加味逍遥丸通过神经递质调节、抗炎抗氧化及肠道菌群调控改善大鼠的抑郁样行为

doi:10.12122/j.issn.1673-4254.2025.02.16

南方医科大学学报 ›› 2025, Vol. 45 ›› Issue (2): 347-358.doi: 10.12122/j.issn.1673-4254.2025.02.16

• • 上一篇

加味逍遥丸通过神经递质调节、抗炎抗氧化及肠道菌群调控改善大鼠的抑郁样行为

刘莹¹^,²(), 李柏睿¹, 李永财¹, 常禄博¹, 王娇¹, 杨琳¹, 颜永刚¹^,², 屈凯³, 刘继平¹^,⁴, 张岗¹(), 沈霞¹^,²()

^1.陕西中医药大学药学院，陕西西咸新区 712046
^2.陕西省秦岭中草药应用开发工程技术研究中心，陕西西咸新区 712046
^3.陕西省中医医院，陕西西安 710003
^4.陕西省中医药管理局中药药效机制与物质基础重点研究室，陕西西咸新区 712046

收稿日期:2024-11-17 出版日期:2025-02-20 发布日期:2025-03-03
通讯作者: 张岗,沈霞 E-mail:farewell991007@163.com;jay_gumling2003@aliyun.com;jxrain@163.com
作者简介:刘莹，在读硕士研究生，E-mail: farewell991007@163.com
基金资助:
陕西省自然科学基础研究计划项目(2022JM-538);重点产业创新链(群)-社会发展领域(2024SF-ZDCYL-03-04);陕西省中医药中青年科技骨干人才项目(11610000MB29656749/2020-00894)

Jiawei Xiaoyao Pills improves depression-like behavior in rats by regulating neurotransmitters, inhibiting inflammation and oxidation and modulating intestinal flora

Ying LIU¹^,²(), Borui LI¹, Yongcai LI¹, Lubo CHANG¹, Jiao WANG¹, Lin YANG¹, Yonggang YAN¹^,², Kai QV³, Jiping LIU¹^,⁴, Gang ZHANG¹(), Xia SHEN¹^,²()

^1.School of Pharmacy, Shaanxi University of Chinese Medicine, Xixian New Area 712046, China
^2.Qinling Chinese Herbal Medicine Application and Development Engineering Technology Research Center, Xixian New Area 712046, China
^3.Shaanxi Provincial Hospital of Traditional Chinese Medicine, Xi'an 710003, China
^4.Key Laboratory of Pharmacodynamic Mechanism and Material Basis of Traditional Chinese Medicine, Shaanxi Provincial Administration of Traditional Chinese Medicine, Xixian New Area 712046, China

Received:2024-11-17 Online:2025-02-20 Published:2025-03-03
Contact: Gang ZHANG, Xia SHEN E-mail:farewell991007@163.com;jay_gumling2003@aliyun.com;jxrain@163.com

摘要/Abstract

摘要：

目的基于网络药理学、代谢组学和16srDNA探究加味逍遥丸（JWXYP）改善抑郁样行为的功效物质及作用机制。方法通过TCMSP、TCMIP等数据库，筛选加味逍遥丸治疗抑郁症的功效物质群、关键靶点及通路。将SD大鼠随机分为空白（Control）组、模型（CUMS）组、阳性组（1.8 mg/kg）组和加味逍遥丸低、中、高（JWXYP，1.44、2.88、4.32 g·kg^-1·d^-1）组（n=6）。通过行为学实验评估大鼠抑郁样行为， HE染色观察肝脏和海马病理变化，ELISA检测血清和脑组织生化指标。血清代谢组学采用OPLS-DA方法筛选差异代谢物并进行通路分析。盲肠内容物进行16srDNA基因测序分析肠道菌群变化。结果网络药理学研究发现，加味逍遥丸中的薄荷酮和丹皮酚等关键成分可通过血脑屏障，调节炎症通路保护神经系统。动物实验显示，加味逍遥丸均显著改善模型大鼠的体质量、糖水偏好率及旷场中央区域活动时间（P<0.05）；明显减少肝脏炎性细胞浸润和细胞坏死，改善海马神经元结构；降低血清中肿瘤坏死因子-α、白细胞介素-1β、白细胞介素-6水平以及脂多糖结合蛋白，上调外周血和大脑中5-羟色胺、血管活性肠肽的浓度（P<0.05），其中高剂量效果最为显著（P<0.01）。代谢组学分析显示，加味逍遥丸干预的差异代谢物包括吲哚-3-乙酰胺、乙酰左旋肉碱、3-甲基-L-组氨酸等，涉及次级胆汁酸生物合成、精氨酸和脯氨酸代谢和D-氨基酸代谢等多条通路（P<0.05）。16srDNA分析发现，加味逍遥丸组的Chao1、Observed和Shannon指数均明显升高，肠道菌群丰富度和均匀度增加，且菌鼠乳杆菌、肠乳酸杆菌的丰度显著提高。结论加味逍遥丸通过调节神经递质、抗炎抗氧化以及肠道菌群三条途径相互作用，改善情绪和认知功能减轻抑郁样症状。

