中国组织工程研究

中国组织工程研究 ›› 2026, Vol. 30 ›› Issue (35): 9217-9230.doi: 10.12307/2026.254

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

郁平神安颗粒对失眠模型小鼠神经元自噬的调控作用

王玉河1，2，谢天宇1，2，马诗佳1，2，王玉娇1，2，李梦婷1，2，谢道俊2，3   

  1. 1安徽中医药大学，安徽省合肥市   230038；2安徽中医药大学第一临床医学院，安徽省合肥市   230031；3安徽中医药大学第一附属医院，安徽省合肥市   230031
  • 收稿日期:2025-09-11 修回日期:2025-11-28 出版日期:2026-12-18 发布日期:2026-04-28
  • 通讯作者: 谢道俊，博士，教授，主任医师，博士生导师，安徽中医药大学第一临床医学院，安徽省合肥市 230031；安徽中医药大学第一附属医院，安徽省合肥市 230031
  • 作者简介:王玉河，男，2000年生，安徽中医药大学在读硕士，主要从事睡眠障碍的临床与基础研究。
  • 基金资助:
    国家自然科学基金资助项目(81874389)，项目负责人：谢道俊

Regulatory effects of Yuping Shen’an Granules on neuronal autophagy in a mouse model of insomnia

Wang Yuhe1,2, Xie Tianyu1,2, Ma Shijia1,2, Wang Yujiao1,2, Li Mengting1,2, Xie Daojun2,3,*   

  1. 1Anhui University of Chinese Medicine, Hefei 230038, Anhui Province, China; 2The First Clinical College of Anhui University of Chinese Medicine, Hefei 230031, Anhui Province, China; 3The First Affiliated Hospital of Anhui University of Chinese Medicine, Hefei 230031, Anhui Province, China
  • Received:2025-09-11 Revised:2025-11-28 Online:2026-12-18 Published:2026-04-28
  • Contact: Xie Daojun, MD, Professor, Chief physician, Doctoral supervisor, The First Clinical College of Anhui University of Chinese Medicine, Hefei 230031, Anhui Province, China; The First Affiliated Hospital of Anhui University of Chinese Medicine, Hefei 230031, Anhui Province, China
  • About author:Wang Yuhe, MS candidate, Anhui University of Chinese Medicine, Hefei 230038, Anhui Province, China; The First Clinical College of Anhui University of Chinese Medicine, Hefei 230031, Anhui Province, China
  • Supported by:
    National Natural Science Foundation of China, No. 81874389 (to XDJ)

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摘要：




文题释义：
自噬：是细胞能量应激时启动的保守循环过程，通过形成直径500-1 000 nm的吞噬泡包裹异常组分，与溶酶体融合后降解内容物，回收氨基酸等原料。核心指标自噬流可通过微管相关蛋白轻链3Ⅱ/微管相关蛋白轻链3Ⅰ比值和P62降解效率评估。该机制对维持稳态和代谢调控至关重要，自噬功能紊乱与神经退行病、癌症及衰老密切相关。
海马体：是位于大脑颞叶内侧的双侧对称结构，成人单侧体积为3.0-3.5 cm³，主要由CA1-CA3区、齿状回和下托等亚区构成，在记忆编码与巩固、空间导航认知、情境情绪调节及睡眠-觉醒周期调控中发挥关键作用，海马体功能实现依赖于高度特化的突触可塑性及神经环路整合机制。

背景：现代医学治疗失眠伴焦虑多采用苯二氮䓬类药物，长期服用易致药物依赖及不良反应；中医药基于辨证论治，疗效确切，安全性高，具有多通路、多靶点综合作用，为该病治疗提供了新的思路与方向。
目的：探讨郁平神安颗粒通过磷脂酰肌醇3激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白信号通路对失眠伴焦虑小鼠海马神经元自噬的影响。 
方法：采用超高效液相色谱-质谱联用鉴定郁平神安颗粒成分，并结合网络药理学预测关键通路。将90只C57BL/6J雄性小鼠随机分为正常组、模型组和郁平神安低、中、高剂量组[3.125，6.25，12.5 g/(kg·d)]、右佐匹克隆组[0.06 mg/(kg·d)]，回复-模型组，回复-模型+磷脂酰肌醇3激酶抑制剂组[10 mg/(kg·d)]，回复-郁平神安[12.5 g/(kg·d)]+抑制剂组[10 mg/(kg·d)]，每组10只。除正常组外，其余各组均暴露于慢性不可预知温和刺激14 d和腹腔注射对氯苯丙氨酸4 d构建失眠伴焦虑小鼠模型。行为学评估采用旷场和戊巴比妥钠睡眠实验；苏木精-伊红染色与尼氏染色观察海马病理变化；酶联免疫吸附测定法检测5-羟色胺、γ-氨基丁酸、多巴胺、去甲肾上腺素水平；免疫荧光检测微管相关蛋白轻链3B表达；蛋白免疫印迹法检测磷脂酰肌醇3激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白通路相关蛋白和自噬相关蛋白(P62、Beclin-1、微管相关蛋白轻链3B)及5-羟色胺受体1A蛋白表达；透射电镜观察自噬小体数量。 
结果与结论：①超高效液相色谱-质谱联用鉴定出郁平神安颗粒的2 276种成分，筛选出35种主要成分；②网络药理学分析显示，磷脂酰肌醇3激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白通路为潜在作用靶点；③与正常组相比，模型组小鼠的探索行为增加、睡眠潜伏期延长(P < 0.01)，海马神经元损伤显著，5-羟色胺、γ-氨基丁酸水平降低，多巴胺、去甲肾上腺素水平升高(P < 0.01)，微管相关蛋白轻链3B荧光及蛋白表达上调(P < 0.01)，磷酸化磷脂酰肌醇3激酶/蛋白激酶B /哺乳动物雷帕霉素靶蛋白、P62、5-羟色胺受体1A表达下调，Beclin-1、微管相关蛋白轻链3B表达上调(P < 0.05)，自噬小体数量增多；④与模型组相比，回复-模型+抑制剂组损伤加重，郁平神安高剂量组及回复-郁平神安+抑制剂组可显著逆转上述变化，改善行为学指标，减轻神经元损伤，升高5-羟色胺、γ-氨基丁酸水平，降低多巴胺、去甲肾上腺素水平(P < 0.05)，回调自噬相关蛋白表达(P < 0.05)，自噬小体减少。结果表明：郁平神安通过激活磷脂酰肌醇3激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白通路、调节神经递质平衡并抑制海马神经元过度自噬，改善失眠伴焦虑症状。

