无忧汤通过激活Nrf2/HO-1通路抑制铁死亡减轻慢性睡眠剥夺模型大鼠海马神经元损伤并改善突触可塑性

doi:10.12122/j.issn.1673-4254.2026.08.04

南方医科大学学报 ›› 2026, Vol. 46 ›› Issue (8): 1754-1763.doi: 10.12122/j.issn.1673-4254.2026.08.04

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无忧汤通过激活Nrf2/HO-1通路抑制铁死亡减轻慢性睡眠剥夺模型大鼠海马神经元损伤并改善突触可塑性

何丽玲¹^,²(), 吴丹¹^,², 赖碧璇¹^,², 牛绮萱¹^,², 陈俊华¹^,², 高建红¹^,³, 龙清华¹^,², 王政喻¹^,²()

^1.湖北民族大学，医学部，湖北恩施 445000
^2.湖北民族大学，风湿病发生与干预湖北省重点实验室，湖北恩施 445000
^3.安徽中医药大学第一临床医学院，安徽合肥 230000

收稿日期:2026-01-12 出版日期:2026-08-20 发布日期:2026-08-01
通讯作者: 王政喻 E-mail:642755743@qq.com;1098796629@qq.com
作者简介:何丽玲，博士，讲师，E-mail: 642755743@qq.com
基金资助:
湖北省自然科学基金联合基金(2023AFD068);湖北省中医药管理局中医药科研项目(ZY20230047);湖北民族大学校内科研项目(XN2316);湖北民族大学研究生创新项目(MYK2023060)

Wuyou Decoction alleviates hippocampal neuronal injury and improves synaptic plasticity in rats with chronic sleep deprivation by inhibiting ferroptosis via activating the Nrf2/HO-1 pathway

Liling HE¹^,²(), Dan WU¹^,², Bixuan LAI¹^,², Qixuan NIU¹^,², Junhua CHEN¹^,², Jianhong GAO¹^,³, Qinghua LONG¹^,², Zhengyu WANG¹^,²()

^1.Department of Medicine, Hefei 23000, China
^2.Hubei Provincial Key Laboratory of Rheumatic Disease Occurrence and Intervention, Hubei Minzu University, Enshi 445000, China, Hefei 23000, China
^3.First Clinical Medical College of Anhui University of Traditional Chinese Medicine, Hefei 23000, China

Received:2026-01-12 Online:2026-08-20 Published:2026-08-01
Contact: Zhengyu WANG E-mail:642755743@qq.com;1098796629@qq.com

摘要/Abstract

摘要：

目的探讨中药方剂无忧汤对慢性睡眠剥夺（CSD）模型大鼠抑制铁死亡改善海马神经元损伤，保护突触可塑性的作用机制。方法将50只SD大鼠随机分为正常组，模型组及无忧汤低、中、高剂量组，10只/组。除正常组外，其余各组大鼠通过水平台睡眠剥夺法建立CSD模型，包括7 d适应性造模及21 d正式造模，并于正式造模开始同步灌胃给药28 d。之后使用莫里斯水迷宫评估认知功能，通过HE染色、尼氏染色和高尔基染色以及免疫荧光评估海马病理学和突触结构。使用透射电子显微镜、铁测定法和生化试剂盒定量铁和氧化应激标志物Fe²⁺、超氧化物歧化酶、丙二醛、谷胱甘肽（GSH）、还原型谷胱甘肽/氧化型谷胱甘肽（GSH/GSSG）。通过蛋白质印迹法检测参与铁调节和铁死亡蛋白铁蛋白重链1、转铁蛋白受体1、酰基辅酶A合成酶长链家族成员4（ACSL4）、溶质载体家族7成员11（SLC7A11）、谷胱甘肽过氧化物酶4、核因子E2相关因子2（Nrf2）、血红素加氧酶1（HO-1）的表达，采用分子对接技术探索无忧汤成分与铁死亡相关蛋白之间的相互作用。结果与正常组相比，模型组大鼠水迷宫逃避潜伏期增加，平台穿越次数及目标象限停留时间均缩短（P<0.01）。神经元结构变差，数目减少，突触结构受损（P<0.01）。增加氧化应激水平及铁沉积，同时上调Nrf2、HO-1的表达（P<0.01）。与模型组相比，无忧汤中剂量组改善了CSD大鼠的空间学习和记忆能力，莫里斯水迷宫中逃逸时间更短和平台穿越次数更高（P<0.01）。无忧汤中剂量组减轻了神经元退化，恢复了突触结构，并减少了海马体中的线粒体损伤（P<0.01，P<0.05）。无忧汤中剂量组减少铁的积累并缓解了CSD大鼠模型中的氧化应激（P<0.01），提高铁死亡相关标志物的表达，并上调Nrf2、HO-1基因表达（P<0.01，P<0.05）。结论无忧汤通过抑制铁死亡、减少神经元损伤以及通过激活Nrf2/HO-1信号通路增强突触修复，减轻了慢性睡眠剥夺引起的认知障碍，而中剂量效果最佳，这可能由于低剂量达不到作用阈值，高剂量超过阈值后，疗效反而可能减弱或出现不良反应有关。

关键词: 无忧汤, 慢性睡眠剥夺, 神经元损伤, 铁死亡, Nrf2/HO-1信号通路

Abstract:

