欧前胡素对支气管哮喘小鼠气道重塑的改善作用及其机制

doi:10.13481/j.1671-587X.20250607

摘要/Abstract

摘要：

目的探讨欧前胡素（IMP）对支气管哮喘小鼠气道重塑的影响，并阐明其可能的机制。方法 40只SFP级雄性BALB/c小鼠随机分为对照组、模型组、低剂量IMP组（IMP-L组）、高剂量IMP组（IMP-H组）和地塞米松组，每组8只。除对照组外，其余各组小鼠腹腔注射卵清蛋白（OVA）混悬液构建小鼠哮喘模型。造模1周后观察各组小鼠每日哮喘症状并进行评分，8周后检测各组小鼠增强呼吸间歇（Penh）值以评估气道反应性，流式细胞术检测各组小鼠肺泡灌洗液（BALF）中嗜酸性粒细胞百分率，酶联免疫吸附试验（ELISA）法检测各组小鼠血清IgE和BALF中白细胞介素（IL）-13、IL-5、IL-4及γ干扰素（IFN-γ）水平，HE、PAS和Masson染色法观察各组小鼠肺组织病理形态表现、杯状细胞数量及胶原沉积情况，免疫组织化学染色法观察各组小鼠肺组织中α-平滑肌肌动蛋白（α-SMA）和无翅型小鼠乳腺肿瘤病毒（MMTV）整合位点家族成员5A（Wnt5A）蛋白表达情况，Western blotting法检测各组小鼠肺组织中Wnt5A、细胞髓细胞瘤基因（c-Myc）、β-连环蛋白（β-catenin）和α-SMA蛋白表达水平，免疫荧光法检测各组小鼠肺组织中α-SMA蛋白表达水平。结果与对照组比较，模型组小鼠哮喘症状评分升高（P<0.01），Penh值明显升高（P<0.01），血清IgE水平及BALF中嗜酸性粒细胞百分率和IL-13、IL-5及IL-4水平均明显升高（P<0.05或P<0.01），IFN-γ水平明显降低（P<0.05），肺组织中α-SMA和Wnt5A蛋白表达水平明显升高（P<0.01），Wnt/β-catenin通路相关蛋白表达明显升高（P<0.01）；免疫荧光法检测，肺组织中α-SMA蛋白表达水平明显升高（P<0.01）。与模型组比较，IMP-L组、IMP-H组和地塞米松组小鼠哮喘症状评分降低（P<0.01），IMP-H组小鼠Penh值降低（P<0.05），IMP-L组、IMP-H组和地塞米松组小鼠血清IgE水平及BALF中嗜酸性粒细胞百分率和IL-13、IL-5及IL-4水平降低（P<0.05或P<0.01），IFN-γ水平明显升高（P<0.05），肺组织中α-SMA和Wnt5A蛋白表达水平降低（P<0.05或P<0.01），Wnt/β-catenin通路相关蛋白表达水平降低（P<0.05或P<0.01）；免疫荧光法检测，肺组织α-SMA蛋白表达水平明显降低（P<0.05或P<0.01）。结论 IMP对支气管哮喘小鼠气道重塑具有改善作用，并可抑制Wnt/β-catenin通路相关蛋白表达水平。

关键词: 欧前胡素, 哮喘, 气道重塑, Wnt/β-连环蛋白信号通路, α-平滑肌肌动蛋白

Abstract:

