Yiqi Zishen Formula ameliorates inflammation in mice with chronic obstructive pulmonary disease by inhibiting the PI3K/Akt/NF-κB signaling pathway

doi:10.12122/j.issn.1673-4254.2025.07.07

Abstract

Abstract:

Objective To investigate pharmacologically active components of Yiqi Zishen Formula (YZF) and their mechanisms for alleviating airway inflammation in mice with chronic obstructive pulmonary disease (COPD). Methods Ultra-high-performance liquid chromatography coupled with quadrupole-orbitrap mass spectrometry was employed to characterize the chemical components in YZF and YZF-medicated rat serum. A compound-disease target network was constructed based on serum components of YZF to screen the key pathways and targets using enrichment analysis. A mouse model of cigarette smoke-induced COPD was used to evaluate the anti-inflammatory effect of YZF and validate the expression of key proteins in network pharmacology-enriched pathways. Fifty male C57BL/6J mice were randomized equally into control group, COPD model group, high- and low-dose YZF treatment groups, and N-acetylcysteine treatment group. Pulmonary function of the mice was assessed using whole-body plethysmography, and lung histopathology, alveolar structure, and airway remodeling were analyzed using HE staining. The levels of IL-1β, IL-6, and TNF‑α in bronchoalveolar lavage fluid (BALF) were determined with ELISA, and pulmonary expressions of PI3K, Akt, phosphorylated Akt (p-Akt), p65, and phosphorylated p65 (p-p65) were detected using immunohistochemistry. Results We identified a total of 156 chemical components (including 26 flavonoids or flavonoid glycosides, 27 alkaloids, and 11 saponins) in YZF and 43 prototype components in medicated rat serum. Network pharmacology revealed 704 YZF-related targets and 1199 COPD-associated targets. Integrated analysis suggested that the anti-COPD effects of YZF were associated with the PI3K-Akt signaling pathway. In mouse models of COPD, YZF treatment significantly increased mean alveolar number and peak expiratory flow (P<0.05), reduced mean linear intercept, bronchial wall thickness, lung coefficient, and BALF cytokine levels, and suppressed the expressions of PI3K, Akt, p-Akt, p65, and p-p65 in the lung tissues. Conclusion YZF alleviates COPD symptoms and airway inflammation in mice possibly by inhibiting the PI3K/Akt/NF‑κB pathway through its multiple components that interact with multiple targets.

Key words: Yiqi Zishen Formula, chronic obstructive pulmonary disease, serum-absorbed components, network pharmacology, inflammation

Liming WANG, Hongrui CHEN, Yan DU, Peng ZHAO, Yujie WANG, Yange TIAN, Xinguang LIU, Jiansheng LI. Yiqi Zishen Formula ameliorates inflammation in mice with chronic obstructive pulmonary disease by inhibiting the PI3K/Akt/NF-κB signaling pathway[J]. Journal of Southern Medical University, 2025, 45(7): 1409-1422.

