摘要
目的研究低氧刺激慢性鼻窦炎伴鼻息肉(CRSwNP)与正常鼻黏膜上皮细胞炎性因子的变化与异同,探讨其在CRSwNP发病机制中的作用。方法选择2015年6月至2018年1月在吉林大学中日联谊医院就诊的68例CRSwNP患者,其中男性36例,女性32例,年龄(45.2±12.5)岁(xˉ±s,下同),患者的鼻息肉黏膜纳入慢性鼻窦炎鼻息肉组(CRS-NP组),下鼻甲黏膜纳入慢性鼻窦炎下鼻甲组(CRS-IT组),并根据组织病理学结果进一步分成嗜酸粒细胞浸润及非浸润两种类型,即嗜酸粒细胞浸润性鼻息肉组(Eos-NP组,n=34)、非嗜酸粒细胞浸润性鼻息肉组(Non-Eos-NP组,n=34)、嗜酸粒细胞浸润性鼻息肉患者的下鼻甲组(Eos-IT组,n=20)和非嗜酸粒细胞浸润性鼻息肉患者的下鼻甲组(Non-Eos-IT组,n=20);同期25例鼻窦囊肿或鼻中隔偏曲患者的下鼻甲黏膜作为对照组(n=25),其中男性14例,女性11例,年龄(42.8±10.2)岁。免疫组织化学染色分析各组黏膜上皮组织中白细胞介素(IL)17A、干扰素γ(IFN-γ)、肿瘤坏死因子α(TNF-α)与乏氧诱导因子(hypoxia-inducible factor 1α,HIF-1α)的表达情况。酶联免疫吸附试验检测低氧刺激0、24和48 h后各组原代鼻黏膜上皮细胞分泌IL-17A、IFN-γ和TNF-α的差异;免疫荧光、固态光源高内涵与免疫印记实验检测原代鼻黏膜上皮细胞HIF-1α的表达情况。应用SPSS 17.0软件对数据进行统计学分析,采用双向方差分析作为主要统计学方法。结果免疫组织化学染色结果显示,IL-17A和TNF-α在对照组中表达最高(光密度值分别为0.37±0.03、0.53±0.02),IFN-γ和HIF-1α在Eos-IT组中表达最高(光密度值分别为0.47±0.03、0.39±0.02)。未接受低氧刺激时,IL-17A与TNF-α在对照组中水平较低;低氧刺激48 h后,对照组的IL-17A与TNF-α分泌量明显高于其他各组。未接受低氧刺激时,Eos-NP组分泌的IFN-γ明显高于对照组[(13.7±1.3)pg/ml比(11.1±1.6)pg/ml,P<0.05];低氧刺激48 h后,IFN-γ在对照组和Eos-NP组中的分泌量没有显著区别。对照组与CRS-IT组表达的HIF-1α随低氧时间延长而增加,Eos-NP组与Non-Eos-NP组表达的HIF-1α随低氧时间延长而减少。对照组与CRS-IT组HIF-1α主要表达于鼻黏膜上皮细胞的细胞质内,CRS-NP组HIF-1α主要表达于鼻黏膜上皮细胞的细胞核内。结论低氧刺激下,CRSwNP患者的鼻息肉、下鼻甲与正常鼻黏膜上皮细胞中IL-17A、TNF-α、IFN-γ的分泌以及HIF-1α的表达水平存在差异,且呈现不同的亚细胞定位,提示其可能参与了CRSwNP发病相关基因的转录与调控。
Objective To investigate the roles of hypoxic stimulation in the pathogenesis of chronic rhinosinusitis with nasal polyps(CRSwNP)by comparing the variation and differences of inflammatory factors secreted from epithelial cells of nasal polyps and normal nasal mucosa under hypoxic stimulation.Methods Sixty-eight patients who were diagnosed with CRSwNP from June 2015 to January 2018 at China-Japan Union Hospital of Jilin University were analyzed,including 36 males and 32 females,aged(45.2±12.5)years.Nasal polyps mucosa was included in CRS-NP group and inferior turbinate mucosa was included in CRS-IT group.According to the degree of eosinophil infiltration in histopathologic results,each of these two groups was further divided into eosinophil infiltration and non-eosinophil infiltration as Eos-NP group(n=34),Non-Eos-NP group(n=34),Eos-IT group(n=20)and Non-Eos-IT group(n=20).The inferior turbinate mucosa of twenty-five patients who were diagnosed with cyst of paranasal sinus or deviation of nasal septum was classified as control group(n=25),including 14 males and 11 females,aged(42.8±10.2)years.The expression of interleukin 17A(IL-17A),interferonγ(IFN-γ),tumor necrosis factorα(TNF-α)and hypoxia-inducible factor 1α(HIF-1α)in each group was analyzed by immunohistochemical staining.After 0,24 and 48 h hypoxic stimulation,the secretion of IL-17A,IFN-γ,TNF-αin primary nasal mucosa epithelial cells of each group was tested by enzyme-linked immune sorbent assay(ELISA)experiment;the expression of HIF-1αwas tested by immunofluorescent staining and imaging and Western blot.SPSS 17.0 software and two-way ANOVA were used for statistical analysis.Results Immunohistochemical staining showed that the expression of IL-17A and TNF-αwas much higher in control group(optical density(OD)value was 0.37±0.03,0.53±0.02,respectively)and the expression of IFN-γand HIF-1αwas much higher in Eos-IT group(OD value was 0.47±0.03,0.39±0.02,respectively).The secretion of IL-17A and TNF-αwas much lower in control group than that in other groups under normal condition.After 48 h hypoxic stimulation,the secretion of IL-17A and TNF-αwas much higher in control group compared with other groups.The secretion of IFN-γin Eos-NP group was much higher than that in control group under normal condition((13.7±1.3)pg/ml vs(11.1±1.6)pg/ml,P<0.05).After 48 h hypoxic stimulation,there was no difference of IFN-γbetween control group and Eos-NP group.The expression of HIF-1αdecreased in Eos-NP group and Non-Eos-NP group while increased in CRS-IT group and control group upon prolonged exposure to hypoxia.HIF-1αwas mostly located at cytoplasm of epithelial cells in control and CRS-IT group while mainly located at nucleus of epithelial cells in CRS-NP group.Conclusions The secretion of IL-17A,TNF-α,IFN-γand the expression of HIF-1αshow significant difference between normal nasal mucosa,polyps and inferior turbinate of CRSwNP under hypoxic stimulation,presenting different subcellular localization.This illustrates the proteins above are involved in transcription and regulation of the gene responsible for the pathogenesis of CRSwNP.
作者
修倩
高奕瑶
朱冬冬
Xiu Qian;Gao Yiyao;Zhu Dongdong(Department of Otorhinolaryngology Head and Neck Surgery,China Japan Hospital of Jilin University,Changchun 130000,China;Department of Scientific Research Center,China-Japan Union Hospital,Changchun 130000,China;Jilin Provincial Key Laboratory of Precise Diagnosis and Treatment of Upper Airway Allergic Diseases,Changchun 130000,China)
出处
《中华耳鼻咽喉头颈外科杂志》
CSCD
北大核心
2021年第3期263-272,共10页
Chinese Journal of Otorhinolaryngology Head and Neck Surgery
基金
国家自然科学基金(81800887,81870701)
吉林省卫生科研人才专项项目(2019SCZ004)
吉林省卫生专项项目(2018scz042)
吉林省科技厅立项(20200201200JC)。
