从印楝花中分离得到的豆甾醇通过下调 HT-22 细胞中的 Cdk5/p35/p25 信号通路来减轻谷氨酸诱导的神经毒性。
Stigmasterol isolated from Azadirachta indica flowers attenuated glutamate-induced neurotoxicity via downregulation of the Cdk5/p35/p25 signaling pathway in the HT-22 cells.
机构信息
Department of Clinical Chemistry, Faculty of Allied Health Sciences, Program in Clinical Biochemistry and Molecular Medicine, Chulalongkorn University, Bangkok 10330, Thailand; Laboratory of Signal Transduction, Research Institute for Veterinary Science, College of Veterinary Medicine, Seoul National University, Seoul 08826, South Korea.
Department of Clinical Chemistry, Faculty of Allied Health Sciences, Program in Clinical Biochemistry and Molecular Medicine, Chulalongkorn University, Bangkok 10330, Thailand.
出版信息
Phytomedicine. 2023 May;113:154728. doi: 10.1016/j.phymed.2023.154728. Epub 2023 Feb 24.
BACKGROUND
Glutamate, an excitatory neurotransmitter, was elevated in the brain of neurodegenerative disease (ND) patients. The excessive glutamate induces Ca influx and reactive oxygen species (ROS) production which exacerbates mitochondrial function, leading to mitophagy aberration, and hyperactivates Cdk5/p35/p25 signaling leading to neurotoxicity in ND. Stigmasterol, a phytosterol, has been reported for its neuroprotective effects; however, the underlying mechanism of stigmasterol on restoring glutamate-induced neurotoxicity is not fully investigated.
PURPOSE
We investigated the effect of stigmasterol, a compound isolated from Azadirachta indica (AI) flowers, on ameliorating glutamate-induced neuronal apoptosis in the HT-22 cells.
STUDY DESIGN
To further understand the underlying molecular mechanisms of stigmasterol, we investigated the effect of stigmasterol on Cdk5 expression, which was aberrantly expressed in glutamate-treated cells. Cell viability, Western blot analysis, and immunofluorescence are employed.
RESULTS
Stigmasterol significantly inhibited glutamate-induced neuronal cell death via attenuating ROS production, recovering mitochondrial membrane depolarization, and ameliorating mitophagy aberration by decreasing mitochondria/lysosome fusion and the ratio of LC3-II/LC3-I. In addition, stigmasterol treatment downregulated glutamate-induced Cdk5, p35, and p25 expression via enhancement of Cdk5 degradation and Akt phosphorylation. Although stigmasterol demonstrated neuroprotective effects on inhibiting glutamate-induced neurotoxicity, the efficiency of stigmasterol is limited due to its poor water solubility. We conjugated stigmasterol to soluble soybean polysaccharides with chitosan nanoparticles to overcome the limitations. We found that the encapsulated stigmasterol increased water solubility and enhanced the protective effect on attenuating the Cdk5/p35/p25 signaling pathway compared with free stigmasterol.
CONCLUSION
Our findings illustrate the neuroprotective effect and the improved utility of stigmasterol in inhibiting glutamate-induced neurotoxicity.
背景
谷氨酸是一种兴奋性神经递质，在神经退行性疾病（ND）患者的大脑中升高。过量的谷氨酸会诱导 Ca 内流和活性氧（ROS）的产生，从而加剧线粒体功能障碍，导致自噬异常，并过度激活 Cdk5/p35/p25 信号通路，导致 ND 中的神经毒性。豆甾醇是一种植物甾醇，已被报道具有神经保护作用；然而，豆甾醇恢复谷氨酸诱导的神经毒性的潜在机制尚未完全研究。
目的
我们研究了从印楝花中分离得到的化合物豆甾醇对改善 HT-22 细胞中谷氨酸诱导的神经元凋亡的作用。
研究设计
为了进一步了解豆甾醇的潜在分子机制，我们研究了豆甾醇对 Cdk5 表达的影响，Cdk5 在谷氨酸处理的细胞中异常表达。采用细胞活力测定、Western blot 分析和免疫荧光法。
结果
豆甾醇通过抑制 ROS 产生、恢复线粒体膜去极化以及通过减少线粒体/溶酶体融合和 LC3-II/LC3-I 比值来改善自噬异常，显著抑制谷氨酸诱导的神经元细胞死亡。此外，豆甾醇通过增强 Cdk5 降解和 Akt 磷酸化来下调谷氨酸诱导的 Cdk5、p35 和 p25 表达。尽管豆甾醇通过抑制谷氨酸诱导的神经毒性显示出神经保护作用，但由于其水溶性差，其效率有限。我们将豆甾醇与壳聚糖纳米粒结合可溶性大豆多糖进行偶联，以克服这些限制。我们发现，与游离豆甾醇相比，包封的豆甾醇增加了水溶解度，并增强了对抑制 Cdk5/p35/p25 信号通路的保护作用。
结论
我们的研究结果表明，豆甾醇具有神经保护作用，并提高了其抑制谷氨酸诱导的神经毒性的应用效果。
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