Protective Effect and Mechanism of Artemitin on Ozone-Induced Pulmonary Inflammation in Mice

doi:10.12300/j.issn.1674-5817.2026.003

Abstract

Abstract:

Objective To investigate the protective effect of artemitin against ozone (O₃)-induced pulmonary inflammation in mice and to explore its underlying mechanism. Methods Thirty-two specific pathogen-free (SPF) male C57BL/6 mice were randomly divided into a blank control group, an artemitin-only control group, an O₃-induced model group, and an artemitin-treated group, with 8 mice in each group. In the artemitin-treated group, animals were injected intraperitoneally with artemitin (20 mg/kg) daily, followed by a 1 h exposure to 0.000 25% O₃. The artemitin-only control group received the same artemitin injection but was exposed to clean air. The O₃-induced model group was injected intraperitoneally with an equal volume of normal saline daily, followed by a 1-h exposure to 0.000 25% O₃. The blank control group received the saline injection and was exposed to clean air. All treatments were administered for 7 consecutive days. Pulmonary function indices of the mice in each group were measured using a whole-body plethysmography system. The levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in bronchoalveolar lavage fluid (BALF) were detected by enzyme-linked immunosorbent assay (ELISA). Hematoxylin-eosin (HE) staining was used to observe the histopathological changes in the lung tissue, and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was employed for untargeted metabolomic analysis to examine the changes in overall metabolic pathways in plasma. Results Compared with the O₃-induced model group, artemitin treatment significantly improved O₃-induced lung function impairments (P<0.05), lowered the levels of TNF-α and IL-1β in BALF (P<0.05) and alleviated inflammatory cell infiltration and structural damage in lung tissues. Metabolomics results indicated that artemitin treatment compensatorily enhanced energy supply and oxidized lipid signaling, while inhibiting purine metabolism and tryptophan-derived pathways. Conclusion Artemitin exhibits a significant protective effect against O₃-induced pulmonary inflammation in mice. It corrects upstream metabolic disorders by synergistically modulating tryptophan metabolism, energy metabolism, and lipid inflammatory mediators, thereby inhibiting downstream inflammatory responses and airway hyperresponsiveness. This study not only reveals the pleiotropic mechanism of artemitin in treating airway inflammation but also provides potential metabolic biomarkers and a theoretical basis for its clinical translation.

Key words: Ozone, Artemitin, Pulmonary inflammation, Tumor necrosis factor-α, Interleukin-1β, C57BL/6 mice

CLC Number:

R-332
R563

LI Yanping,HUANG Shan,YAO Jengyuan,et al. Protective Effect and Mechanism of Artemitin on Ozone-Induced Pulmonary Inflammation in Mice[J]. Laboratory Animal and Comparative Medicine, 2026, 46(5): 640-649. DOI: 10.12300/j.issn.1674-5817.2026.003.

Figures/Tables 5

Figure 1 Experimental flowchart

Note： RR， respiratory rate； TV， tidal volume； MV， minute ventilation； Penh， enhanced pause； PAU， pause； PIF， peak inspiratory flow； PEF， peak expiratory flow； Ti， inspiratory time； Te， expiratory time； EF₅₀， expiratory flow at 50% tidal volume； TNF-α， tumor necrosis factor -α； IL-1β， interleukin-1β.

Figure 2 Pulmonary function indicators and levels of inflammatory factors TNF-<strong>α</strong> and IL-1<strong>β</strong> in bronchoalveolar lavage fluid of mice

Note： A-J， Detection results of various pulmonary function indices （Penh， PAU， Te， MV， EF₅₀， PEF， RR， TV， PIF， Ti） of mice in the blank control group， artemitin-only control group， model group， and artemitin-treated group； K-L， Levels of inflammatory factors TNF-α and IL-1β in bronchoalveolar lavage fluid of mice in the blank control group， artemitin-only control group， model group， and artemitin-treated group. The artemitin-treated group was first intraperitoneally injected with artemitin （20 mg/kg） each day， and then continuously exposed to 0.000 25% O₃ for 1 h； the artemitin-only control group was first intraperitoneally injected with artemitin （20 mg/kg） each day， and then exposed to clean air； the O₃-induced model group was first intraperitoneally injected with an equal volume of normal saline each day， and then exposed to 0.000 25% O₃ for 1 h； the blank control group was first injected with an equal volume of normal saline each day， and then exposed to clean air； all the above groups were continuously treated for 7 d. Penh， enhanced pause； PAU， pause； Te， expiratory time； MV， minute ventilation； EF₅₀， expiratory flow at 50% tidal volume； PEF， peak expiratory flow； RR， respiratory rate； TV， tidal volume； PIF， peak inspiratory flow； Ti， inspiratory time； TNF-α， tumor necrosis factor-α； IL-1β， interleukin-1β. n=8； ^ns P>0.05， ^** P<0.01， ^*** P<0.001， ^**** P<0.000 1.

