State Key Laboratory of Bioactive Substances and Functions of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100050, China.
Beijing Key Laboratory of Drug Targets Identification and Drug Screening, National Center for Pharmaceutical Screening, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100050, China.
Acta Pharmacol Sin. 2024 Mar;45(3):480-489. doi: 10.1038/s41401-023-01187-3. Epub 2023 Nov 22.
Dopaminergic neurons in the substantia nigra (SN) expressing SUR1/Kir6.2 type ATP-sensitive potassium channels (K-ATP) are more vulnerable to rotenone or metabolic stress, which may be an important reason for the selective degeneration of neurons in Parkinson's disease (PD). Baicalein has shown neuroprotective effects in PD animal models. In this study, we investigated the effect of baicalein on K-ATP channels and the underlying mechanisms in rotenone-induced apoptosis of SH-SY5Y cells. K-ATP currents were recorded from SH-SY5Y cells using whole-cell voltage-clamp recording. Drugs dissolved in the external solution at the final concentration were directly pipetted onto the cells. We showed that rotenone and baicalein opened K-ATP channels and increased the current amplitudes with EC values of 0.438 μM and 6.159 μM, respectively. K-ATP channel blockers glibenclamide (50 μM) or 5-hydroxydecanoate (5-HD, 250 μM) attenuated the protective effects of baicalein in reducing reactive oxygen species (ROS) content and increasing mitochondrial membrane potential and ATP levels in rotenone-injured SH-SY5Y cells, suggesting that baicalein protected against the apoptosis of SH-SY5Y cells by regulating the effect of rotenone on opening K-ATP channels. Administration of baicalein (150, 300 mg·kg·d, i.g.) significantly inhibited rotenone-induced overexpression of SUR1 in SN and striatum of rats. We conducted surface plasmon resonance assay and molecular docking, and found that baicalein had a higher affinity with SUR1 protein (K = 10.39 μM) than glibenclamide (K = 24.32 μM), thus reducing the sensitivity of K-ATP channels to rotenone. Knockdown of SUR1 subunit reduced rotenone-induced apoptosis and damage of SH-SY5Y cells, confirming that SUR1 was an important target for slowing dopaminergic neuronal degeneration in PD. Taken together, we demonstrate for the first time that baicalein attenuates rotenone-induced SH-SY5Y cell apoptosis through binding to SUR1 and activating K-ATP channels.
在黑质（SN）中表达 SUR1/Kir6.2 型三磷酸腺苷敏感钾通道（K-ATP）的多巴胺能神经元对鱼藤酮或代谢应激更敏感，这可能是帕金森病（PD）中神经元选择性退化的一个重要原因。黄芩素在 PD 动物模型中显示出神经保护作用。在这项研究中，我们研究了黄芩素对鱼藤酮诱导的 SH-SY5Y 细胞凋亡中 K-ATP 通道的影响及其潜在机制。使用全细胞膜片钳记录技术从 SH-SY5Y 细胞中记录 K-ATP 电流。药物在终浓度下溶解在外部溶液中，并直接吸入细胞中。我们表明，鱼藤酮和黄芩素均能打开 K-ATP 通道，并分别以 EC 值为 0.438μM 和 6.159μM 增加电流幅度。K-ATP 通道阻滞剂格列本脲（50μM）或 5-羟癸酸（5-HD，250μM）可减弱黄芩素降低鱼藤酮损伤 SH-SY5Y 细胞内活性氧（ROS）含量、增加线粒体膜电位和 ATP 水平的保护作用，表明黄芩素通过调节鱼藤酮对 K-ATP 通道开放的影响来防止 SH-SY5Y 细胞凋亡。黄芩素（150、300mg·kg·d，ig）给药可显著抑制鱼藤酮诱导的大鼠 SN 和纹状体中 SUR1 的过度表达。我们进行了表面等离子体共振分析和分子对接，发现黄芩素与 SUR1 蛋白的亲和力（K=10.39μM）高于格列本脲（K=24.32μM），从而降低了 K-ATP 通道对鱼藤酮的敏感性。SUR1 亚基敲低可减少鱼藤酮诱导的 SH-SY5Y 细胞凋亡和损伤，证实 SUR1 是减缓 PD 中多巴胺能神经元变性的重要靶标。综上所述，我们首次证明黄芩素通过与 SUR1 结合并激活 K-ATP 通道来减轻鱼藤酮诱导的 SH-SY5Y 细胞凋亡。
