The Eighth School of Clinical Medicine (Foshan Hospital of Traditional Chinese Medicine), Guangzhou University of Chinese Medicine, Foshan, 528000, China.
Basic Medical College Guangzhou University of Chinese Medicine, Guangzhou, 510000, China.
J Ethnopharmacol. 2023 Aug 10;312:116505. doi: 10.1016/j.jep.2023.116505. Epub 2023 Apr 18.
Dachaihu decoction (DCH), a classic formula for Yangming and Shaoyang Syndrome Complex recorded in "Treatise on Cold Damage", has been widely used in treating intestinal disorders and inflammatory diseases with few side effects in China. However, the mechanism of DCH on septic intestinal injury (SII) remains to be explored.
This study aimed to clarify the mechanism of DCH on SII.
SII model of rat, established by cecal ligation and puncture (CLP), was used to study the effect of DCH on SII. 24 h mortality was recorded. Histological changes were observed by H&E staining. The expression of tight junction protein ZO-1 (ZO-1) and mucin2 (MUC2) was determined by immunohistochemical analysis. Secretory IgA (sIgA), diamine oxidase (DAO) and intestinal fatty acid binding protein (iFABP) were determined by enzyme-linked immunosorbent assay (ELISA). IL-1β, IL-6 and TNF-α were measured by ELISA and quantitative Real-time PCR (RT-qPCR). The gut microbiota was analyzed by 16S rRNA sequencing. The potential targets and pathways of DCH in treating SII were analyzed by integrative analysis of transcriptomic and metabolomic methods. Total glutathione (T-GSH), GSH, GSSG (reduced form of GSH), GSH peroxidase (GPX), superoxide dismutase (SOD), malonaldehyde (MDA) and indicators of hepatic and renal function were measured by biochemical kits.
Medium dose of DCH improved 24 h mortality of SII rats, reduced the pathological changes of ileum, and increased the expression levels of ZO-1, MUC2 and sIgA. DCH decreased DAO, iFABP of serum and IL-1β, IL-6, TNF-α of ileum. DCH improved α- and β-diversity and modulated the structure of gut microbiota, with Escherichia_Shigella decreased and Bacteroides and Ruminococcus increased. GSH metabolism was identified as the key pathway of DCH on SII by integrative analysis of transcriptome and metabolome. GSH/GSSG and the most common indicators of oxidative stress, were validated. Antioxidative T-GSH, GSH, GPX and SOD were increased, while MDA, the mark of lipid peroxidation was downregulated by DCH. Eventually, DCH was proved to be safe and hepato- and nephro-protective.
DCH ameliorated septic intestinal injury possibly by modulating the gut microbiota and enhancing glutathione metabolism of SII rats, without hepatotoxicity and nephrotoxicity.
ETHNOPHARMACOLOGICAL 相关性：大柴胡汤（DCH）是《伤寒论》中记载的阳明少阳综合征的经典方剂，在中国已广泛用于治疗肠道疾病和炎症性疾病，副作用较少。然而，DCH 对脓毒性肠损伤（SII）的作用机制仍有待探索。
本研究旨在阐明 DCH 对 SII 的作用机制。
采用盲肠结扎穿孔（CLP）建立大鼠 SII 模型，研究 DCH 对 SII 的影响。记录 24 小时死亡率。通过 H&E 染色观察组织学变化。通过免疫组织化学分析测定紧密连接蛋白 ZO-1（ZO-1）和粘蛋白 2（MUC2）的表达。通过酶联免疫吸附试验（ELISA）测定分泌型免疫球蛋白 A（sIgA）、二胺氧化酶（DAO）和肠脂肪酸结合蛋白（iFABP）。通过 ELISA 和定量实时 PCR（RT-qPCR）测定 IL-1β、IL-6 和 TNF-α。通过 16S rRNA 测序分析肠道微生物群。通过整合转录组和代谢组学方法分析 DCH 治疗 SII 的潜在靶点和途径。通过生化试剂盒测定总谷胱甘肽（T-GSH）、GSH、GSSG（GSH 的还原形式）、谷胱甘肽过氧化物酶（GPX）、超氧化物歧化酶（SOD）、丙二醛（MDA）和肝肾功能指标。
中剂量 DCH 可提高 SII 大鼠 24 小时死亡率，减轻回肠病理变化，增加 ZO-1、MUC2 和 sIgA 的表达水平。DCH 降低了血清中 DAO、iFABP 和回肠中的 IL-1β、IL-6、TNF-α。DCH 改善了 α-和 β-多样性，并调节了肠道微生物群的结构，埃希氏菌-志贺氏菌减少，拟杆菌和真杆菌增加。通过整合转录组和代谢组学分析，确定了 GSH 代谢是 DCH 对 SII 的关键途径。验证了 GSH/GSSG 和最常见的氧化应激标志物。DCH 增加了抗氧化 T-GSH、GSH、GPX 和 SOD，同时降低了 MDA，即脂质过氧化的标志物。最终，证明 DCH 对 SII 大鼠具有安全性，且具有肝保护和肾保护作用。
DCH 通过调节 SII 大鼠的肠道微生物群和增强谷胱甘肽代谢来改善脓毒性肠损伤，无肝毒性和肾毒性。
