Xie Meixia, Huang Huilin, Lu Yingxin, Chen Lei, Wang Shumei, Xian Minghua
Guangdong Pharmaceutical University, Guangzhou, 510006, China; Key Laboratory of Digital Quality Evaluation of Chinese Materia Medica of State Administration of TCM, School of Chinese Materia Medica, Guangdong Pharmaceutical University, Guangzhou, 510006, China; Engineering & Technology Research Center for Chinese Materia Medica Quality of the Universities of Guangdong Province, School of Chinese Materia Medica, Guangdong Pharmaceutical University, Guangzhou, 510006, China.
Guangdong Pharmaceutical University, Guangzhou, 510006, China; Key Laboratory of Digital Quality Evaluation of Chinese Materia Medica of State Administration of TCM, School of Chinese Materia Medica, Guangdong Pharmaceutical University, Guangzhou, 510006, China; Engineering & Technology Research Center for Chinese Materia Medica Quality of the Universities of Guangdong Province, School of Chinese Materia Medica, Guangdong Pharmaceutical University, Guangzhou, 510006, China.
J Ethnopharmacol. 2025 Jan 31;340:119247. doi: 10.1016/j.jep.2024.119247. Epub 2024 Dec 17.
Ischemic stroke (IS) is a leading cause of death and disability in China. Danhong Injection (DHI) is a traditional Chinese medicine preparation made from Salvia miltiorrhiza var. miltiorrhiza and Carthamus tinctoriusL. It is used for treating stroke in China with proven safety and efficacy. Microglia M1/M2 polarization is a key factor in IS inflammatory response. However, the key transcription factors that regulate microglia polarization are unknown. It is also not clear how DHI exerts its mechanism in the treatment of IS.
This research aimed to investigate the effect of DHI on microglial polarization and neuroinflammation associated with IS and to elucidate the underlying mechanisms, with an emphasis on the JUNB/NF-κB signaling pathway.
An oxygen-glucose deprivation (OGD) damage cell model and a permanent middle cerebral artery occlusion (pMCAO) model in C57BL/6 mice were employed. Neurological deficits, cerebral infarct volume, and microglial activation were assessed. Non-targeted metabolomics analysis with UHPLC-QE-MS and molecular biology methods, including RT-qPCR and Western blot, were applied to investigate the mechanisms.
In vivo, DHI decreased inflammation, reduced brain damage, and enhanced neurological function. DHI also ameliorated microglial activation and OGD-induced apoptosis in vitro. Metabolomics analysis identified significant metabolic changes, particularly in amino acid metabolism. Additionally, DHI treatment decreased the upregulated mRNA levels of ASS1 and ASL after OGD, indicating an influence on the arginine biosynthesis pathway, which is crucial for microglial function. DHI modulated the M1 to M2 phenotypes of microglial polarization and regulated microglial polarization through the JUNB/NF-κB signaling pathway. This was confirmed by JUNB silencing experiments.
DHI exhibits neuroprotective effects via suppressing ASS1 through the JUNB/NF-κB pathway, promoting the M2 state of microglia, and lowering the expression of inflammatory cytokines. This research unveils the potential therapeutic target of JUNB for IS treatment and sheds light on the novel intervention mechanism of DHI in microglial cells.
缺血性中风（IS）是中国死亡和残疾的主要原因。丹红注射液（DHI）是一种由丹参和红花制成的中药制剂。在中国，它被用于治疗中风，安全性和有效性已得到证实。小胶质细胞M1/M2极化是IS炎症反应的关键因素。然而，调节小胶质细胞极化的关键转录因子尚不清楚。DHI在治疗IS中如何发挥其作用机制也不明确。
本研究旨在探讨DHI对与IS相关的小胶质细胞极化和神经炎症的影响，并阐明其潜在机制，重点关注JUNB/NF-κB信号通路。
采用氧糖剥夺（OGD）损伤细胞模型和C57BL/6小鼠永久性大脑中动脉闭塞（pMCAO）模型。评估神经功能缺损、脑梗死体积和小胶质细胞激活情况。应用超高效液相色谱-四极杆飞行时间质谱（UHPLC-QE-MS）进行非靶向代谢组学分析以及包括逆转录定量聚合酶链反应（RT-qPCR）和蛋白质免疫印迹法（Western blot）在内的分子生物学方法来研究其机制。
在体内，DHI减轻炎症、减少脑损伤并增强神经功能。DHI在体外也改善了小胶质细胞激活和OGD诱导的细胞凋亡。代谢组学分析确定了显著的代谢变化，尤其是在氨基酸代谢方面。此外，DHI处理降低了OGD后ASS1和ASL上调的mRNA水平，表明对精氨酸生物合成途径有影响，而精氨酸生物合成途径对小胶质细胞功能至关重要。DHI调节小胶质细胞极化的M1至M2表型，并通过JUNB/NF-κB信号通路调节小胶质细胞极化。JUNB沉默实验证实了这一点。
DHI通过JUNB/NF-κB途径抑制ASS1、促进小胶质细胞的M2状态并降低炎性细胞因子的表达，从而发挥神经保护作用。本研究揭示了JUNB作为IS治疗潜在靶点的可能性，并阐明了DHI在小胶质细胞中的新型干预机制。