Figures/Tables 10

References 34

[1]	Agustí A, Celli BR, Criner GJ, et al. Global initiative for chronic obstructive lung disease 2023 report: gold executive summary[J]. Eur Respir J, 2023, 61(4): 2300239. doi：10.1183/13993003.00239-2023
[2]	丁莉莉, 冯淬灵, 赵克明. 慢性阻塞性肺疾病中医研究进展[J]. 辽宁中医药大学学报, 2023, 25(3): 164-8.
[3]	李建生, 余学庆. 慢性阻塞性肺疾病中医分期分级防治策略[J]. 中医杂志, 2019, 60(22): 1895-9.
[4]	Wang MH, Li JS, Li SY, et al. Effects of comprehensive therapy based on traditional Chinese medicine patterns on older patients with chronic obstructive pulmonary disease: a subgroup analysis from a four-center, randomized, controlled study[J]. Front Med, 2014, 8(3): 368-75. doi：10.1007/s11684-014-0360-0
[5]	田燕歌, 李亚, 李建生, 等. 调补肺肾法对COPD大鼠JAK/STAT信号转导的影响及远后效应[J]. 中国中药杂志, 2013, 38(1): 75-80. doi：10.4268/cjcmm20130117
[6]	Qin YQ, Chen YL, Zhao P, et al. Tiaobu Feishen therapy inhibits inflammation induced by cigarette smoke extracts in a human monocyte/macrophage cell line[J]. J Tradit Chin Med, 2021, 41(3): 360-6. doi：10.19852/j.cnki.jtcm.2021.03.003
[7]	Dong HR, Liu XF, Zheng WC, et al. Three Tiaobu Feishen formulae reduces cigarette smoke-induced inflammation in human airway epithelial cells[J]. J Tradit Chin Med, 2020, 40(3): 386-92.
[8]	Chen YL, Wu YS, Li JS, et al. Three Tiaobu Feishen therapies protect human alveolar epithelial cells against cigarette smoking and tumor necrosis factor: induced inflammation by nuclear factor-kappa B pathway[J]. J Tradit Chin Med, 2019, 39(1): 45-9.
[9]	李高峰, 刘淑娟, 李亚, 等. 调补肺肾三法通过抑制ERK1/2信号通路改善COPD大鼠气道黏液高分泌[J]. 中国实验动物学报, 2024, 32(4): 411-22.
[10]	陈旺宇, 节笑笑, 陈甜甜, 等. 中药血清药物化学研究现状[J]. 中国医药科学, 2023, 13(4): 26-9, 86.
[11]	Arifin WN, Zahiruddin WM. Sample size calculation in animal studies using resource equation approach[J]. Malays J Med Sci, 2017, 24(5): 101-5. doi：10.21315/mjms2017.24.5.11
[12]	国家药典委员会. 中华人民共和国药典-三部: 2020年版[M]. 北京: 中国医药科技出版社, 2020.
[13]	Wu JY, Cai BR, Zhang A, et al. Chemical identification and antioxidant screening of Bufei yishen formula using an offline DPPH ultrahigh-performance liquid chromatography Q-extractive orbitrap MS/MS[J]. Int J Anal Chem, 2022, 2022: 1423801. doi：10.1155/2022/1423801
[14]	Shao D, Liu XG, Wu JY, et al. Identification of the active compounds and functional mechanisms of Jinshui Huanxian formula in pulmonary fibrosis by integrating serum pharmacochemistry with network pharmacology[J]. Phytomedicine, 2022, 102: 154177. doi：10.1016/j.phymed.2022.154177
[15]	梅晓峰, 赵鹏, 卢瑞龙, 等. 慢性阻塞性肺疾病小鼠模型制备方法的比较研究[J]. 中国病理生理杂志, 2022, 38(1): 178-85. doi：10.3969/j.issn.1000-4718.2022.01.023
[16]	Ye JB, Ren G, Li WY, et al. Characterization and identification of prenylated flavonoids from Artocarpus heterophyllus lam. roots by quadrupole time-of-flight and linear trap quadrupole orbitrap mass spectrometry[J]. Molecules, 2019, 24(24): 4591. doi：10.3390/molecules24244591
[17]	慢性阻塞性肺疾病中西医结合管理专家共识写作组, 何权瀛, 冯淬灵. 慢性阻塞性肺疾病中西医结合管理专家共识(2023版)[J]. 中国全科医学, 2023, 26(35): 4359-71.
[18]	徐砚通, 唐雅楠. 方剂“组-谱-效-机” 研究模式探讨[J]. 中国中药杂志, 2025, 50(3): 840-5. doi：10.19540/j.cnki.cjcmm.20241017.601
