Dll4-Notch3信号对哮喘大鼠气道炎症及气道重塑的影响

发布时间：2018-04-28 23:04

本文选题：哮喘 + 肺组织　；参考：《桂林医学院》2017年硕士论文

【摘要】：目的:探讨Dll4-Notch3信号对哮喘大鼠气道炎症、气道重塑的影响。方法:选用SPF级雌性SD大鼠,雾化吸入OVA及用Notch3、Dll4两种抗体在体干预制作哮喘模型。哮喘模型成功后,测定每组大鼠气道阻力;常规HE染色运用图像分析软件分析气道壁的嗜酸性粒细胞(EOS)浸润情况;测定相关指标观察气道壁厚度(支气管管腔内周长、管壁面积、支气管平滑肌面积及平滑肌细胞核数);测定气道平滑肌向气道上皮基底膜迁移情况;免疫组织化学方法(IHC)观察各组大鼠气道壁Notch3、Dll4的表达情况;采用实时荧光定量PCR(RT-qPCR)、蛋白免疫印迹法(Western-blot)检测各组大鼠肺组织Notch3、Dll4 m RNA及蛋白的表达变化。结果:(1)(1)哮喘1周组与哮喘4周组大鼠的气道反应性、气道壁EOS浸润数量、气道壁壁厚相关指标都较正常对照组显著增高(P0.05),哮喘4周组较哮喘1周组上述指标显著增高(P0.05),且随致敏天数延长上述指标逐渐增高(P0.05)。(2)哮喘1周组与哮喘4周组相对于正常对照组,平滑肌距基底膜的距离减小(P0.05),哮喘4周组与哮喘1周组相比距离减小(P0.05)。(3)Notch3、Dll4在哮喘1周组、哮喘4周组的mRNA表达水平均高于正常对照组(P0.05),且此两组间有显著性差异(P0.05)。Western-blot及IHC所测结果基本同RT-qPCR结果一致。(2)(1)哮喘1周组、哮喘4周组的气道反应性、气道壁EOS浸润数量、气道壁壁厚相关指标高于正常对照组(P0.05),哮喘4周组的上述指标亦高于哮喘1周组(P0.05)。Notch3治疗1周组、Notch3治疗4周组的上述指标均高于正常对照组(P0.05),Notch3治疗4周组的上述指标高于Notch3治疗1周组(P0.05)。Dll4治疗1周组、Dll4治疗4周组的上述指标均高于正常对照组(P0.05),Dll4治疗4周组的上述指标高于Dll4治疗1周组(P0.05)。哮喘4周组上述指标高于Notch3治疗1周组和Notch3治疗4周组(P0.05),哮喘4周组上述指标高于Dll4治疗4周组,无组间对比意义(P0.05)。(2)哮喘1周组、哮喘4周组的支气管平滑肌距基底膜的距离较正常对照组减小(P0.05),哮喘4周组较哮喘1周组上述指标减小(P0.05)。Notch3治疗1周组、Notch3治疗4周组的以上指标较正常对照组减小(P0.05),Notch3治疗4周组的以上指标较Notch3治疗1周组减小(P0.05)。Dll4治疗1周组、Dll4治疗4周组的以上指标较正常对照组减小(P0.05),Dll4治疗4周组的以上指标较Dll4治疗1周组减小(P0.05)。哮喘4周组上述指标小于Notch3治疗1周组和Notch3治疗4周组(P0.05),哮喘4周组上述指标大于Dll4治疗4周组,无组间对比意义(P0.05)。(3)哮喘1周组、哮喘4周组Notch3、Dll4 mRNA表达水平均高于正常对照组(P0.05),哮喘4周组的Notch3、Dll4表达水平高于哮喘1周组(P0.05)。Notch3治疗1周组、Notch3治疗4周组的以上指标均高于正常对照组(P0.05),Notch3治疗4周组的以上指标高于Notch3治疗1周组(P0.05)。Dll4治疗1周组、Dll4治疗4周组的以上指标均高于正常对照组(P0.05),Dll4治疗4周组的以上指标高于Dll4治疗1周组(P0.05)。哮喘4周组上述指标高于Notch3治疗1周组和Notch3治疗4周组(P0.05),哮喘4周组上述指标高于Dll4治疗4周组,无组间对比意义(P0.05)。Western-blot及IHC所测结果基本同RT-qPCR结果一致。结论:Dll4在哮喘大鼠气道上皮细胞有特异性表达,Notch3在整个哮喘肺组织中呈高表达。抑制Dll4-Notch3信号可改善哮喘气道炎症及气道重构。表明Dll4-Notch3信号参与了哮喘的发病。
[Abstract]:Objective: To investigate the effect of Dll4-Notch3 signal on airway inflammation and airway remodeling in asthmatic rats. Methods: using SPF grade female SD rats, inhaling OVA and using Notch3, Dll4 two kinds of antibodies to make asthma model in vivo. After the asthma model was successful, the airway resistance was measured in each group of rats; the airway wall was analyzed by the routine HE staining software and the image analysis software was used to analyze the airway wall. The infiltration of eosinophil (EOS), the thickness of the airway wall (perimeter of the bronchus, the area of the tube wall, the area of the bronchial smooth muscle and the number of the nucleus of the smooth muscle of the bronchial smooth muscle), the migration of the airway smooth muscle to the basal membrane of the airway epithelium, and the expression of Notch3 and Dll4 in the airway wall of the rats were observed by the immunohistochemical method (IHC). The changes in the expression of Notch3, Dll4 m RNA and protein in lung tissues of each group were detected by real time fluorescence quantitative PCR (RT-qPCR) and protein immunoblotting (Western-blot). Results: (1) (1) the airway responsiveness of the 1 week asthma group and the 4 week group of asthma, the number of infiltration of the airway wall EOS and the wall thickness of the airway were significantly higher than that of the normal control group. High (P0.05), 4 weeks asthma group was significantly higher than that of the 1 week asthma group (P0.05), and the above index increased with the number of sensitization days (P0.05). (2) the distance between the 1 week asthma group and the 4 week asthma group compared with the normal control group, the distance of the smooth muscle from the basilar membrane decreased (P0.05), the 4 week asthma group was compared with the 1 week group of asthma (P0.05). (3) Notch3, Dll4 In the 1 week asthma group, the expression level of mRNA in the 4 week asthma group was higher than that in the normal control group (P0.05), and there was