Objective To explore the effects of imperatorin （IMP） on airway remodeling in the bronchial asthma mice， and to elucidate the possible mechanisms. Methods Forty SFP male BALB/c mice were randomly divided into control group， model group， low dose of IMP group （IMP-L group）， high dose of IMP group （IMP-H group） and dexamethasone group， with 8 mice in each group. Except for contol group， the mice in the other groups were injected with an ovalbumin （OVA） suspension intraperitoneally to induce the asthma models. After one week， the daily asthma symptoms of the mice were observed and scored. After 8 weeks， the enhanced pause（Penh） values of the mice in various groups were detected to evaluate the airway reactivities. The percentages of eosinophils in the bronchoalveolar lavage fluid （BALF） of the mice in various groups were detected by flow cytometry. The levels of serum IgE， interleukin interferon-gamma（IL）-13， IL-5， IL-4 and interferon-gamma（IFN-γ） in BALF of the mice in various groups were measured by enzyme-linked immunosorbent assay （ELISA） method. HE， PAS and Masson staining were applied to observe the pathomorphology， the number of goblet cells and collagen deposition of the lung tissue of the mice in various groups.Immunohisto chemistry method was applied to detect the expressions of α-smooth muscle actin （α-SMA） and mouse mammary tumor virus（MMTV） wingless type MMTV intergration site family member 5A （Wnt5A） proteins in lung tissue of the mice in various groups. The expression levels of Wnt5A， cellular myelocytomatosis oncogene （c-Myc）， β-catenin and α-SMA in lung tissue of the mice in various groups were detected by Western blotting method. The expression levels of α-SMA protein in lung tissue of the mice in various groups were detected by immunofluorescence method. Results Compared with control group， the score of asthma symptoms of the mice in model group was increased （P<0.01）； the Penh value was significantly increased （P<0.01）； the serum IgE levels and the levels of IL-13， IL-5， IL-4 in BALF， as well as the percentage of eosinophils （EOS） in BALF were significantly increased （P<0.05 or P<0.01）， and the level of IFN-γ was reduced （P<0.05）； the expression levels of α-SMA and Wnt5A proteins in lung tissue were markedly increased （P<0.01）； the expression levels of proteins associated with the Wnt/β-catenin signaling pathway in the lung tissue were significantly increased （P<0.01）； the immofluorescence method results showed the expression level of α-SMA protein in lung tissue was significantly increased （P<0.01）. Compared with model group， the scores of asthma symphtoms of the mice in IMP-L group， IMP-H group， and dexamethasone group were decereased （P<0.01）， and the Penh values of the mice in IMP-H group were decreased （P<0.05）； the serum IgE levels and the levels of IL-13， IL-5， IL-4 in BALF， as well as the percentages of EOS in BALF of the mice in IMP-L group， IMP-H group， and dexamethasone group were decreased （P<0.05 or P<0.01）， and the levels of IFN-γ were increased （P<0.05）； the expression levels α-SMA and Wnt5A proteins in lung tissue were decreased （P<0.05 or P<0.01）； the expression levels of proteins related to the Wnt/β-catenin signaling pathway in the lung tissue were decreased （P<0.05 or P<0.01）； the immunofluorescence method results showed that expression levels of the α-SMA protein in the lung tissue were reduced （P<0.05 or P<0.01）. Conclusion IMP has an improving effect on airway remodeling in the asthmatic mice and can inhibit the expression levels of Wnt/β-catenin pathway-related proteins.

Key words: Imperatorin, Asthma, Airway remodeling, Wnt/β-catenin signaling pathway, α-smooth muscle actin

中图分类号:

R562.25

阎煜,黄丹丹,洪春玲,魏博雯,杨媛媛,延光海,宋艺兰,咸哲民. 欧前胡素对支气管哮喘小鼠气道重塑的改善作用及其机制[J]. 吉林大学学报(医学版), 2025, 51(6): 1508-1517.

Yu YAN,Dandan HUANG,Chunling HONG,Bowen WEI,Yuanyuan YANG,Guanghai YAN,Yilan SONG,Zhemin XIAN. Improvement effect of imperatorin on airway remodeling in bronchial asthma mice and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2025, 51(6): 1508-1517.