[19]	祁建宏, 董芳旭. 黄酮类化合物药理作用研究进展[J]. 北京联合大学学报, 2020, 34(3): 89-92. doi：10.16255/j.cnki.ldxbz.2020.03.014
[20]	Hosawi S. Current update on role of hesperidin in inflammatory lung diseases: chemistry, pharmacology, and drug delivery approaches[J]. Life (Basel), 2023, 13(4): 937. doi：10.3390/life13040937
[21]	Liu CL, Zhu XM, Aga EB, et al. Ebeiedinone and peimisine inhibit cigarette smoke extract-induced oxidative stress injury and apo-ptosis in BEAS-2B cells[J]. Cell Stress Chaperones, 2024, 29(6): 697-708. doi：10.1016/j.cstres.2024.10.001
[22]	李凡凡, 郑越超. 天然生物碱抗菌活性及机理研究[J]. 医学信息, 2021, 34(17): 49-52. doi：10.3969/j.issn.1006-1959.2021.17.013
[23]	Wang LN, Zhang YX, Song ZM, et al. Ginsenosides: a potential natural medicine to protect the lungs from lung cancer and inflammatory lung disease[J]. Food Funct, 2023, 14(20): 9137-66. doi：10.1039/d3fo02482b
[24]	Zeng JM, Liao SD, Liang Z, et al. Schisandrin A regulates the Nrf2 signaling pathway and inhibits NLRP3 inflammasome activation to interfere with pyroptosis in a mouse model of COPD[J]. Eur J Med Res, 2023, 28(1): 217. doi：10.1186/s40001-023-01190-8
[25]	Guo Q, Jin YZ, Chen XY, et al. NF-κB in biology and targeted therapy: new insights and translational implications[J]. Signal Transduct Target Ther, 2024, 9(1): 53. doi：10.1038/s41392-024-01757-9
[26]	Liu YH, Kong HB, Cai HP, et al. Progression of the PI3K/Akt signaling pathway in chronic obstructive pulmonary disease[J]. Front Pharmacol, 2023, 14: 1238782. doi：10.3389/fphar.2023.1238782
[27]	杨洋, 李竹英, 田春燕, 等. 中医药干预慢性阻塞性肺疾病相关信号通路的研究进展[J]. 中国实验方剂学杂志, 2024, 30(15): 244-55.
[28]	张钰, 涂星, 张燕, 等. 文王一支笔的抗炎作用机制: 基于网络药理学、分子对接及实验验证[J]. 南方医科大学学报, 2023, 43(3): 383-92. doi：10.12122/j.issn.1673-4254.2023.03.07
[29]	白莉敏, 吕行, 任引刚, 等. miR-22-3p通过激活PTEN/PI3K/AKT/NF-κB信号通路加重慢性阻塞性肺疾病[J]. 现代生物医学进展, 2023, 23(10): 1835-42.
[30]	Thongin S, Den-Udom T, Uppakara K, et al. Beneficial effects of capsaicin and dihydrocapsaicin on endothelial inflammation, nitric oxide production and antioxidant activity[J]. Biomed Pharmacother, 2022, 154: 113521. doi：10.1016/j.biopha.2022.113521
[31]	Dong Y, Zou YZ, Li T, et al. Schisandrol a alleviates allergic asthma in mice via regulating the NF-κB/IκBα and Nrf2/HO-1 signaling pathways[J]. J Med Food, 2025, 28(1): 28-37. doi：10.1089/jmf.2024.k.0117
[32]	Chen JM, Li M, Chen R, et al. Gegen Qinlian standard decoction alleviated irinotecan-induced diarrhea via PI3K/AKT/NF-κB axis by network pharmacology prediction and experimental validation combination[J]. Chin Med, 2023, 18(1): 46. doi：10.1186/s13020-023-00747-3
[33]	Kim JS, Yi HK. Schisandrin C enhances mitochondrial biogenesis and autophagy in C2C12 skeletal muscle cells: potential invo-lvement of anti-oxidative mechanisms[J]. Naunyn Schmiedebergs Arch Pharmacol, 2018, 391(2): 197-206. doi：10.1007/s00210-017-1449-1
[34]	Qin YQ, Yang JF, Li HB, et al. Recent advances in the therapeutic potential of nobiletin against respiratory diseases[J]. Phyto-medicine, 2024, 128: 155506. doi：10.1016/j.phymed.2024.155506