https://orcid.org/0009-0007-9526-3303 (谢道俊) 


中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

关键词: 郁平神安颗粒, 失眠伴焦虑, 网络药理学, 磷脂酰肌醇3激酶(PI3K), 蛋白激酶B(AKT), 哺乳动物雷帕霉素靶蛋白(mTOR), 自噬

Abstract: BACKGROUND: Modern medicine often uses benzodiazepines for the treatment of insomnia complicated by anxiety; however, long-term use can lead to drug dependence and adverse reactions. Traditional Chinese medicine, based on syndrome differentiation and holistic treatment, demonstrates definite efficacy and high safety, offering multi-pathway and multi-target comprehensive effects, thereby providing new ideas and directions for the treatment of this condition.
OBJECTIVE: To investigate the effects of Yuping Shen’an Granules on hippocampal neuronal autophagy in a mouse model of insomnia complicated by anxiety via the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway.
METHODS: The components of Yuping Shen’an Granules were identified using ultra-performance liquid chromatography-mass spectrometry, and key pathways were predicted through network pharmacology. Ninety male C57BL/6J mice were randomly divided into nine groups (n=10 per group): normal group, model group, low-, medium-, and high-dose Yuping Shen’an groups [3.125, 6.25, and 12.5 g/(kg·d)], eszopiclone group [0.06 mg/(kg·d)], verification experiment-model group, verification experiment-model + PI3K inhibitor [10 mg/(kg·d)] group, and verification experiment-Yuping Shen’an [12.5 g/(kg·d)] + PI3K inhibitor [10 mg/(kg·d)] group. All groups except for the normal group underwent 2 weeks of chronic unpredictable mild stress and 4 days of intraperitoneal injection of p-chlorophenylalanine to establish a mouse model of insomnia complicated by anxiety. Behavioral assessments were conducted using the open field test and pentobarbital sodium-induced sleep test. Pathological changes in the hippocampus were observed via hematoxylin-eosin staining and Nissl staining. Serotonin, γ-aminobutyric acid, dopamine, and norepinephrine levels were measured using enzyme-linked immunosorbent assay. LC3B expression was detected through immunofluorescence staining. The Western blot assay was employed to measure the expression of proteins associated with the PI3K/Akt/mTOR pathway and autophagy-related proteins (P62, Beclin-1, LC3B), as well as the serotonin 1A receptor. The number of autophagosomes was quantified using a transmission electron microscope.  
RESULTS AND CONCLUSION: Ultra-performance liquid chromatography-mass spectrometry identified 2 276 components, with 35 major components screened. Network pharmacology analysis indicated the PI3K/Akt/mTOR pathway as a potential target. Compared with the normal group, the model group exhibited increased exploratory behavior, prolonged sleep latency (P < 0.01), significant hippocampal neuronal damage, decreased levels of serotonin and γ-aminobutyric acid, and increased levels of dopamine and norepinephrine (P < 0.01). Additionally, there was upregulation of LC3B fluorescence and protein expression (P < 0.01), downregulation of phosphorylated PI3K/Akt/mTOR, P62, and the serotonin 1A receptor, and upregulation of Beclin-1 and LC3B (P < 0.05), along with an increased number of autophagosomes. Compared with the model group, the verification experiment-model + PI3K inhibitor group exhibited aggravated damage, while the high-dose Yuping Shen’an group and the verification experiment-Yuping Shen’an + PI3K inhibitor group significantly reversed these changes. They improved behavioral indicators, alleviated neuronal damage, increased levels of serotonin and γ-aminobutyric acid, and decreased levels of dopamine and norepinephrine (P < 0.05). Additionally, they modulated the expression of autophagy-related proteins (P < 0.05) and reduced the number of autophagosomes. These findings indicate that Yuping Shen’an Granules improve symptoms of insomnia complicated by anxiety by activating the PI3K/Akt/mTOR pathway, regulating neurotransmitter balance, and inhibiting excessive autophagy in hippocampal neurons.

Key words: Yuping Shen’an Granules, insomnia with anxiety, network pharmacology, phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), mammalian target of rapamycin (mTOR), autophagy

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