a significant difference between the two groups (P0.05) the results of.Western-blot and IHC were basically the same as the results of RT-qPCR. (1) the 1 weeks of asthma, the airway reactivity of the 4 week group of asthma, the number of EOS infiltration in the airway wall and the index of the wall thickness of the airway were higher than that of the normal. In the control group (P0.05), the above indexes in the 4 week asthma group were also higher than those in the 1 week group of asthma (P0.05) for 1 weeks, and the above indexes in the 4 week group were higher than those in the normal control group (P0.05). The above indexes in the 4 week group of Notch3 treatment were higher than those in the 1 week group of Notch3 (P0.05) for 1 weeks, and the above indexes in the 4 weeks group were all higher than those of the normal control. Group (P0.05), the above index in the 4 week group of Dll4 treatment was higher than that of the group of Dll4 for 1 weeks (P0.05). The above index in the 4 week group of asthma was higher than that in the 1 week group of Notch3 and 4 weeks in the Notch3 treatment group (P0.05). The above index in the 4 week group of asthma was higher than that in the 4 week group of Dll4 (P0.05). (2) the bronchial smooth muscle of the bronchial smooth muscle from the 1 week group of asthma and the 4 weeks of asthma was from the basilar membrane. The distance from the normal control group decreased (P0.05), the 4 week asthma group was less than that of the 1 week asthma group (P0.05) in the 1 week group, and the above index in the 4 week group decreased (P0.05). The above index in the 4 week group of Notch3 treatment was lower than that in the 1 week group of Notch3 (P0.05).Dll4 therapy 1 weeks, and Dll4 was used to treat the above indexes in the 4 week group. Compared with the normal control group (P0.05), the above index in the 4 week group of Dll4 treatment was less than that of the 1 week group (P0.05). The above indexes in the 4 week group of asthma were less than that of the 1 weeks group and the 4 week group (P0.05), the 4 week asthma group was more than the 4 week group with Dll4 treatment, and there was no comparison between the groups (P0.05). (3) the asthmatic 1 weeks group, 4 weeks of asthma, Notch3, Dll4 M The expression level of RNA was higher than that of the normal control group (P0.05). The expression of Notch3 and Dll4 in the 4 week asthma group was higher than that in the 1 week group of asthma (P0.05).Notch3 treatment 1 weeks, and the above indexes in the 4 week group of Notch3 treatment were higher than that of the normal control group (P0.05). The above index in the 4 week group of Notch3 treatment was higher than that of the 1 week group of Notch3 treatment (P0.05) for 1 weeks, and 4 weeks for the treatment. The above index in the group was higher than that of the normal control group (P0.05). The above index in the 4 week group of Dll4 treatment was higher than that of the 1 week group (P0.05). The above indexes in the 4 week group of asthma were higher than that in the 1 week group of Notch3 and the 4 week group of Notch3 (P0.05). The above indexes in the 4 week group of asthma were higher than those in the 4 week group with Dll4, and there was no comparison significance (P0.05).Western-blot and IHC. The results are basically the same as the results of RT-qPCR. Conclusion: Dll4 has a specific expression in the airway epithelial cells of asthmatic rats, and Notch3 is highly expressed in the lung tissue of the whole asthma. The inhibition of Dll4-Notch3 signal can improve the airway inflammation and airway remodeling in asthma. It indicates that the Dll4-Notch3 signal is involved in the onset of asthma.

【学位授予单位】：桂林医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R562.25

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