关键词: 加味逍遥丸, 网络药理学, 肠道菌群, 血清代谢组, 抑郁样行为

Abstract:

Objective To explore the bioactive components in Jiawei Xiaoyao Pills (JWXYP) and their mechanisms for alleviating depression-like behaviors. Methods The active compounds, key targets, and pathways of JWXYP were identified using TCMSP and TCMIP databases. Thirty-six SD rats were randomized equally into 6 groups including a control group and 5 chronic unpredictable mild stress (CUMS)-induced depression groups. After modeling, the 5 model groups were treated with daily gavage of normal saline, 1.8 mg/kg fluoxetine hydrochloride (positive control drug), or JWXYP at 1.44, 2.88, and 4.32 g/kg. The depression-like behaviors of the rats were evaluated using behavioral tests, and pathological changes in the liver and hippocampus were examined with HE staining. The biochemical indicators in the serum and brain tissues were detected using ELISA. Serum metabolomics analysis was performed to identify the differential metabolites using OPLS-DA, and gut microbiota changes were analyzed using 16S rDNA sequencing. Results Network pharmacology revealed that menthone and paeonol in JWXYP were capable of penetrating the blood-brain barrier to regulate inflammatory pathways and protect the nervous system. In the rat models subjected to CUMS, treatment with JWXYP significantly improved body weight loss, sucrose preference and open field activities, reduced liver inflammation, alleviated structural changes in the hippocampal neurons, decreased serum levels of TNF‑α, IL-1β, IL-6 and LBP, and increased 5-HT and VIP concentrations in the serum and brain tissue, and these effects were the most pronounced in the high-dose group. Metabolomics analysis showed changes in such metabolites as indole-3-acetamide and acetyl-L-carnitine in JWXYP-treated rats, involving the pathways for bile acid biosynthesis and amino acid metabolism. 16S rDNA analysis demonstrated increased gut microbiota diversity and increased abundance of Lactobacillus species in JWXYP-treated rats. Conclusion JWXYP alleviates depression-like symptoms in rats by regulating the neurotransmitters, inhibiting inflammation and oxidation, and modulating gut microbiota.

Key words: Jiawei Xiaoyao Pills, network pharmacology, intestinal flora, serum metabolome, depression-like behavior

刘莹, 李柏睿, 李永财, 常禄博, 王娇, 杨琳, 颜永刚, 屈凯, 刘继平, 张岗, 沈霞. 加味逍遥丸通过神经递质调节、抗炎抗氧化及肠道菌群调控改善大鼠的抑郁样行为[J]. 南方医科大学学报, 2025, 45(2): 347-358.

Ying LIU, Borui LI, Yongcai LI, Lubo CHANG, Jiao WANG, Lin YANG, Yonggang YAN, Kai QV, Jiping LIU, Gang ZHANG, Xia SHEN. Jiawei Xiaoyao Pills improves depression-like behavior in rats by regulating neurotransmitters, inhibiting inflammation and oxidation and modulating intestinal flora[J]. Journal of Southern Medical University, 2025, 45(2): 347-358.

图/表 8

图1 网络药理学结果

Fig.1 Network pharmacology results. A: JWXYP and depression intersection genes. B: MCODE algorithm for screening the key targets. C: Chinese medicine-component-target network diagram. D: GO analysis of the key genes. E: Bubble map of KEGG analysis of the key genes.

