Liao Wei, Wu Xinyi, Gong Jianping, Li Shengwei
Department of Hepatobiliary Surgery, The Second Affiliated Hospital of Chongqing Medical University, Yuzhong District, Chongqing, China.
Shock. 2026 Jan 1;65(1):93-103. doi: 10.1097/SHK.0000000000002673. Epub 2025 Dec 30.
Liver ischemia-reperfusion injury (LIRI) is a main cause of complication development regarding the liver. This study examines how Oridonin mitigates oxidative stress and pyroptosis in macrophages during LIRI by inhibiting the mitochondrial reactive oxygen species (ROS)/thioredoxin-interacting protein (TXNIP)/NOD-like receptor protein 3 (NLRP3) signaling pathway.
LIRI mouse models were treated with Oridonin at doses of 1, 5, and 10 mg/kg. Liver damage was evaluated through serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) levels enzyme linked immunosorbent assay (ELISA), histological analysis (HE staining and Suzuki scoring), and immunofluorescence for NLRP3 and macrophage markers (F4/80, inducible nitric oxide synthase [iNOS]). Inflammatory cytokines (Tumor necrosis factor alpha, interleukin-1beta [IL-1β], IL-18, and IL-6) and oxidative stress markers (malondialdehyde [MDA], total superoxide dismutase [T-SOD], and glutathione peroxidase [GSH-Px]) were measured using ELISA. M1 macrophage markers (iNOS, CD86, and CD80) were assessed via quantitative real-time polymerase chain reaction (qRT-PCR). Primary hepatic macrophages were isolated for flow cytometry, cell-counting kit (CCK)-8 assays, and ROS detection. Protein expression was analyzed using Western Blot and immunohistochemistry.
Oridonin dose-dependently reduced liver damage, lowering ALT/AST levels and improving histological scores. Depletion of Kupffer cells with clodronate liposomes abolished Oridonin's protective effects, indicating macrophage mediation. LIRI increased M1 macrophages, F4/80⁺/iNOS⁺ cells, and inflammatory cytokines, all of which were partially reversed by Oridonin. In vitro, Oridonin reduced oxidative stress and pyroptosis in hypoxia-reoxygenation-exposed hepatic macrophages, evidenced by decreased ROS and lower levels of pyroptosis-related proteins (gasdermin D, Cleaved-Caspase-1, and IL-1β). Mechanistically, Oridonin may ameliorate oxidative stress and pyroptosis in hepatic macrophages of LIRI mice by inhibiting the ROS/TXNIP/NLRP3 pathway.
Oridonin effectively protects against LIRI by decreasing macrophage M1 polarization, oxidative stress, and pyroptosis through inhibition of the mitochondrial ROS/TXNIP/NLRP3 pathway.
肝缺血再灌注损伤（LIRI）是肝脏并发症发生的主要原因。本研究探讨冬凌草甲素如何通过抑制线粒体活性氧（ROS）/硫氧还蛋白相互作用蛋白（TXNIP）/NOD样受体蛋白3（NLRP3）信号通路减轻LIRI期间巨噬细胞中的氧化应激和焦亡。
用1、5和10mg/kg剂量的冬凌草甲素处理LIRI小鼠模型。通过血清丙氨酸氨基转移酶（ALT）/天冬氨酸氨基转移酶（AST）水平酶联免疫吸附测定（ELISA）、组织学分析（苏木精-伊红染色和铃木评分）以及NLRP3和巨噬细胞标志物（F4/80、诱导型一氧化氮合酶[iNOS]）的免疫荧光评估肝损伤。使用ELISA测量炎性细胞因子（肿瘤坏死因子α、白细胞介素-1β[IL-1β]、IL-18和IL-6）和氧化应激标志物（丙二醛[MDA]、总超氧化物歧化酶[T-SOD]和谷胱甘肽过氧化物酶[GSH-Px]）。通过定量实时聚合酶链反应（qRT-PCR）评估M1巨噬细胞标志物（iNOS、CD86和CD80）。分离原代肝巨噬细胞用于流式细胞术、细胞计数试剂盒（CCK）-8测定和ROS检测。使用蛋白质免疫印迹和免疫组织化学分析蛋白质表达。
冬凌草甲素剂量依赖性地减轻肝损伤，降低ALT/AST水平并改善组织学评分。用氯膦酸脂质体消耗枯否细胞消除了冬凌草甲素的保护作用，表明巨噬细胞起介导作用。LIRI增加了M1巨噬细胞、F4/80⁺/iNOS⁺细胞和炎性细胞因子，所有这些均被冬凌草甲素部分逆转。在体外，冬凌草甲素降低了缺氧复氧暴露的肝巨噬细胞中的氧化应激和焦亡，表现为ROS降低和焦亡相关蛋白（gasdermin D、裂解的半胱天冬酶-1和IL-1β）水平降低。机制上，冬凌草甲素可能通过抑制ROS/TXNIP/NLRP3途径改善LIRI小鼠肝巨噬细胞中的氧化应激和焦亡。
冬凌草甲素通过抑制线粒体ROS/TXNIP/NLRP3途径减少巨噬细胞M1极化、氧化应激和焦亡，从而有效预防LIRI。
