SNP-9对Aβ_(25-35)导致bEnd.3细胞损伤的作用及其机制

Effects and mechanisms of SNP-9 on Aβ_(25-35)-induced damage in bEnd.3 cells

为了研究来源于丝素蛋白水解物的神经保护活性多肽SNP-9对于阿尔茨海默病(Alzheimer′s disease,AD)中血脑屏障损伤的作用,使用Aβ_(25-35)损伤脑微血管内皮细胞bEnd.3建立AD损伤模型,并给药干预。通过MTT实验检测SNP-9和Aβ_(25-35)对细胞活力的影响;RT-qPCR法检测SNP-9和Aβ_(25-35)对细胞紧密连接(tightjunctions,TJs)相关的ZO-1、occludin和claudin-5 mRNA水平的影响;Westernblot检测SNP-9和Aβ_(25-35)对细胞TNF-α、磷酸化NF-κB、NF-κB、IκBα和RAGE蛋白水平的影响。实验结果显示,SNP-9给药减少了Aβ_(25-35)诱导的bEnd.3细胞损伤,改善了模型细胞中ZO-1、occludin和claudin-5 mRNA水平的异常情况,缓解了Aβ_(25-35)导致的TNF-α、磷酸化NF-κB、IκBα和RAGE蛋白水平异常。研究结果表明,SNP-9可能通过影响RAGE/NF-κB通路,调控模型细胞炎症因子TNF-α水平,改善TJs相关指标异常,缓解Aβ_(25-35)诱导的bEnd.3细胞损伤。

In order to investigate the effects of neuroprotective peptide SNP-9 which is derived from silk fibroin hydrolysate on the injury of the blood-brain barrier in Alzheimer′s disease(AD),Aβ_(25-35) was used to damage brain microvascular endothelial cells bEnd.3 to establish AD injury model and drug intervention was performed.MTT assay was used to detect the effects of SNP-9 and Aβ_(25-35) on cell viability.RT-qPCR was used to determine the effects of SNP-9 and Aβ_(25-35) on the mRNA levels of tight junctions(TJs)-related ZO-1,occludin and claudin-5.Western blot was used to detect the effects of SNP-9 and Aβ_(25-35) on the protein levels of TNF-α,phosphorylated NF-κB,NF-κB,IκBαand RAGE.The results showed that SNP-9 reduced bEnd.3 cell damage induced by Aβ_(25-35),and improved the abnormal mRNA levels of ZO-1,occludin and claudin-5 in model cells.It alleviated the abnormal protein levels of TNF-α,phosphorylated NF-κB,IκBαand RAGE induced by Aβ_(25-35).These results suggest that SNP-9 may regulate the levels of TNF-αin model cells by influencing RAGE/NF-κB pathway,and then ameliorate TJs-related abnormalities and alleviate bEnd.3 cell injury induced by Aβ_(25-35).