NO.	Compound	Retention time (min)	Formula	MS/MS	Error (ppm)
1	Synephrine	1.6	C₉H₁₃NO₂	150.0917	1.20
2	Histidinyl-asparagine	1.62	C₁₀H₁₃N₄O₃	192.0785; 120.0650; 103.0390	-5.53
3	Fructose-phenylalanine	1.62	C₁₅H₂₁NO₇	310.1298; 292.1188	-1.52
4	Chlorogenic acid	12.03	C₁₆H₁₈O₉	163.0389; 145.0285; 135.0439; 117.0336	-6.19
5	p-Coumaric acid O-glycoside (a)	12.69	C₁₅H₁₆O₈	163.0390	1.24
6	Schizonepetoside A	12.72	C₁₆H₂₆O₇	169.1223	-1.71
7	Neostemonine or its isomer (a)	13.15	C₁₈H₂₅NO₄	246.1492	0.50
8	Croomine	13.19	C₁₈H₂₇NO₄	248.1645	-1.09
9	Neostemonine or its isomer (b)	13.43	C₁₈H₂₅NO₄	246.1491	0.50
10	Verticine-N-Oxide	14.55	C₂₇H₄₅NO₄	430.3311	-1.44
11	Diosmetin-6-C-glucoside	15.06	C₂₂H₂₂O₁₁	445.1081; 397.0879; 367.0805	-1.22
12	Verticinone-N-Oxide or its isomer (a)	15.51	C₂₇H₄₃NO₄	428.3118; 410.3059	-1.60
13	Verticinone-N-Oxide or its isomer (b)	15.83	C₂₇H₄₃NO₄	428.3174; 410.3051	-1.60
14	Stemoninine or its isomer (a)	16.78	C₂₂H₃₁NO₅	372.2172; 316.0912	-2.08
15	Stemoninine or its isomer (b)	17.4	C₂₂H₃₁NO₅	372.2175; 316.0909	-3.26
16	Peimisine	17.75	C₂₇H₄₁O₃N	410.3039	-1.85
17	Meranzin	18	C₁₅H₁₆O₄	189.0547	-0.53
18	Peimine	19.18	C₂₇H₄₅O₃N	414.3372	-0.33
19	TuberostemonineK/Neotuberostemonine	19.73	C₂₂H₃₃NO₄	358.2334; 348.2545; 302.2115; 250.1802	0.21
20	Peiminine	21.25	C₂₇H₄₃NO₃	412.3218; 396.2900	-0.91
21	Peimine isomer	23.46	C₂₇H₄₅O₃N	414.3374	-0.75
22	Gomisin R	23.74	C₂₂H₂₄O₇	383.1506;	-1.28
23	Dihydrocapsaicin	23.76	C₁₈H₂₉NO₃	290.2121;262.2171; 192.1391;136.0762;122.0602	-0.90
24	Peiminine isomer	23.97	C₂₇H₄₃NO₃	412.3222	-2.26
25	Hapepunine	24.31	C₂₈H₄₇NO₂	412.3602	-1.44
26	Fritillarizine/Puqiedinone	24.4	C₂₇H₄₃NO₂	396.3281	-1.08
27	Gentrogenin or its isomer (a)	25.1	C₂₇H₄₀O₄	411.2920	5.86
28	Gomisin S	25.2	C₂₃H₃₀O₇	401.1963; 369.1695	-1.18
29	Gentrogenin or its isomer (b)	25.44	C₂₇H₄₀O₄	411.2888	-3.86
30	Methyl eugenol	26.14	C₁₁H₁₄O₂	151.0752; 116.0011	0.16
31	Schisandrol A	26.43	C₂₄H₃₂O₇	415.2115; 400.1886; 384.1931; 369.1700; 338.1512	-1.69
32	Nobiletin	26.65	C₂₁H₂₂O₈	388.1158; 373.0921; 358.0685; 355.0820; 327.0869; 211.0245	-3.12
33	Schisanhenol or its isomer (a)	28.39	C₂₃H₃₀O₆	388.1881; 371.1856; 340.1665; 325.1430	-2.42
34	Schisantherin A	28.85	C₃₀H₃₂O₉	415.1725; 268.9779; 91.0565	-6.24
35	Schisanhenol or its isomer (b)	28.87	C₂₃H₃₀O₆	388.1876; 371.1868; 340.1668; 325.1438	-2.42
36	(-)-Gomisin L2	28.87	C₂₂H₂₆O₆	371.1465	-1.69
37	9-Tetradecenoic acid	29.8	C₁₄H₂₆O₂	209.1927	-1.06
38	Schisandrin A	30.09	C₂₄H₃₂O₆	402.2043; 386.2090; 371.1852; 347.1491; 316.1307; 301.1073	-1.23
39	Schisandrin B	30.88	C₂₃H₂₈O₆	386.1725; 370.1776; 355.1534; 300.0990; 285.0758	-1.31
40	Wuweizisu C	31.53	C₂₂H₂₄O₆	285.0756; 257.0808; 227.0703; 212.0466	3.93
41	Gondoic acid or its isomer (a)	32.83	C₂₀H₃₈O₂	265.2524; 247.2425	-0.57
42	Gondoic acid or its isomer (b)	33.96	C₂₀H₃₈O₂	293.2838	-1.94
43	Gondoic acid or its isomer (c)	34.48	C₂₀H₃₈O₂	293.2824	-3.41