Figure 3 Results of HE staining of lung tissues in mice

Note： The artemitin-treated group was first intraperitoneally injected with artemitin （20 mg/kg） each day， and then continuously exposed to 0.000 25% O₃ for 1 h； the artemitin-only control group was first intraperitoneally injected with artemitin （20 mg/kg） each day， and then exposed to clean air； the O₃-induced model group was first intraperitoneally injected with an equal volume of normal saline each day， and then exposed to 0.000 25% O₃ for 1 h； the blank control group was first injected with an equal volume of normal saline each day， and then exposed to clean air； all the above groups were continuously treated for 7 d.

Figure 4 Multivariate statistical and KEGG enrichment analysis in metabolomics

Note： A， PCA score plots of plasma from the blank control， O₃-induced model， and artemitin-treated groups in ESI⁺ mode； B， PCA score plots of plasma from the blank control， O₃-induced model， and artemitin-treated groups in ESI⁻ mode； C， OPLS-DA score plots of plasma from the O₃-induced model and artemitin-treated groups in ESI⁺ mode； D， OPLS-DA score plots of plasma from the O₃-induced model and artemitin-treated groups in ESI⁻ mode； E， Volcano plot of differential metabolites between the O₃-induced model and artemitin-treated groups； F， Top 10 enriched KEGG pathways of differential metabolites. ESI， electrospray ionization； PCA， principal component analysis； OPLS-DA， orthogonal partial least squares discriminant analysis； FC， fold change. n=8.

Table 1

Differential metabolites in plasma between the artemitin-treated group and the O₃-induced model group

序号No.	质荷比 m/z	化合物 Compound	KEGG编号 KEGG ID	HMDB编号 HMDB ID	log₂FC	-log₁₀ P	AUC	保留时间/min Retention time/min	离子化模式 Ionization mode
1	103.040 818 5	3-Hydroxybutyric acid	C01089	HMDB0000011	1.8	10.1	1.00	0.76	negative
2	361.203 087 3	Cortisol	C00735	HMDB0000063	2.4	7.3	1.00	1.08	negative
3	363.218 000 8	Leukotriene A4	C00909	HMDB0001337	3.2	6.8	1.00	1.34	negative
4	160.077 502 2	Phenylacetaldehyde	C00601	HMDB0006236	-2.2	6.0	0.98	0.93	negative
5	307.115 643 5	Adenosine	C00212	HMDB0000050	-2.0	7.8	0.97	0.77	negative
6	160.041 370 6	Quinoline-4,8-diol	C05637	HMDB0060289	-1.2	6.2	0.97	0.90	negative
7	131.048 605 8	N1-Acetyl-N2-formyl-5-methoxykynuramine	C05642	HMDB0004259	-1.6	5.6	0.96	0.94	positive
8	496.274 317 6	Taurocholic acid	C05122	HMDB0000036	1.7	4.6	0.95	3.07	negative
9	311.224 870 9	12,13-EpOME	C14826	HMDB0004702	1.0	5.2	0.95	3.25	negative
10	360.071 196 1	N-Glycolylneuraminic acid	C03410	HMDB0000833	-2.7	5.1	0.93	0.87	negative
11	283.083 160 1	6-Hydroxymelatonin	C05643	HMDB0004081	2.0	4.6	0.95	0.80	negative
12	235.083 009 7	5-Formiminotetrahydrofolic acid	C00664	HMDB0001534	1.8	7.3	0.95	0.77	negative
13	147.012 513 4	Oxoglutaric acid	C00026	HMDB0000208	2.1	3.8	0.89	8.24	positive
14	187.008 039 0	3-Fumarylpyruvate	C02514	HMDB0060371	1.5	4.0	0.86	0.84	negative
15	208.062 677 7	Homogentisic acid	C00544	HMDB0000130	1.5	3.8	0.82	0.77	negative

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