中医药学报

基于EphA4/ephrinA3信号通路研究安寐丹对星形胶质细胞和神经元氧糖剥夺/复氧损伤的保护作用及机制

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吴艺璇 ¹ , 李娅琪 ¹ , 黎柱江 ¹ , 姜杰程 ¹ , 张卓 ¹ , 王平 ¹ , 谢光璟 ¹^,²^,^* , 黄攀攀 ¹^,²

中医药学报 | 实验研究 2026,54(10): 13-20

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中医药学报 |实验研究 2026 , 54 (10) : 13 -20

基于EphA4/ephrinA3信号通路研究安寐丹对星形胶质细胞和神经元氧糖剥夺/复氧损伤的保护作用及机制

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吴艺璇¹, 李娅琪¹, 黎柱江¹, 姜杰程¹, 张卓¹, 王平¹, 谢光璟¹^,²^,^*, 黄攀攀¹^,²

作者信息

^1.湖北中医药大学老年脑健康中医药防护技术与新产品研发教育部工程研究中心，湖北　武汉　430065

^2.湖北时珍实验室，湖北　武汉　430065

通讯作者:

^* 谢光璟（1991-），男，博士，副教授，硕士生导师，从事中医药防治老年病研究。

作者简介:

吴艺璇（2000-），女，硕士研究生，从事中医延缓衰老理论及应用研究。

Study on the Protective Effect and Mechanism of Anmei Dan on Astrocyte and Neuron Oxygen-Glucose Deprivation/Reoxygenation Injury Based on the EphA4/ephrinA3 Signaling Pathway

Yixuan WU¹, Yaqi LI¹, Zhujiang LI¹, Jiecheng JIANG¹, Zhuo ZHANG¹, Ping WANG¹, Guangjing XIE¹^,²^,^*, Panpan HUANG¹^,²

Affiliations

^1.Engineering Research Center of Ministry of Education for Traditional Chinese Medicine Prevention and Protection Technologies and New Product Development for Senile Brain Health, Hubei University of Chinese Medicine, Wuhan 430065, China

^2.Hubei Shizhen Laboratory, Wuhan 430065, China

出版时间: 2026-10-20 doi: 10.19664/j.cnki.1002-2392.260196

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摘要

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目的：

探究安寐丹通过调控酪氨酸激酶肾上腺素受体4（EphA4）/酪氨酸蛋白激酶A3受体（ephrinA3）信号通路对星形胶质细胞和神经元氧糖剥夺/复氧（OGD/R）的保护作用及机制。

方法：

体外建立小鼠星形胶质细胞（C8-D1A细胞）/海马神经元（HT22细胞）氧糖剥夺/复氧（OGD/R）模型，同时制备对照血清及安寐丹含药血清（AMD）。设立对照组、OGD/R组、OGD/R+AMD组及OGD/R+褪黑素组。通过细胞增殖与活性检测（CCK-8）法观察细胞的增殖和存活情况，筛选安寐丹含药血清最佳干预浓度；利用流式细胞术检测HT22/C8-D1A细胞凋亡率，免疫荧光染色检测神经元标志物NeuN及星形胶质细胞标志物GFAP、神经发生标志物5′-溴脱氧尿嘧啶核苷（Brdu）的荧光强度，蛋白免疫印迹技术（Western blot）检测EAAT1、EAAT2、ephrinA3、EphrA4及SYN蛋白的表达，透射电镜观察细胞超微结构。

结果：

与对照组相比，OGD/R组HT22/C8-D1A细胞标志物荧光强度减弱（P<0.01），神经元形态异常，EphA4/ephrinA3蛋白表达水平上调（P<0.01），EAAT1、EAAT2蛋白表达量下降（P<0.05，P<0.01）。与OGD/R组相比，OGD/R+AMD组GFAP和NeuN平均荧光强度增强，神经元形态有所改善，星形胶质细胞EphA4/ephrinA3水平降低（P<0.01），EAAT1、EAAT2、SYN蛋白表达量上调（P<0.05），细胞凋亡率显著下降（P<0.01）。

