She Yong, Shao Chong-Yu, Liu Yuan-Feng, Huang Ying, Yang Jiehong, Wan Hai-Tong
School of Life Sciences, Zhejiang Chinese Medical University, Hangzhou, Zhejiang, China.
School of Basic Medical Sciences, Zhejiang Chinese Medical University, Hangzhou, Zhejiang, China.
Front Pharmacol. 2024 Jun 27;15:1415445. doi: 10.3389/fphar.2024.1415445. eCollection 2024.
Ischemic Stroke (IS) stands as one of the primary cerebrovascular diseases profoundly linked with inflammation. In the context of neuroinflammation, an excessive activation of microglia has been observed. Consequently, regulating microglial activation emerges as a vital target for neuroinflammation treatment. Catalpol (CAT), a natural compound known for its anti-inflammatory properties, holds promise in this regard. However, its potential to modulate neuroinflammatory responses in the brain, especially on microglial cells, requires comprehensive exploration. In our study, we investigated into the potential anti-inflammatory effects of catalpol using lipopolysaccharide (LPS)-stimulated BV2 microglial cells as an experimental model. The production of nitric oxide (NO) by LPS-activated BV2 cells was quantified using the Griess reaction. Immunofluorescence was employed to measure glial cell activation markers. RT-qPCR was utilized to assess mRNA levels of various inflammatory markers. Western blot analysis examined protein expression in LPS-activated BV2 cells. NF-κB nuclear localization was detected by immunofluorescent staining. Additionally, molecular docking and molecular dynamics simulations (MDs) were conducted to explore the binding affinity of catalpol with key targets. Catalpol effectively suppressed the production of nitric oxide (NO) induced by LPS and reduced the expression of microglial cell activation markers, including Iba-1. Furthermore, we observed that catalpol downregulated the mRNA expression of proinflammatory cytokines such as IL-6, TNF-α, and IL-1β, as well as key molecules involved in the NLRP3 inflammasome and NF-κB pathway, including NLRP3, NF-κB, caspase-1, and ASC. Our mechanistic investigations shed light on how catalpol operates against neuroinflammation. It was evident that catalpol significantly inhibited the phosphorylation of NF-κB and NLRP3 inflammasome activation, both of which serve as upstream regulators of the inflammatory cascade. Molecular docking and MDs showed strong binding interactions between catalpol and key targets such as NF-κB, NLRP3, and IL-1β. Our findings support the idea that catalpol holds the potential to alleviate neuroinflammation, and it is achieved by inhibiting the activation of NLRP3 inflammasome and NF-κB, ultimately leading to the downregulation of pro-inflammatory cytokines. Catalpol emerges as a promising candidate for the treatment of neuroinflammatory conditions.
缺血性中风（IS）是与炎症密切相关的主要脑血管疾病之一。在神经炎症的背景下，已观察到小胶质细胞过度激活。因此，调节小胶质细胞激活成为神经炎症治疗的关键靶点。梓醇（CAT）是一种以其抗炎特性而闻名的天然化合物，在这方面具有潜力。然而，其调节大脑神经炎症反应的潜力，特别是对小胶质细胞的影响，需要全面探索。在我们的研究中，我们以脂多糖（LPS）刺激的BV2小胶质细胞为实验模型，研究了梓醇的潜在抗炎作用。使用格里斯反应对LPS激活的BV2细胞产生的一氧化氮（NO）进行定量。采用免疫荧光法测量胶质细胞激活标志物。利用RT-qPCR评估各种炎症标志物的mRNA水平。蛋白质印迹分析检测LPS激活的BV2细胞中的蛋白质表达。通过免疫荧光染色检测NF-κB核定位。此外，进行了分子对接和分子动力学模拟（MD）以探索梓醇与关键靶点的结合亲和力。梓醇有效抑制了LPS诱导的一氧化氮（NO）产生，并降低了包括Iba-1在内的小胶质细胞激活标志物的表达。此外，我们观察到梓醇下调了促炎细胞因子如IL-6、TNF-α和IL-1β的mRNA表达，以及参与NLRP3炎性小体和NF-κB途径的关键分子，包括NLRP3、NF-κB、caspase-并且ASC。我们的机制研究揭示了梓醇如何对抗神经炎症。很明显，梓醇显著抑制了NF-κB的磷酸化和NLRP3炎性小体的激活，这两者都是炎症级联反应的上游调节因子。分子对接和MD显示梓醇与NF-κB、NLRP3和IL-1β等关键靶点之间有强烈的结合相互作用。我们的研究结果支持梓醇具有减轻神经炎症的潜力这一观点，这是通过抑制NLRP3炎性小体和NF-κB的激活来实现的，最终导致促炎细胞因子的下调。梓醇成为治疗神经炎症性疾病的有希望的候选药物。