Objective To investigate the mechanism of Wuyou Decoction (WYD) for ameliorating hippocampal neuron damage and protecting synaptic plasticity in rats with chronic sleep deprivation (CSD). Methods Fifty SD rats were randomly divided into normal control group, CSD model group, and low-, medium-, and high-dose WYD treatment groups (n=10). Except for those in the control group, all the rats were subjected to CSD modeling using the horizontal platform method. Cognitive function of the rats was assessed with Morris water mazetest, and hippocampal pathology, synaptic structure, iron content, oxidative stress markers, and ferroptosis-related protein expressions were evaluated. Molecular docking study was performed to explore the interactions between WYD components and ferroptosis-related proteins. Results CSD induced significant cognitive impairment and neuronal and synaptic damage in the rat models, causing also increased oxidative stress and iron deposition and upregulated Nrf2 and HO-1 expressions. Treatment with WYD, especially at the medium dose, significantly improved learning and memory abilities of the rat models, alleviated neuronal degeneration, restored synaptic structure, reduced iron accumulation and oxidative stress, and upregulated ferroptosis-related markers as well as Nrf2 and HO-1 expressions. Conclusion WYD, optimally at the medium dose, alleviates CSD-induced cognitive impairment in rats by inhibiting ferroptosis, reducing neuronal damage, and enhancing synaptic repair via activation of the Nrf2/HO-1 signaling pathway.

Key words: Wuyou Decoction, chronic sleep deprivation, neuron damage, ferroptosis, Nrf2/HO-1 signaling pathway

何丽玲, 吴丹, 赖碧璇, 牛绮萱, 陈俊华, 高建红, 龙清华, 王政喻. 无忧汤通过激活Nrf2/HO-1通路抑制铁死亡减轻慢性睡眠剥夺模型大鼠海马神经元损伤并改善突触可塑性[J]. 南方医科大学学报, 2026, 46(8): 1754-1763.

Liling HE, Dan WU, Bixuan LAI, Qixuan NIU, Junhua CHEN, Jianhong GAO, Qinghua LONG, Zhengyu WANG. Wuyou Decoction alleviates hippocampal neuronal injury and improves synaptic plasticity in rats with chronic sleep deprivation by inhibiting ferroptosis via activating the Nrf2/HO-1 pathway[J]. Journal of Southern Medical University, 2026, 46(8): 1754-1763.

图/表 10

图1 大鼠在MWM期间的学习记忆水平

Fig.1 Statistical charts of the performance of the rats in Morris water maze test. A: Representative trajectories in the navigation test in each group. B: Progression of escape latency during the 5-day training. C: Statistical chart of final escape latency. D: Number of platform crossings. E: Duration of rats' stay in the target quadrant. F: Swimming speed during spatial exploration (n=10). **P<0.01 vs NC group, ##P<0.01 vs Model group.

图2 各组大鼠海马CA1与CA3区的HE染色

Fig.2 HE staining of the hippocampal CA1 and CA3 regions of the rats in each group (Original magnification: ×200, n=3).

图3 各组大鼠海马CA1与CA3区Nissl染色情况

Fig.3 Nissl staining of the hippocampal CA1 and CA3 regions of the rats in each group (×200, n=3). **P<0.01 vs NC group, #P<0.05, ##P<0.01 vs Model group.

图4 各组大鼠海马组织免疫组化检测

Fig.4 Immunohistochemical detection of hippocampal tissue of the rats in each group (×200, n=3). **P<0.01 vs NC group, #P<0.05, ##P<0.01 vs Model group.

图5 海马突触的相关检测结果

Fig.5 Result of hippocampal synapses. A-C: Golgi staining. D-F: Immunofluorescence intensity of PSD95 and SYN1 (×200, n=3). **P<0.01 vs NC group, #P<0.05, ##P<0.01 vs Model group.

图6 透射电镜观察大鼠海马线粒体结构

Fig.6 Mitochondrial structure in rat hippocampus detected by transmission electron microscopy (×10 000).

图7 各组大鼠氧化应激水平及铁代谢相关蛋白FTH1、TfR1的含量

Fig.7 Oxidative stress levels (A) and contents of iron metabolism-related proteins FTH1 and TfR1 (B) in rats from different groups (n=6 or 3). **P<0.01 vs NC group; #P<0.05, ##P<0.01 vs Model group.

图8 蛋白免疫印迹法检的含量铁死亡相关蛋白ACSL4、GPX4、SCL7A11及通路蛋白Nrf2、HO-1表达

Fig.8 Expression levels of ferroptosis-related proteins (ACSL4, GPX4, and SCL7A11) and signaling pathway proteins (Nrf2 and HO-1) detected by Western blotting (n=3). **P<0.01 vs NC group, #P<0.05, ##P<0.01 vs Model group.

图9 阿魏酸、芍药苷与Nrf2、HO-1和GPX4的分子对接图

Fig.9 Molecular docking diagrams of paeonol, paeonol glycosides, Nrf2, HO-1, and GPX4.

表1 阿魏酸、芍药苷与Nrf2、HO-1和GPX4的结合能

Tab.1 The binding energy of ferulic acid, paeoniflorin with Nrf2, HO-1, and GPX4

Key compound	Affinity (kcal/mol)
Key compound	Nfr2	HO-1	GPX4
Ferulic acid	-7.1	-5.7	-6.6
Paeoniflorin	-9.6	-7.2	-7.2

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无忧汤通过激活Nrf2/HO-1通路抑制铁死亡减轻慢性睡眠剥夺模型大鼠海马神经元损伤并改善突触可塑性

Wuyou Decoction alleviates hippocampal neuronal injury and improves synaptic plasticity in rats with chronic sleep deprivation by inhibiting ferroptosis via activating the Nrf2/HO-1 pathway

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