NO.	Compound	Retention time (min)	Formula	MS/MS	Error (ppm)
1	Synephrine	1.6	C₉H₁₃NO₂	150.0917	1.20
2	Histidinyl-asparagine	1.62	C₁₀H₁₃N₄O₃	192.0785; 120.0650; 103.0390	-5.53
3	Fructose-phenylalanine	1.62	C₁₅H₂₁NO₇	310.1298; 292.1188	-1.52
4	Chlorogenic acid	12.03	C₁₆H₁₈O₉	163.0389; 145.0285; 135.0439; 117.0336	-6.19
5	p-Coumaric acid O-glycoside (a)	12.69	C₁₅H₁₆O₈	163.0390	1.24
6	Schizonepetoside A	12.72	C₁₆H₂₆O₇	169.1223	-1.71
7	Neostemonine or its isomer (a)	13.15	C₁₈H₂₅NO₄	246.1492	0.50
8	Croomine	13.19	C₁₈H₂₇NO₄	248.1645	-1.09
9	Neostemonine or its isomer (b)	13.43	C₁₈H₂₅NO₄	246.1491	0.50
10	Verticine-N-Oxide	14.55	C₂₇H₄₅NO₄	430.3311	-1.44
11	Diosmetin-6-C-glucoside	15.06	C₂₂H₂₂O₁₁	445.1081; 397.0879; 367.0805	-1.22
12	Verticinone-N-Oxide or its isomer (a)	15.51	C₂₇H₄₃NO₄	428.3118; 410.3059	-1.60
13	Verticinone-N-Oxide or its isomer (b)	15.83	C₂₇H₄₃NO₄	428.3174; 410.3051	-1.60
14	Stemoninine or its isomer (a)	16.78	C₂₂H₃₁NO₅	372.2172; 316.0912	-2.08
15	Stemoninine or its isomer (b)	17.4	C₂₂H₃₁NO₅	372.2175; 316.0909	-3.26
16	Peimisine	17.75	C₂₇H₄₁O₃N	410.3039	-1.85
17	Meranzin	18	C₁₅H₁₆O₄	189.0547	-0.53
18	Peimine	19.18	C₂₇H₄₅O₃N	414.3372	-0.33
19	TuberostemonineK/Neotuberostemonine	19.73	C₂₂H₃₃NO₄	358.2334; 348.2545; 302.2115; 250.1802	0.21
20	Peiminine	21.25	C₂₇H₄₃NO₃	412.3218; 396.2900	-0.91
21	Peimine isomer	23.46	C₂₇H₄₅O₃N	414.3374	-0.75
22	Gomisin R	23.74	C₂₂H₂₄O₇	383.1506;	-1.28
23	Dihydrocapsaicin	23.76	C₁₈H₂₉NO₃	290.2121;262.2171; 192.1391;136.0762;122.0602	-0.90
24	Peiminine isomer	23.97	C₂₇H₄₃NO₃	412.3222	-2.26
25	Hapepunine	24.31	C₂₈H₄₇NO₂	412.3602	-1.44
26	Fritillarizine/Puqiedinone	24.4	C₂₇H₄₃NO₂	396.3281	-1.08
27	Gentrogenin or its isomer (a)	25.1	C₂₇H₄₀O₄	411.2920	5.86
28	Gomisin S	25.2	C₂₃H₃₀O₇	401.1963; 369.1695	-1.18
29	Gentrogenin or its isomer (b)	25.44	C₂₇H₄₀O₄	411.2888	-3.86
