| 引用本文: |
谭惠中, 刘竹暄, 唐洁, 谢乐, 张秀丽, 朱新华, 伍大华.丹参多酚酸B通过NLRP3/Caspase-1/GSDMD通路抑制OGD诱导海马神经元焦亡的分子机制研究[J].湖南中医药大学学报,2025,45(4):638-646[点击复制] |
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| 丹参多酚酸B通过NLRP3/Caspase-1/GSDMD通路抑制OGD诱导海马神经元焦亡的分子机制研究 |
| 谭惠中,刘竹暄,唐洁,谢乐,张秀丽,朱新华,伍大华 |
| (湖南中医药大学, 湖南 长沙 410208;湖南省中西医结合医院, 湖南 长沙 410006;湖南中医药大学第一附属医院, 湖南 长沙 410007) |
| 摘要: |
| 目的 通过网络药理学和实验验证，研究了丹参多酚酸B（Sal B）治疗VD的潜在分子机制。方法 采用网络药理学研究Sal B干预VD的潜在靶点，构建蛋白质-蛋白质相互作用（PPI）网络及GO和KEGG通路富集分析获得相关靶点通路，分子对接评估Sal B和靶点的相互作用力。CCK-8法筛选Sal B给药浓度，显微镜观察细胞形态，酶标仪检测细胞上清中乳酸脱氢酶（LDH）释放率，ELISA法测炎症因子，免疫荧光检测NOD样受体热蛋白结构域相关蛋白3（NLRP3）蛋白表达，Western blot检测NLRP3、凋亡相关蛋白样蛋白（ASC）、半胱氨酸蛋白酶-1（Caspase-1）和消皮素D（GSDMD）蛋白表达水平。结果 生物信息学分析获得Sal B干预VD靶点共37个，网络药理学研究筛选到Caspase-1、NLRP3、TNFR1等核心靶基因，并通过KEGG和GO富集到NOD样受体信号通路（NLRs）等可能涉及这些靶基因。体外实验采用经典的NLRP3/Caspase-1/GSDMD焦亡途径进行。建立OGD-诱导海马神经元焦亡模型，发现Sal B能显著提高OGD后HT22细胞存活率（P＜0.05，P＜0.01），减轻细胞损伤，减少LDH释放量（P＜0.05，P＜0.01），降低OGD损伤HT22细胞IL-1β和IL-18水平（P＜0.01），抑制NLRP3炎症小体激活，并降低了NLRP3、GSDMD-N、cleaved-Caspase-1、ASC蛋白表达水平（P＜0.05，P＜0.01）。结论 Sal B可能通过NLRP3/Caspase-1/GSDMD通路抑制OGD诱导的海马神经元细胞焦亡，为Sal B防治VD提供了实验依据。 |
| 关键词: 血管性痴呆 丹参多酚酸B 氧糖剥夺 细胞焦亡 NLRP3/Caspase-1/GSDMD信号通路 网络药理学 |
| DOI：10.3969/j.issn.1674-070X.2025.04.008 |
| 投稿时间：2024-12-22 |
| 基金项目:国家自然科学基金项目（82374441）；湖南省自然科学基金项目（2024JJ5317）；湖南省卫生健康委卫生科研课题（W20243137）；湖南省研究生科研创新项目（CX20230809）。 |
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| Molecular mechanism of salvianolic acid B inhibiting OGD-induced pyroptosis of hippocampal neurons via NLRP3/Caspase-1/GSDMD pathway |
| TAN Huizhong, LIU Zhuxuan, TANG Jie, XIE Le, ZHANG Xiuli, ZHU Xinhua, WU Dahua |
| (Hunan University of Chinese Medicine, Changsha, Hunan 410208, China;Hunan Hospital of Integrated Traditional Chinese and Western Medicine, Changsha, Hunan 410006, China;The First Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410007, China) |
| Abstract: |
| Objective To explore the potential molecular mechanisms of salvianolic acid B (Sal B) in treating vascular dementia (VD) through network pharmacology and experimental verification. Methods Network pharmacology was used to identify the potential targets of Sal B intervention in VD. A protein-protein interaction (PPI) network was constructed, followed by GO and KEGG pathway enrichment analyses to determine biologically relevant target pathways. Molecular docking was performed to evaluate the interaction between Sal B and its targets. The optimal concentration of Sal B was determined via CCK-8 assay, the cell morphology was observed under a microscope, and the release rate of lactate dehydrogenase (LDH) in the cell supernatant was measured using a microplate reader. The levels of inflammatory factors were measured by ELISA, the protein expression of NLRP3 was examined by immunofluorescence, and the protein expression levels of NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC), cysteinyl aspartate specificproteinase 1 (Caspase-1), and GSDMD were checked by Western blot. Results A total of 37 targets for Sal B intervention in VD were obtained by bioinformatics analysis. Core target genes such as Caspase-1, NLRP3, and TNFR1 were screened by network pharmacology, and the pathways including the NOD-like receptors (NLRs) signaling pathway that may involve these target genes were identified through KEGG and GO analyses. In vitro experiments were performed using the classic NLRP3/Caspase-1/GSDMD pyroptosis pathway. The OGD-induced pyroptosis model of hippocampal neurons was established, and it was found that Sal B could significantly enhance the survival rate of HT22 cells after OGD (P＜0.05, P＜0.01), alleviate cell damage, reduce LDH release (P＜0.05 or P＜0.01), decrease the levels of IL-1β and IL-18 in OGD-damaged HT22 cells (P＜0.01), inhibit the activation of NLRP3 inflammasome, and reduce the protein expression levels of NLRP3, GSDMD-N, cleaved Caspase-1, and ASC (P＜0.05, P＜0.01). Conclusion Sal B may inhibit OGD-induced pyroptosis of hippocampal neurons through NLRP3/Caspase-1/GSDMD pathway, providing experimental evidence for preventing and treating VD with Sal B. |
| Key words: vascular dementia salvianolic acid B oxygen-glucose deprivation pyroptosis NLRP3/Caspase-1/GSDMD signaling pathway network pharmacology |
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