Kang Xiaoyu, Cheng Enhao, Guo Peixian, Xi Leying, Xiang Chunjie, Xu Shuo, Zhang Yajie, Yang Yuee, Zhang Junfeng, Long Hongyan
Department of Pediatrics, Nanjing Hospital of Chinese Medicine Affiliated to Nanjing University of Chinese Medicine, Nanjing, 210022, China; Nanjing University of Chinese Medicine, Nanjing, 210023, China.
Department of Pediatrics, The First Affiliated Hospital of Henan University of Chinese Medicine, Zhengzhou, 450046, China.
Phytomedicine. 2025 Sep;145:157009. doi: 10.1016/j.phymed.2025.157009. Epub 2025 Jun 23.
BACKGROUND: Jing An decoction (JA), a traditional Chinese medicine formulation, has demonstrated notable clinical efficacy in the treatment of Tourette syndrome (TS) over the past two decades. The gut-brain axis (GBA) critically regulates neuroinflammation and neurodevelopmental processes, suggesting its potential as a therapeutic target in TS. However, the mechanisms by which JA alleviates TS via the GBA remain unclear. PURPOSE: This study investigated gut microbiota and its metabolites to explore the potential mechanisms by which JA alleviates TS, a chronic neurodevelopmental disorder. METHODS: A TS rat model was established via intraperitoneal injection of 3,3'-iminodipropionitrile (IDPN). Behavioral assessments, including stereotyped behavior, motor behavior, open-field testing, and novel object recognition, were conducted to evaluate the effect of JA. Gut microbiota was assessed by 16S rRNA sequencing, while short-chain fatty acids were quantified via GC-MS. Inflammatory cytokine levels were measured using ELISA. Gene expression related to tight junctions, key bacterial taxa, and metabolic enzymes was quantified using qPCR. Immunohistochemistry, immunofluorescence, and western blotting were employed to evaluate microglial polarization, barrier integrity, and the expression of key signaling pathway proteins. Additionally, an LPS-induced primary microglial inflammation model was used to investigate the role of butyrate in regulating microglia-mediated neuroinflammation. RESULTS: JA significantly reduced stereotypic behaviors, hyperactivity, and cognitive impairments in TS rats. It restored gut microbiota diversity by increasing the abundance of butyrate-producing bacteria-particularly the Lachnospiraceae NK4A136 group-and upregulating the expression of butyrate-metabolizing enzymes (Buk and ButCOA). Butyrate levels in the colon and striatum were elevated in JA-treated rats, correlating with reduced neuroinflammation and enhanced intestinal and blood-brain barrier integrity. JA promoted M2 microglial polarization, suppressed HDAC3 expression, and inhibited the TLR4/NF-κB pathway. In primary microglial cells, butyrate attenuated LPS-induced neuroinflammation-an effect comparable to that of TLR4/NF-κB inhibitors (TAK-242, PDTC)-but this effect was reversed by an MCT1 inhibitor and HDAC3 activator (AZD3965, ITSA-1). CONCLUSIONS: JA alleviates TS by regulating the GBA axis through butyrate-producing bacteria. Butyrate alleviates neuroinflammation by inhibiting the TLR4/HDAC3/NF-κB pathway, thereby promoting M2 microglial polarization.
背景：静安汤（JA）是一种中药配方，在过去二十年中已证明在治疗抽动秽语综合征（TS）方面具有显著的临床疗效。肠-脑轴（GBA）对神经炎症和神经发育过程起着关键调节作用，提示其可能成为TS的治疗靶点。然而，JA通过GBA轴减轻TS的机制尚不清楚。 目的：本研究调查肠道微生物群及其代谢产物，以探索JA减轻TS（一种慢性神经发育障碍）的潜在机制。 方法：通过腹腔注射3,3'-亚氨基二丙腈（IDPN）建立TS大鼠模型。进行行为评估，包括刻板行为、运动行为、旷场试验和新物体识别，以评估JA的效果。通过16S rRNA测序评估肠道微生物群，同时通过气相色谱-质谱联用（GC-MS）对短链脂肪酸进行定量。使用酶联免疫吸附测定（ELISA）测量炎症细胞因子水平。使用实时定量聚合酶链反应（qPCR）对与紧密连接、关键细菌分类群和代谢酶相关的基因表达进行定量。采用免疫组织化学、免疫荧光和蛋白质免疫印迹法评估小胶质细胞极化、屏障完整性和关键信号通路蛋白的表达。此外，使用脂多糖（LPS）诱导的原代小胶质细胞炎症模型研究丁酸盐在调节小胶质细胞介导的神经炎症中的作用。 结果：JA显著降低了TS大鼠的刻板行为、多动和认知障碍。它通过增加产丁酸盐细菌的丰度，特别是毛螺菌科NK4A136组，并上调丁酸盐代谢酶（Buk和ButCOA）的表达，恢复了肠道微生物群的多样性。JA处理的大鼠结肠和纹状体中的丁酸盐水平升高，这与神经炎症减轻以及肠道和血脑屏障完整性增强相关。JA促进M2小胶质细胞极化，抑制组蛋白去乙酰化酶3（HDAC3）表达，并抑制Toll样受体4（TLR4）/核因子κB（NF-κB）通路。在原代小胶质细胞中，丁酸盐减轻了LPS诱导的神经炎症，其效果与TLR4/NF-κB抑制剂（TAK-242、吡咯烷二硫代氨基甲酸盐（PDTC））相当，但这种效果被单羧酸转运体1（MCT1）抑制剂和HDAC3激活剂（AZD3965、ITSA-1）逆转。 结论：JA通过产丁酸盐细菌调节GBA轴减轻TS。丁酸盐通过抑制TLR4/HDAC3/NF-κB通路减轻神经炎症，从而促进M2小胶质细胞极化。
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