30	Methyl eugenol	26.14	C₁₁H₁₄O₂	151.0752; 116.0011	0.16
31	Schisandrol A	26.43	C₂₄H₃₂O₇	415.2115; 400.1886; 384.1931; 369.1700; 338.1512	-1.69
32	Nobiletin	26.65	C₂₁H₂₂O₈	388.1158; 373.0921; 358.0685; 355.0820; 327.0869; 211.0245	-3.12
33	Schisanhenol or its isomer (a)	28.39	C₂₃H₃₀O₆	388.1881; 371.1856; 340.1665; 325.1430	-2.42
34	Schisantherin A	28.85	C₃₀H₃₂O₉	415.1725; 268.9779; 91.0565	-6.24
35	Schisanhenol or its isomer (b)	28.87	C₂₃H₃₀O₆	388.1876; 371.1868; 340.1668; 325.1438	-2.42
36	(-)-Gomisin L2	28.87	C₂₂H₂₆O₆	371.1465	-1.69
37	9-Tetradecenoic acid	29.8	C₁₄H₂₆O₂	209.1927	-1.06
38	Schisandrin A	30.09	C₂₄H₃₂O₆	402.2043; 386.2090; 371.1852; 347.1491; 316.1307; 301.1073	-1.23
39	Schisandrin B	30.88	C₂₃H₂₈O₆	386.1725; 370.1776; 355.1534; 300.0990; 285.0758	-1.31
40	Wuweizisu C	31.53	C₂₂H₂₄O₆	285.0756; 257.0808; 227.0703; 212.0466	3.93
41	Gondoic acid or its isomer (a)	32.83	C₂₀H₃₈O₂	265.2524; 247.2425	-0.57
42	Gondoic acid or its isomer (b)	33.96	C₂₀H₃₈O₂	293.2838	-1.94
43	Gondoic acid or its isomer (c)	34.48	C₂₀H₃₈O₂	293.2824	-3.41

Ingredient	Degree	Closeness centrality	Betweenness centrality
Gomisin S	46	0.4087	0.1222
Schisandrin A	45	0.4026	0.1143
(-)-Gomisin L2	44	0.4011	0.1029
Wuweizisu C	38	0.3923	0.0869
Schisantherin A	35	0.3894	0.0765
Dihydrocapsaicin	34	0.3894	0.1012
Schisandrol A	34	0.3881	0.0748
Gentrogenin	33	0.3759	0.0782
9-Tetradecenoic acid	32	0.3881	0.0940
Hapepunine	31	0.3853	0.0848

Ingredient	Degree	Closeness centrality	Betweenness centrality
Gomisin S	46	0.4087	0.1222
Schisandrin A	45	0.4026	0.1143
(-)-Gomisin L2	44	0.4011	0.1029
Wuweizisu C	38	0.3923	0.0869
Schisantherin A	35	0.3894	0.0765
Dihydrocapsaicin	34	0.3894	0.1012
Schisandrol A	34	0.3881	0.0748
Gentrogenin	33	0.3759	0.0782
9-Tetradecenoic acid	32	0.3881	0.0940
Hapepunine	31	0.3853	0.0848