表1 化学成分理化性质表

Tab.1 Physicochemical properties of the bioactive compounds in Jiawei Xiaoyao Pills （JWXYP）

Compound	MW	OB(%)	BBB	DL
Quercetin	302.25	46.43	-0.77	0.28
Luteolin	286.25	36.16	-0.84	0.25
Kaempferol	286.25	41.88	-0.55	0.24
Thymol	150.24	41.47	1.68	0.03
Stigmasterol	412.77	43.83	1	0.76
Paeonol	166.19	28.79	0.84	0.04
Oleanolic acid	456.78	29.02	0.07	0.76
Saikosaponin A	781.1	32.39	-2.93	0.09
Pulegone	152.26	51.6	1.74	0.03
Isoliquiritigenin	256.27	85.32	-0.41	0.15
Paeoniflorin	480.51	53.87	-1.86	0.79
Saikogenin F	472.78	25.88	-0.71	0.63
Licochalcone A	338.43	40.79	-0.21	0.29

图2 大鼠模型建立及行为学测试结果测定

Fig.2 Establishment of depression rat models and behavioral test results of the rats. A: Body weight curves of rats in each group. B: Sugar water preference test. C: External hair of rats before modeling (1), after modeling (2) and after drug treatment (3). D: Open- field test of the rats (activity time, distance of the edge, total distance, and number of activities). E: Trajectory diagram of the rats in open field test. *P<0.05, **P<0.01, ***P<0.001 vs model group (CUMS); #P<0.05 vs control group.

图3 ELISA结果、肝脾指数、肝脏照片及HE染色结果

Fig.3 ELISA results, liver and spleen index, liver and hippocampus HE staining results. A, D: Content of VIP and 5-HT in the brain tissues. B, C, E-H: Serum levels of IL-6, TNF-α, VIP, 5-HT, ILβ and LBP. J, K: Thymus index and spleen index. L: HE staining of the liver tissues in each group (Original magnification: ×100). M: HE staining of rat hippocampus (×100). N: HE staining of hippocampal CA1 region of the rats (×200) . *P<0.05, **P<0.01, ***P<0.001 vs model group (CUMS); #P<0.05, ##P<0.01 vs control group.

图4 差异代谢物的鉴定及多元统计分析

Fig.4 Identification of differential metabolites and multivariate statistical analysis. A: Principal component analysis. B: Partial least squares discriminant analysis (OPLS-DA). C: Differential metabolite volcano plot analysis. The threshold of the difference was VIP≥1 and T-test P<0.05 in the OPLS-DA model.

图5 血清代谢物的影响及代谢通路分析

Fig.5 Effects of serum metabolites and metabolic pathway analysis. A: VIP (importance of variables in projection) diagram of OPLS-DA in POS mode. B: VIP diagram of OPLS-DA in NEG mode. C: KEGG enrichment circle diagram (CUMS vs JWJWXYP). D: KEGG enriched bubble diagram of the top 20 pathways with the smallest Q value. E: Metabolic pathway and classification between JWXYP and CUMS groups.

图6 肠道菌群物种丰度及差异物种

Fig.6 Intestinal flora species abundance and differential species. A: Distribution of samples in PC1 and PC2 dimensions. B: PCA analysis and PCoA analysis of the distribution of samples in PCo1 and PCo2 dimensions. C: β diversity box plot. D: Shaanon diversity index: ACE, Sob, Chao1 and Shannon index of Control, CIHMS and JWXYP groups. E: Distribution of Shannon index. F: Relative abundance of species at the phylum, class and species levels in control, CUMS and JWXYP groups.

图7 组内分析采用Spearman分析对加味逍遥丸干预CUMS诱导的抑郁相关成分、大鼠体内差异代谢物、差异肠道菌群

Fig.7 Spearman analysis of depression-related components, differential metabolites and differential intestinal flora in JWXYP-treated rats with CUMS-induced depression. P value and color depth represent the degree of correlation.*P<0.05, **P<0.01,***P<0.001, JW vs CUMS group.

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加味逍遥丸通过神经递质调节、抗炎抗氧化及肠道菌群调控改善大鼠的抑郁样行为

Jiawei Xiaoyao Pills improves depression-like behavior in rats by regulating neurotransmitters, inhibiting inflammation and oxidation and modulating intestinal flora

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