结论：

安寐丹可能通过下调EphA4/ephrinA3信号通路，介导谷氨酸转运，缓解OGD/R诱导的HT22/C8-D1A细胞损伤，这可能是其改善睡眠剥夺相关神经损伤的作用机制。

关键词

睡眠剥夺 / 氧糖剥夺/复氧 / 安寐丹 / 星形胶质细胞 / 酪氨酸激酶肾上腺素受体4/酪氨酸蛋白激酶A3受体信号通路

Abstract

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Objective:

To investigate the protective effects and underlying mechanisms of Anmeidan (AMD) against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced injury in astrocytes and neurons by regulating the EphA4/ephrinA3 signaling pathway.

Methods:

An invitro OGD/R model was established using mouse astrocytes (C8-D1A) and hippocampal neurons (HT22 cells). Control serum and AMD-containing serum were prepared. Cells were divided into control, OGD/R, OGD/R+AMD, and OGD/R+melatonin groups. Cell proliferation and viability were assessed via CCK-8 assay to determine the optimal AMD serum concentration. Apoptosis rates were measured by flow cytometry. Immunofluorescence staining was performed to evaluate fluorescence intensity of neuronal marker NeuN, astrocytic marker GFAP, and neurogenesis marker BrdU. Protein expression of ephrinA3, EphA4, EAAT1, EAAT2, and SYN was analyzed by Western blot. Cellular ultrastructure was observed via transmission electron microscopy (TEM).

Results:

Compared with the control group, the OGD/R group showed reduced fluorescence intensity of NeuN and GFAP (P<0.01), abnormal neuronal morphology, upregulated EphA4/ephrinA3 protein expression (P<0.01), and decreased EAAT1/EAAT2 levels (P<0.05, P<0.01). AMD treatment significantly enhanced GFAP and NeuN fluorescence intensity, improved neuronal morphology, downregulated EphA4/ephrinA3 expression (P<0.01), upregulated EAAT1, EAAT2, SYN (P<0.05), and reduced apoptosis rates (P<0.01).

Conclusion:

Anmeidan alleviates OGD/R-induced astrocyte and neuronal injury by downregulating the EphA4/ephrinA3 signaling pathway and modulating glutamate transport, which may underlie its therapeutic potential in sleep deprivation-related neural damage.

Key words

Sleep deprivation / Oxygen-glucose deprivation/reoxygenation / Anmei Dan / Astrocytes / Tyrosine kinase adrenergic receptor 4/tyrosine protein kinase A3 receptor signaling pathway

引用本文

吴艺璇, 李娅琪, 黎柱江, 姜杰程, 张卓, 王平, 谢光璟, 黄攀攀. 基于EphA4/ephrinA3信号通路研究安寐丹对星形胶质细胞和神经元氧糖剥夺/复氧损伤的保护作用及机制. 中医药学报, 2026 , 54 (10) : 13 -20 . DOI: 10.19664/j.cnki.1002-2392.260196

Yixuan WU, Yaqi LI, Zhujiang LI, Jiecheng JIANG, Zhuo ZHANG, Ping WANG, Guangjing XIE, Panpan HUANG. Study on the Protective Effect and Mechanism of Anmei Dan on Astrocyte and Neuron Oxygen-Glucose Deprivation/Reoxygenation Injury Based on the EphA4/ephrinA3 Signaling Pathway[J]. Acta Chinese Medicine and Pharmacology, 2026 , 54 (10) : 13 -20 . DOI: 10.19664/j.cnki.1002-2392.260196

基金

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国家自然科学基金项目(82305354; 82374310; 82174247)

参考文献

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排序方式：

[1]

KONAKANCHI S, RAAVI V, ML H K, et al. Effect of chronic sleep deprivation and sleep recovery on hippocampal CA3 neurons, spatial memory and anxiety-like behavior in rats[J]. Neurobiol Learn Mem, 2022, 187: 107559.

