Page 14 - 南京医科大学学报自然科学版

南京医科大学学报（自然科学版） 第43卷第6期
·756 · Journal of Nanjing Medical University（Natural Sciences） 2023年6月

·基础研究·

羟氯喹抑制MEK/ERK/ROS通路减少中性粒细胞胞外诱捕网形

成改善葡聚糖硫酸钠诱导的小鼠结肠炎

江婧月，张红杰 *
南京医科大学第一附属医院消化内科，江苏 南京 210029

［摘 要］ 目的：探讨羟氯喹（hydroxychloroquine，HCQ）对葡聚糖硫酸钠（dextran sulfate sodium salt，DSS）诱导的结肠炎模型鼠
肠道炎症的影响及其影响中性粒细胞胞外诱捕网（neutrophil extracellular trap，NET）形成的机制。方法：雄性C57BL/6鼠随机分为
正常组、DSS组（自由饮用3%DSS溶液+200 μL纯水每天灌胃）、DSS+HCQ组［自由饮用3%DSS溶液+200 μL HCQ（60 mg/kg）每天
灌胃］。评估小鼠疾病活动指数（disease activity index，DAI），造模7 d后处死小鼠，分析小鼠结肠长度，肠组织HE 染色评估肠
道炎症水平，荧光显微镜及荧光酶标仪检测各组小鼠结肠组织NET水平。收集志愿者外周血提取中性粒细胞进行细胞实验，
荧光显微镜及荧光酶标仪检测 NET 水平，DCFH⁃DA 荧光探针孵育后荧光酶标仪检测活性氧（reactive oxygen species，ROS）水
平，蛋白质印迹法检测p⁃MEK、MEK、p⁃ERK、ERK蛋白水平。结果：HCQ减轻DSS诱导的小鼠结肠炎并可抑制结肠炎模型鼠肠
组织中NET形成。在细胞实验中，HCQ可抑制体外培养中性粒细胞ROS释放和NET形成，同时抑制MEK/ERK信号通路的激
活。MEK 激动剂 C16⁃PAF 可逆转 HCQ 对 MEK/ERK 信号通路的抑制作用，增加 ROS 的释放，并增加 NET 的形成。结论：
HCQ 可减轻 DSS 小鼠肠道炎症，机制可能是通过抑制中性粒细胞 MEK/ERK 信号通路，减少 ROS 释放，从而减轻 NET 导致
的肠道损伤。
［关键词］ 溃疡性结肠炎；羟氯喹；中性粒细胞胞外诱捕网；活性氧
［中图分类号］ R574.62 ［文献标志码］ A ［文章编号］ 1007⁃4368（2023）06⁃756⁃08
doi：10.7655/NYDXBNS20230602

Hydroxychloroquine alleviates intestinal inflammation in dextran sulfate sodium salt
induced colitis mice by inhibiting MEK/ERK pathway and reducing the formation of ROS⁃
dependent neutrophil extracellular traps

JIANG Jingyue，ZHANG Hongjie *
Department of Gastroenterology，the First Affiliated Hospital of Nanjing Medical University，Nanjing 210029，China

［Abstract］ Objective：The current study aims to explore the effect and mechanism of hydroxychloroquine （HCQ）on
lipopolysaccharide（LPS）induced neutrophil extracelluar traps（NET）in dextran sulfate sodium salt（DSS）induced colitis mice.
Methods：Mice were randomly divided into control group，DSS group（3% DSS drinking water+200 μL pure water by gavage），and
DSS+HCQ group（3% DSS drinking water+200 μL HCQ of 60 mg/kg by gavage）. The disease activity index（DAI）score was evaluated
every day. The mice were killed after seven days，colon length of mice in each group were counted. The pathological changes of colon
tissue of mice in each group were observed by HE staining；the NET level was detected by fluorescence microscope and fluorescence
microplate reader. The peripheral blood of volunteers was collected to extract neutrophils for experiments in vitro，and the level of NET
with fluorescence microscope and fluorescence microplateanalyzer was detected. The level of reactive oxygen species（ROS）was
detected by fluorescence microplate analyzer after incubation with DCFH⁃DA fluorescence probe. The protein expression levels
of p⁃MEK，MEK，p⁃ERK and ERK were detected by Western blot. Results：HCQ alleviated intestinal inflammation in DSS model mice
by reducing the formation of NET in vivo and in vitro. In vitro，HCQ decreased the release of ROS，and inhibited the activation of MEK/
ERK signaling pathway. The MEK agonist C16⁃PAF reversed the inhibition of HCQ on MEK/ERK signaling pathway，enhanced the
release of ROS，and then increased the formation of NET. Conclusion：HCQ can reduce DSS induced colitis in mice through reducing the

［基金项目］ 国家自然科学基金（82070568）
∗
通信作者（Corresponding author），E⁃mail：hjzhang06@163.com