School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing 102488, PR China.
Chinese Herbal Medicine Industry Development Center of Ningqiang County, Hanzhong 724400, PR China.
Phytomedicine. 2024 Dec;135:156229. doi: 10.1016/j.phymed.2024.156229. Epub 2024 Nov 10.
Studies have demonstrated the potential of gastrodin in the treatment of Alzheimer's disease (AD), however, its mechanism of action remains elusive. Currently, the Amyloid-β (Aβ) cascade hypothesis continues to be the prevailing theory regarding AD etiology. The ubiquitination of P-glycoprotein (P-gp) at the blood-brain barrier (BBB) contributes to the accumulation of Aβ in the brain during AD.
To investigate the mechanism of gastrodin intervention in AD.
The molecular docking, molecular dynamics simulations, and microscale thermophoresis (MST) were employed to identify the action target of gastrodin. The western blot (WB) was performed to detect the protein expression level, the ubiquitination level of P-gp was determined using co-immunoprecipitation (CO-IP) assay. P-gp transport activity was detected using an NBD-CSA fluorescence assay. Trans-Epithelial Electrical Resistance (TEER) was used to detect cell resistance. Fluorescein-labeled dextran experiments were performed to determine the individual cell permeability. The immunofluorescence (IF) was employed to detect Aβ deposition, the Morris Water Maze test was used to assess behavioral changes in APP/PS1 mice and the levels of Aβ40 and Aβ42 expression were quantified using enzyme-linked immunosorbent assay.
The FBXO15 was the target of gastrodin-mediated inhibition of P-gp ubiquitination. Gastrodin increased the P-gp expression, cell resistance, and P-gp transport activity of BEND.3 cells upon treatment with Aβ40 through mechanisms involving the reduction of FBXO15 and P-gp binding and the inhibition of P-gp ubiquitination. And gastrodin could effectively improve memory function and increase number of neurons in APP/PS1 mice, reduce the accumulation of Aβ40 and Aβ42, and enhance P-gp expression in a dose-dependent manner.
Aβ40 induces the ubiquitination and proteasomal degradation of BBB P-gp, however, gastrodin inhibits the ubiquitination of P-gp by binding to FBXO15, thereby increasing P-gp protein expression and enhancing its transport function.
研究表明天麻素在治疗阿尔茨海默病（AD）方面具有潜力，但其作用机制仍不清楚。目前，β淀粉样蛋白（Aβ）级联假说仍然是 AD 病因的主流理论。血脑屏障（BBB）上 P-糖蛋白（P-gp）的泛素化导致 AD 时大脑中 Aβ的积累。
研究天麻素干预 AD 的机制。
采用分子对接、分子动力学模拟和微量热泳动（MST）技术鉴定天麻素的作用靶点。采用 Western blot（WB）检测蛋白表达水平，采用免疫共沉淀（CO-IP）检测 P-gp 的泛素化水平。采用 NBD-CSA 荧光法检测 P-gp 转运活性。采用跨上皮电阻（TEER）检测细胞电阻。采用荧光素标记的葡聚糖实验测定细胞通透性。采用免疫荧光（IF）检测 Aβ沉积，采用 Morris 水迷宫试验评估 APP/PS1 小鼠的行为变化，采用酶联免疫吸附试验（ELISA）定量检测 Aβ40 和 Aβ42 的表达水平。
FBXO15 是天麻素介导的 P-gp 泛素化抑制的作用靶点。天麻素通过降低 FBXO15 与 P-gp 的结合以及抑制 P-gp 泛素化，增加 BEND.3 细胞 Aβ40 处理时的 P-gp 表达、细胞电阻和 P-gp 转运活性。天麻素能有效改善 APP/PS1 小鼠的记忆功能，增加神经元数量，减少 Aβ40 和 Aβ42 的积累，并呈剂量依赖性增加 P-gp 表达。
Aβ40 诱导 BBB P-gp 的泛素化和蛋白酶体降解，而天麻素通过与 FBXO15 结合抑制 P-gp 的泛素化，从而增加 P-gp 蛋白表达，增强其转运功能。