[2]

CREERY J D, BRANG D J, ARNDT J D, et al. Electrophysiological markers of memory consolidation in the human brain when memories are reactivated during sleep[J]. Proc Natl Acad Sci U S A, 2022, 119(44): e2123430119.

[3]

TANASIC S, MATTUSCH C, WAGNER E M, et al. Desipramine targets astrocytes to attenuate synaptic plasticity via modulation of the ephrinA3/EphA4 signalling[J]. Neuropharmacology, 2016, 105: 154-163.

[4]

IRWIN M R. Sleep and inflammation: partners in sickness and in health[J]. Nat Rev Immunol, 2019, 19(11): 702-715.

[5]

ATROOZ F, SALIM S. Sleep deprivation, oxidative stress and inflammation[M]//Inflammatory Disorders, Part A. Amsterdam: Elsevier, 2020: 309-336.

[6]

MOHD SHAFIE A S, ABD AZIZ C B, LONG I, et al. Impact of rapid eye movement sleep deprivation on pain behaviour and oxidative stress in the thalamus: role of tualang honey supplementation[J]. Malays J Med Sci, 2022, 29(2): 69-79.

[7]

YUAN Y, ZHAI Y Y, CHEN J J, et al. Kaempferol ameliorates oxygen-glucose deprivation/reoxygenation-induced neuronal ferroptosis by activating Nrf2/SLC7A11/GPX4axis[J]. Biomolecules, 2021, 11(7): 923.

[8]

XUE R, WAN Y H, SUN X Q, et al. Nicotinic mitigation of neuroinflammation and oxidative stress after chronic sleep deprivation[J]. Front Immunol, 2019, 10: 2546.

[9]

YANG X, GONG R, QIN L W, et al. Trafficking of NMDA receptors is essential for hippocampal synaptic plasticity and memory consolidation[J]. Cell Rep, 2022, 40(7): 111217.

[10]

徐波, 王平, 夏婧, 等. 基于OX1R/PLCβ-1/PKCα/ERK1/2信号通路探讨安寐丹对失眠模型大鼠肝脏神经递质和昼夜节律的影响及机制[J]. 中国实验方剂学杂志, 2024, 30(15): 11-20.

[11]

李非洲, 谢光璟, 徐波, 等. 安寐丹调控ROS/PARP-1/AIF通路改善睡眠剥夺模型神经元损伤[J]. 中国中西医结合杂志, 2024, 44(8): 952-958.

[12]

谢光璟, 徐波, 夏婧, 等. 安寐丹对睡眠剥夺大鼠海马CA1区细胞结构及神经元损伤的保护作用[J]. 中国实验方剂学杂志, 2022, 28(7): 26-32.

[13]

DENG Y, DUAN R, DING W L, et al. Astrocyte-derived exosomal nicotinamide phosphoribosyltransferase (Nampt) ameliorates ischemic stroke injury by targeting AMPK/mTOR signaling to induce autophagy[J]. Cell Death Dis, 2022, 13: 1057.

[14]

SUN B, OU H, REN F, et al. Propofol inhibited autophagy through Ca²⁺/CaMKKβ/AMPK/mTOR pathway in OGD/R-induced neuron injury[J]. Mol Med, 2018, 24(1): 58.

[15]

张军路, 孙康, 吴艺璇, 等. 基于Cyt C调节细胞凋亡途径探讨安寐丹改善老年睡眠剥夺模型致认知损伤的作用机制[J]. 中国实验方剂学杂志, 2024, 30(19): 1-9.

[16]

XU S, LI Y, CHEN J P, et al. Oxygen glucose deprivation/re-oxygenation-induced neuronal cell death is associated with Lnc-D63785 m6A methylation and miR-422a accumulation[J]. Cell Death Dis, 2020, 11: 816.

[17]

RIEMANN D, KRONE L B, WULFF K, et al. Sleep, insomnia, and depression[J]. Neuropsychopharmacology, 2020, 45(1): 74-89.

[18]

ZHANG R, LAHENS N F, BALLANCE H I, et al. A circadian gene expression atlas in mammals: Implications for biology and medicine[J]. Proc Natl Acad Sci U S A, 2014, 111(45): 16219-16224.

