one of the important components of this recipe on the oxidative stress injury by H2O2 induced brain microvascular endothelial cells(bEnd.3 cells).Methods:1.Screening for optimal damage concentration of H2O2 to bEnd.3 cells,and establish a stable cellular oxidative stress model.2.Screening for the safety of raspberry decoction;The effects of different concentrations of Raspberry decoction on H2O2 induced bEnd.3 cell activity were checked by MTT assay.3.The expression of Bcl-2,Bax and Cytochrome-c(Cyt-c)in mitochondrial pathway,Caspase-3 and activated Cleaved caspase-3,Caspase-12 and activated Cleaved caspase-12 in endoplasmic reticulum stress pathway,and Caspase-8 and Cleaved caspase-8 in the death receptor pathway were determined by Western blot.4.It is verified that the protective effect of Raspberry decoction on H2O2 induced bEnd.3 cells apoptosis by Hoechst 33258.Results:1.The bEnd.3 cells were treated with H2O2 of 50-400umol/ml concentrations for 6h,and found that the viability rate of cells was 52.7±6.81%with 300 umol/ml H2O2,and the difference was statistically significant with control(P<0.05).2.There was no significant difference between the cell viability with 0.1-5.0 mg/ml concentration Raspberry decoction(P>0.05)3.The cells was preprotected with a safe Raspberry decoction,and were induced were treated with of 300umol/ml concentrations for 6h.The cell survival rate of H2O2 model group was 49.9± 1.14%,The cells were protectied with the concentration of Raspberry decoction.There were significant differences between the model group and the control,the drug treatment group and the model.(P<0.05).Acorrding to the viability rate,the concentration of 0.1.0.5.5.0 mg/ml Raspberry decoction were used to the experimental.4.The expression of apoptosis-pathways protein by Western blot showed the radio of Cyt-c/?-actin,Cleaved caspase-3/Caspase-3,.Cleaved caspase-8/Caspase-8,Cleaved caspase-12/Caspase-12 was decreased and Bc12/Bax was increased after protreated by Raspberr y decoction.Comparing with model group,the difference was statistically significant(P<0.05).5.Hoechst33258 staining showed that there were mang nuclei consolidation and apo-ptosis bodies in the model H2O2 group,which showed strong fluorescence staining.The protective effect of Raspberry decoction can decreased the apoptosis bodies and fluoresce nce and were correlative with concentration.Conclusion:The oxidative stress damage model was constructed by H2O2 induced bEnd.3 cells in the experiment.The Raspberry decoction can protect bEnd.3 cells from H2O2 induced oxidative stress injury.This mechanism may be related to the induction of apoptosis by regulating the expression of related apoptotic proteins.It provides a new direction for the prevention of AD.