灯盏乙素介入IP3R-Ca2+途径抑制β淀粉样蛋白诱导的神经细胞凋亡

目的：观察灯盏乙素（Scu）对β淀粉样蛋白（Aβ）诱导的细胞模型中1，4，5-三磷酸肌醇受体（IP₃R）-Ca²⁺途径的影响，探讨其在阿尔茨海默病（AD）病程中可能发挥的积极作用。方法：选用神经母细胞瘤细胞分为对照组、Scu处理组、Aβ处理组、Aβ+Scu （高、中、低）处理组及Aβ+IP₃R拮抗剂组，用CCK-8法筛选药物浓度并检测各组细胞生存率；用酶联免疫吸附法检测各组细胞中1，4，5-三磷酸肌醇（IP₃）的含量；用蛋白印迹和实时荧光定量聚合酶链反应方法检测各组细胞IP₃R和凋亡相关因子Caspase-3、Bcl-2、Bax的蛋白及mRNA的表达水平；用激光共聚焦显微镜观察各组细胞内Ca²⁺浓度的变化；用AnnexinV/PI双染法测定各组细胞的凋亡率。结果：与对照组和Scu处理组相比，Aβ处理组细胞存活率下降，IP₃含量升高，IP₃R、Bax和Caspase-3的蛋白及mRNA表达上调，Bcl-2蛋白及mRNA的表达下调，细胞胞浆内Ca²⁺浓度及细胞凋亡率升高；Aβ+Scu处理组细胞中各检测指标的变化与Aβ处理组的结果正好相反，IP₃R通道下游指标的变化与Aβ+IP₃R拮抗剂组基本一致。结论：Scu能够下调通路蛋白IP₃、IP₃R的表达，抑制Aβ介导的Ca²⁺内流所致的细胞凋亡，可能通过对IP₃R-Ca²⁺途径的调控来影响AD病程。

Scutellarin inhibits β-amyloid induced neuronal apoptosis by IP3R-Ca2+ pathway

OBJECTIVE To observe the effect of Scu on the inositol 1,4,5-trisphosphate receptor (IP₃R)-Ca²⁺ pathway in the β-amyloid (Aβ)-induced cell model and explore its possible role in the pathogenesis of Alzheimer's disease (AD).METHODS The SH-SY5Y cells were divided into control group,Scu treatment group,Aβ treatment group,Aβ+Scu (High、Medium、Low) treatment group and Aβ+IP₃R antagonist group.The CCK-8 method was used to screen the drug concentration and detect the survival rate of each group.The content of 1,4,5-trisphosphoinositide (IP₃) in each group was detected by enzyme-linked immunosorbent assay.The protein and mRNA expression levels of IP₃R and apoptosis-related factors Caspase-3,Bcl-2 and Bax were detected by Western blot and Real-time quantitative polymerase chain reaction respectively.Laser confocal microscopy was applied to observe intracellular Ca²⁺ concentration changes.In addition,annexinV/PI double staining was performed to determine the apoptosis rate of each group.RESULTS Compared with the control group and the Scu treatment group,the cell survival rate of the Aβ treatment group decreased,the content of IP₃ increased,the protein and mRNA expression of the IP₃R,Bax,and Caspase-3 were up-regulated while the expression of the Bcl-2 protein and mRNA was down-regulated.Internal Ca²⁺ concentration and apoptosis rate increased;Compared with the Aβ-treated group,the changes in the detection parameters of the Aβ+Scu-treated group were opposite to those in the Aβ-treated group,and played the same role with the Aβ+IP₃R antagonist group in the downstream indicators of the IP₃R channel.and the changes of IP3R channel downstream indicators were basically consistent with those in Aβ+IP3R antagonist group.CONCLUSION Scu can down-regulate the expression of pathway protein IP₃、IP₃R and inhibit Aβ-mediated apoptosis induced by Ca²⁺ influx,which may affect the course of AD through the regulation of IP₃R-Ca²⁺ pathway.