[19]

丁灵, 王欣, 上官惠子, 等. 酸枣仁的化学成分及治疗失眠的作用机制研究进展[J]. 现代中医药, 2025, 45(3): 14-23.

[20]

许红蕾, 王庭芳, 张川. 丹参素及其衍生物药理作用机制和应用研究进展[J]. 中国现代应用药学, 2021, 38(2): 237-243.

[21]

冯鹏, 王明露, 王宇彤, 等. 石菖蒲挥发油化学成分及药理作用研究进展[J]. 中药药理与临床, 2025, 41(6): 100-109.

[22]

刘继艳, 王冰, 李超宇, 等. 五味子非药用部位活性成分和药理作用研究进展[J]. 中草药, 2024, 55(9): 3179-3189.

[23]

HRISTOVSKA I, ROBERT M, COMBET K, et al. Sleep decreases neuronal activity control of microglial dynamics in mice[J]. Nat Commun, 2022, 13: 6273.

[24]

ZHU Z M, TAO X, DAI T T, et al. Cognitive-exercise dual-task attenuates chronic cerebral ischemia-induced cognitive impairment by activating cAMP/PKA pathway through inhibiting EphrinA3/EphA4[J]. Exp Neurol, 2024, 372: 114617.

[25]

PUENTES-MESTRIL C, DELORME J, WANG L J, et al. Sleep loss drives brain region-specific and cell type-specific alterations in ribosome-associated transcripts involved in synaptic plasticity and cellular timekeeping[J]. J Neurosci, 2021, 41(25): 5386-5398.

[26]

LI J G, LIU N H, WANG Y, et al. Inhibition of EphA4 signaling after ischemia-reperfusion reduces apoptosis of CA1 pyramidal neurons[J]. Neurosci Lett, 2012, 518(2): 92-95.

[27]

ZHOU Z X, XU L J, WANG H N, et al. EphA4/ephrinA3 reverse signaling mediated downregulation of glutamate transporter GLAST in Müller cells in an experimental glaucoma model[J]. Glia, 2023, 71(3): 720-741.

[28]

BRANDEBURA A N, PAUMIER A, ONUR T S, et al. Astrocyte contribution to dysfunction, risk and progression in neurodegenerative disorders[J]. Nat Rev Neurosci, 2023, 24(1): 23-39.

[29]

GROC L, CHOQUET D. Linking glutamate receptor movements and synapse function[J]. Science, 2020, 368(6496): eaay4631.

[30]

PIETRANCOSTA N, DJIBO M, DAUMAS S, et al. Molecular, structural, functional, and pharmacological sites for vesicular glutamate transporter regulation[J]. Mol Neurobiol, 2020, 57(7): 3118-3142.

[31]

XU L J, GAO F, CHENG S, et al. Activated ephrinA3/EphA4 forward signaling induces retinal ganglion cell apoptosis in experimental glaucoma[J]. Neuropharmacology, 2020, 178: 108228.

[32]

LIU J H, ZHANG M, WANG Q, et al. Distinct roles of astroglia and neurons in synaptic plasticity and memory[J]. Mol Psychiatry, 2022, 27(2): 873-885.

2026年第54卷第10期

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文章信息

doi: 10.19664/j.cnki.1002-2392.260196

接收时间：2025-07-10
首发时间：2026-10-14
出版时间：2026-10-20

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收稿日期：2025-07-10
修回日期：2026-05-11

基金

国家自然科学基金项目(82305354; 82374310; 82174247)

作者信息

^1.湖北中医药大学老年脑健康中医药防护技术与新产品研发教育部工程研究中心，湖北　武汉　430065

^2.湖北时珍实验室，湖北　武汉　430065

通讯作者:

^* 谢光璟（1991-），男，博士，副教授，硕士生导师，从事中医药防治老年病研究。

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https://castjournals.cast.org.cn/joweb/zyyxb/CN/10.19664/j.cnki.1002-2392.260196

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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