图/表 12

图1

图2

图3

图4

表1

表2

图5

图6

图7

图8

图9

图10

参考文献 24

[1]	SAVIN I A， ZENKOVA M A， SEN’KOVA A V. Bronchial asthma， airway remodeling and lung fibrosis as successive steps of one process［J］. Int J Mol Sci， 2023， 24（22）： 16042.
[2]	刘健. 支气管哮喘气道重塑机制的研究进展［J］. 中国现代医学杂志， 2022， 32（12）： 51-54.
[3]	TIOTIU A， STEIROPOULOS P， NOVAKOVA S， et al. Airway remodeling in asthma： mechanisms， diagnosis， treatment， and future directions［J］. Arch De Bronconeumología， 2025， 61（1）： 31-40.
[4]	HAYAT R， MANZOOR M， HUSSAIN A. Wnt signaling pathway： a comprehensive review［J］. Cell Biol Int， 2022， 46（6）： 863-877.
[5]	LIU Y L， LI J N， CHEN R C， et al. Airway epithelial cells promote in vitro airway smooth muscle cell proliferation by activating the Wnt/β-catenin pathway［J］. Respir Physiol Neurobiol， 2025， 331： 104368.
[6]	HUSSAIN M， XU C Y， LU M P， et al. Wnt/β-catenin signaling links embryonic lung development and asthmatic airway remodeling［J］. Biochim Biophys Acta BBA Mol Basis Dis， 2017， 1863（12）： 3226-3242.
[7]	DENG M， XIE L， ZHONG L， et al. Imperatorin： a review of its pharmacology， toxicity and pharmacokinetics［J］. Eur J Pharmacol， 2020， 879： 173124.
[8]	李晓强，谭余庆，李慧杰，等. 欧前胡素药理作用及机制研究进展［J］. 中国实验方剂学杂志， 2020， 26（18）： 196-201.
[9]	LIU F， SHANG Y X. Sirtuin 6 attenuates epithelial-mesenchymal transition by suppressing the TGF-β1/Smad3 pathway and c-Jun in asthma models［J］. Int Immunopharmacol， 2020， 82： 106333.
[10]	何婷，钱佩瑶，洪敏，等.诱发支气管哮喘动物模型气道重塑特征的方法和评价［J］. 中国实验动物学报， 2022， 30（1）： 117-123.
[11]	杨昆，龚新月，伍文彬，等. 三子养亲汤对支气管哮喘模型大鼠的作用机制研究［J］. 中药药理与临床， 2018， 34（2）： 21-24.
[12]	袁欣，王安琪，王思宇，等. 痰瘀互结证哮喘大鼠模型的建立及评价［J］. 中国实验动物学报， 2024， 32（10）： 1243-1251.
[13]	HABIB N， PASHA M A， TANG D D. Current understanding of asthma pathogenesis and biomarkers［J］. Cells， 2022， 11（17）： 2764.
[14]	HAMMAD H， LAMBRECHT B N. The basic immunology of asthma［J］. Cell， 2021， 184（6）： 1469-1485.
[15]	LUO W H， HU J D， XU W F， et al. Distinct spatial and temporal roles for Th1， Th2， and Th17 cells in asthma［J］. Front Immunol， 2022， 13： 974066.
[16]	JI T F， LI H Q. T-helper cells and their cytokines in pathogenesis and treatment of asthma［J］. Front Immunol， 2023， 14： 1149203.
[17]	KOOPMANS T， HESSE L， NAWIJN M C， et al. Smooth-muscle-derived WNT5A augments allergen-induced airway remodelling and Th2 type inflammation［J］. Sci Rep， 2020， 10： 6754.
[18]	YAO L， ZHAO H， TANG H， et al. Blockade of β-catenin signaling attenuates toluene diisocyanate-induced experimental asthma［J］. Allergy， 2017， 72（4）： 579-589.
[19]	ZHANG T， HUANG H， LIANG L H， et al. Long non-coding RNA （LncRNA） non-coding RNA activated by DNA damage （NORAD） knockdown alleviates airway remodeling in asthma via regulating miR-410-3p/RCC2 and inhibiting Wnt/β-catenin pathway［J］. Heliyon， 2024， 10（1）： e23860.
[20]	FANG Y N， JIN W W， GUO Z， et al. Quercetin alleviates asthma-induced airway inflammation and remodeling through downregulating periostin via blocking TGF-β1/smad pathway［J］. Pharmacology， 2023， 108（5）： 432-443.
[21]	XU T T， WU Z Z， YUAN Q， et al. Proline is increased in allergic asthma and promotes airway remodeling［J］. JCI Insight， 2023， 8（16）： e167395.
[22]	SONG Y L， WANG Z G， JIANG J Z， et al. DEK-targeting aptamer DTA-64 attenuates bronchial EMT-mediated airway remodelling by suppressing TGF-β1/Smad， MAPK and PI3K signalling pathway in asthma［J］. J Cellular Molecular Medi， 2020， 24（23）： 13739-13750.
[23]	REN Y， ZHONG X H， WANG H Y， et al. Chloroquine attenuates asthma development by restoring airway smooth muscle cell phenotype via the ROS-AKT pathway［J］. Front Pharmacol， 2022， 13： 916508.
[24]	BRADDING P， PORSBJERG C， CÔTÉ A， et al. Airway hyperresponsiveness in asthma： The role of the epithelium［J］. J Allergy Clin Immunol， 2024， 153（5）： 1181-1193.

Group	Total IgE	OVA specific IgE
Control	95.03±2.51	2.16±1.34
Model	389.36±9.46^*	193.24±3.05^*
IMP-L	309.57±8.08^△	178.32±3.52^△
IMP-H	295.10±6.88^△	176.48±3.95^△
Dexamethasone	281.51±6.56^△	170.44±6.25^△

Group	IL-13	IL-5	IL-4	IFN-γ
Control	23.37±2.99	20.70±2.51	15.61±1.52	82.24±8.41
Model	85.24±9.01^*	63.69±6.84^*	79.91±8.99^*	20.61±2.64^*
IMP-L	64.82±7.51^△	44.90±8.55^△	60.82±8.08^△	40.30±2.64^△
IMP-H	46.95±9.01^△	35.60±6.01^△	41.61±8.53^△	61.25±4.40^△
Dexamethasone	37.10±7.23^△	28.85±4.16^△	32.77±6.80^△	71.18±2.59^△