MiR-155-5p介导Treg/Th17免疫失衡致变应性鼻炎机制研究

The study on the mechanism of allergic rhinitis induced by miR-155-5p-mediated Treg/Th17 imbalance

为探讨miR-155-5p异常表达对变应性鼻炎(allergic rhinitis, AR)小鼠Th17/Treg免疫平衡的影响及机制,将6周龄BALB/c小鼠随机平均分成6组:A.对照组(Control)、B.模型组(AR)、C.miR-155-5p空载对照组(AR+Vector)、D.miR-155-5p过表达组(AR+miR-155-5p)、E.miR-155-5p阴性对照组(AR+NC)、F.miR-155-5p沉默组(AR+抗miR-155-5p抗体)。采用卵清蛋白(ovalbumin, OVA)诱导建立AR小鼠模型,鼻腔内给予miR-155-5p过表达或沉默慢病毒进行干预。实验末对各组小鼠过敏症状进行评分,HE染色观察鼻黏膜组织病理学变化,real-time PCR、Western blotting检测鼻黏膜组织中miR-155-5p、Foxp3、RORγt的表达水平,ELISA检测血清中IgE及炎性因子TGF-β1、IL-10、IL-17A、IL-6表达,流式细胞术检测外周血Treg和Th17百分比。结果发现,与对照组比较,模型组小鼠鼻炎症状评分明显增加,Foxp3、RORγt mRNA及蛋白表达水平明显升高,血清IgE、IL-17A、IL-6及Th17百分比显著上升,TGF-β1、IL-10及Treg百分比明显降低。MiR-155-5p过表达加重模型组小鼠鼻炎症状,同时促进IgE及Th17相关细胞因子的表达,抑制Treg相关细胞因子的表达,而miR-155-5p沉默后Treg/Th17免疫失衡得到缓解,小鼠鼻炎症状评分显著下降。MiR-155-5p可以显著加重AR小鼠的鼻炎症状,可能与影响Foxp3、RORγt表达,调节Treg/Th17免疫平衡有关。

To investigate the effect and mechanism of abnormal expression of miR-155-5 p on the immune balance of Th17/Treg in allergic rhinitis(AR)mice,BALB/c mice were randomly divided into normal control group,model group,AR+Vector group,AR+miR-155-5 p group,AR+NC group and AR+anti-miR-155-5 p group.The model of allergic rhinitis mice was established by sensitization and stimulation with ovalbumin,and intranasal administration of miR-155-5 p over-expression or silencing lentivirus.The allergic symptoms of mice in each group were evaluated,the pathological changes of nasal mucosa were observed by HE staining,the expressions of miR-155-5 p,Foxp3,RORγt were measured by real-time PCR and Western blotting,and the concentrations of IgE,inflammatory factors including TGF-β1,IL-10,IL-17 A and IL-6 in serum were detected by ELISA.The percentages of peripheral blood Treg and Th17 were analyzed by flow cytometry.The results showed that,compared with control group,the rhinitis symptom score of mice in the AR group was significantly increased,the expressions of Foxp3,RORγt at both mRNA and protein levels were significantly increased,the levels of serum IgE,Il-17 A,IL-6 and Th17 percentage were significantly increased,the levels of TGF-β1,IL-10 and Treg percentage were significantly decreased.MiR-155-5 p over-expression increased the rhinitis symptoms of mice in AR group,promoted the expression of IgE and Th17 related cytokines,and inhibited the expression of Treg related cytokines.We found that the Treg/Th17 immune imbalance was relieved after silencing of miR-155-5 p,and the rhinitis symptom score of mice significantly decreased.Therefore,miR-155-5 p could effectively exacerbate inflammatory symptoms of nasal mucosa in allergic rhinitis mice,which may be attributed to its effect on Foxp3 and RORγt expression and regulation of Treg/Th17 immune balance.