神经源性和白三烯依赖性炎症在上下气道过敏性疾病中的作用研究

Roles of neurogenic and leukotriene-dependent inflammation in upper and lower airway allergic diseases

目的探讨神经源性和白三烯依赖性炎症在上下气道过敏性疾病中的作用。方法选择雌性BALB/c小鼠39只,采用随机数字表法将小鼠分为单纯上气道变应性疾病(SU)小鼠、单纯下气道变应性疾病(SL)小鼠、联合上下气道变应性疾病(CUL)小鼠各12只和对照组小鼠3只。将SU、SL和CUL小鼠再分为模型组、神经肽P物质受体拮抗剂干预(NI)组、白三烯受体拮抗剂干预(LI)组和神经肽P物质受体拮抗剂与白三烯受体拮抗剂混合干预(NLI组),每组3只,合计12组,均给予腹腔注射致敏液400μL,并分别予以鼻腔、气管及鼻腔与气管联合滴入激发液建立相应动物模型。9个药物干预组再分别予神经肽P物质受体拮抗剂、孟鲁司特钠及受体拮抗剂与孟鲁司特钠联合行药物干预。分组后第33天对13组小鼠灌注获得鼻腔灌洗液(NLF)和肺泡灌洗液(BALF),随后过量麻醉处死并取出鼻黏膜和肺组织。采用HE染色观察小鼠鼻黏膜和肺组织嗜酸性粒细胞浸润情况,免疫组化检测小鼠鼻黏膜和肺组织中半胱氨酸白三烯受体1(CysLTR1)和神经肽P物质受体(NPR)的表达水平。采用ELISA法评估NLF和BALF中半胱氨酸白三烯(CysLTs)、神经营养因子和神经递质的水平。结果与对照组比较,SU-模型组、CUL-模型组在鼻黏膜和肺组织均可见嗜酸性粒细胞明显浸润,而SL-模型组在肺组织中可见嗜酸性粒细胞浸润,NPR和CysLTR1在3个模型组的鼻黏膜和肺组织中表达水平高于对照组。在NLF中,SU-模型组、CUL-模型组CysLTs、脑源性神经营养因子(BDNF)、神经生长因子(NGF)、降钙素基因相关肽(CGRP)、神经激肽A(NKA)、神经激肽B(NKB)和神经肽P物质(NP)水平高于对照组,SU-NI组、SU-LI组、SU-NLI组CysLTs、BDNF、NGF、CGRP、NKA和NP水平低于SU-模型组,SU-NLI组CGRP、NP水平低于SU-NI组、SU-LI组,SL-模型组小鼠NP水平高于对照组,SL-NI组、SL-LI组和SL-NLI组NP水平低于SL-模型组,CUL-NI组、CUL-LI组和CUL-NLI组BDNF和NGF水平低于CUL-模型组,CUL-LI组和CUL-NLI组CGRP和NKA水平低于CUL-模型组,CUL-NLI组NKA水平低于CUL-LI组,CUL-NI组、CUL-LI组和CUL-NLI组NKB低于CUL-模型组,差异均有统计学意义(均P<0.05)。在BALF中,SL-模型组、CUL-模型组CysLTs水平高于对照组,SL-模型组NGF、CGRP、NKA、NP水平高于对照组,CUL-模型组CGRP水平高于对照组,SL-NI组、SL-LI组和SL-NLI组CysLTs、NGF、CGRP、NKA和NP水平均低于SL-模型组,SL-NLI组CysLTs水平低于SL-NI组和SL-LI组,CUL-NI组、CUL-LI组和CUL-NLI组的CGRP水平低于CUL-模型组,差异均有统计学意义(均P<0.05)。结论在过敏性疾病中,上气道炎症可以通过神经源性炎症和白三烯依赖性炎症引起下气道炎症,而下气道炎症几乎不能影响上气道。NP受体或白三烯受体的拮抗作用有助于缓解气道炎症性疾病。

Objective To explore the roles of neurogenic and leukotriene-dependent inflammation in upper and lower airway allergic diseases.Methods Thirty nine female BALB/c mice were randomly divided into the upper airway allergic disease(SU)group(n=12),lower airway allergic disease(SL)group(n=12),combined upper and lower airway allergic disease(CUL)group(n=12)and control group(n=3).The upper,lower and combined upper and lower airway allergic disease models were induced by intraperitoneal injection of ovalbumin(OVA),the nasal and tracheal administration of OVA,and both intraperitoneal and nasal trachea administration of OVA,respectively.Each of the three allergic mice groups were further divided into model group,NI group,LI group and NLI group with 3 mice in each,and the last 3 groups were treated with substance p receptor antagonist(NI),leukotriene receptor antagonist(LI)and NI plus LI,respectively.On the d33 animals were sacrificed,the nasal lavage fluid(NLF)and bronchoalveolar lavage fluid(BALF)samples and the nasal mucosa and lung tissue samples were taken.Eosinophilic infiltration in nasal mucosa and lung tissue was observed by HE staining;the expression of CysLTR1 and NPR in the nasal mucosa and lungs was detected with immunohistochemistry;the levels of Cysteine leukotrienes,neurotrophic factors and neurotransmitters in NLF and BALF were measured by ELISA.Results Compared with the control group,the SU-model group and CUL-model group showed increased infiltration of eosinophils in the nasal mucosa and lungs,while eosinophils infiltration was observed in lung tissue of SL-model group.The expressions of NPR and CysLTR1 in nasal mucosa and lung tissue of the 3 model groups were higher than those of the control group.The levels of CysLTs,BDNF,NGF,CGRP,NKA,NKB and NP in NLF of SU-model and CU-model groups were higher than those in control group;while the levels in SU-NI,SU-LI and SU-NLI groups were lower than those in SU-model group.The levels of CGRP and NP in SU-NLI group were lower than those in SU-NI group and SU-LI group,the levels of NP in SL-model group were higher than those in control group,and the levels of NP in SL-NI,SL-LI and SL-NLI groups were lower than those in SL-model group.The levels of BDNF and NGF in CUL-NI,CUL-LI and CUL-NLI groups were lower than those in CUL-model group,the levels of CGRP and NKA in CUL-LI group and CUL-NLI group were lower than those in CUL-model group,and the levels of NKA in CUL-NLI group were lower than those in CUL-LI group.The NKB of CUL-NI,CUL-LI and CUL-NLI groups was lower than that of CUL-model group(all P<0.05).The NLF levels of CysLTs in SL-model and CULmodel groups were higher than those in control group;the levels of NGF,CGRP,NKA and NP in SL-model group were higher than that in control group;the level of CGRP in CUL-model group was higher than that in control group.The levels of CysLTs,NGF,CGRP,NKA and NP in SL-NI,SL-LI and SL-NLI groups were lower than those in SL-model group,and the levels of CysLTs in SL-NLI group were lower than those in SL-NI and SL-LI groups.The levels of CGRP in CUL-NI,CUL-LI and CUL-NLI groups were lower than those in CUL-model group(all P<0.05).Conclusion Upper airway inflammation can cause lower airway inflammation through neurogenic inflammation and leukotriene-dependent inflammation in allergic diseases,while lower airway inflammation hardly affects the upper airway.The intervention with neuropeptide receptors or leukotriene receptors antagonists may be able to treat airway inflammatory diseases.