不同类型过敏性结膜炎患者眼表组织中白细胞介素-4、5和13的表达

摘要

背景 过敏性结膜炎的发病机制尚未完全明确.研究表明,白细胞介素-4(IL-4)、IL-5和IL-13在过敏性结膜炎中发挥重要作用,但这些因子对不同类型的过敏性结膜炎患者的作用是否一致尚有待证明.目的 探讨IL-4、IL-5和IL-13在不同类型过敏性结膜炎患者眼表组织中的表达及其在不同类型过敏性结膜炎发病中的作用.方法 采用前瞻性队列研究设计.纳入2013年4月至2014年9月在山西省眼科医院就诊的春季角结膜炎(VKC)患者20例40眼、季节性过敏性结膜炎(SAC)患者20例40眼和常年性过敏性结膜炎(PAC)患者20例40眼,并纳入同期健康志愿者20人40眼作为正常对照.用消毒刮刀在上睑结膜刮取结膜上皮细胞,分别采用免疫组织化学法、实时荧光定量PCR法检测结膜上皮细胞中IL-4、IL-5和IL-13蛋白及其mRNA的表达;采用1μl毛细玻璃管于受试者下睑颞侧睑缘处吸取泪液4μl,Luminex液相芯片测定受试者泪液中IL-4、IL-5和IL-13的表达.结果 免疫组织化学法染色表明,VKC组、SAC组和PAC组患者结膜上皮细胞的细胞质中均可见到IL-4、IL-5和IL-13的阳性表达,而在正常对照组受试者结膜上皮细胞中未发现IL-4、IL-5和IL-13的表达.以正常对照组IL-4、IL-5和IL-13 mRNA的表达量为对照,VKC组、SAC组和PAC组中IL-4 mRNA的相对表达量分别为4.11±1.24、2.71±0.71和2.00±0.80,IL-5 mRNA的相对表达量分别为4.02±0.43、2.07±0.45和1.47±0.50,IL-13 mRNA的相对表达量分别为6.44±0.66、4.35±1.26和2.39±0.86,4个组间IL-4、IL-5和IL-13 mRNA相对表达量的总体比较差异均有统计学意义(F=51.32、220.18、162.49,均P＜0.01);其中VKC组IL-4、IL-5和IL-13 mRNA的相对表达量明显高于SAC组和PAC组,SAC组IL-4、IL-5、IL-13 mRNA的相对表达量高于PAC组,差异均有统计学意义(均P＜0.05).正常对照组受试者泪液中未检测到IL-4、IL-5和IL-13表达,VKC组泪液中IL-4、IL-5和IL-13蛋白质量浓度分别为(14.06±3.50)、(10.88±1.82)和(34.28±8.42) pg/ml,SAC组分别为(7.71±0.65)、(5.10±1.33)、(23.77±6.29) pg/ml,PAC组分别为(3.30±1.50)、(2.43±1.28)和(17.67±4.28) pg/ml,组间总体比较差异均有统计学意义(F=200.29、260.49、128.23,均P＜0.01);其中VKC组IL-4、IL-5、IL-13蛋白质量浓度明显高于SAC组和PAC组,SAC组IL-4、IL-5和IL-13蛋白质量浓度明显高于PAC组,差异均有统计学意义(均P＜0.01).结论 IL-4、IL-5和IL-13参与多种类型过敏性结膜炎的发病,但在不同类型过敏性结膜炎患者中眼表的表达均有差异.%Background The pathogenesis of allergic conjunctivitis has not been clearly established.Current researchers indicate that interleukin-4 (IL-4), IL-5 and IL-13 may play an important role in allergic conjunctivitis.But whether the roles of these inflammatory factors are same in different types of allergic conjunctivitis remains unclear.Objective This study was to investigate the expressions of IL-4, IL-5 and IL-13 in ocular surface with different types of allergic conjunctivitis.Methods A prospective cohort study was designed.Eighty individuals were recruited in Shanxi Eye Hospital from April 2013 to September 2014, including 20 patients with vernal keratoconjunctivitis (VKC), 20 patients with seasonal allergic conjunctivitis (SAC), 20 patients with perennial allergic conjunctivitis (PAC) and 20 normal healthy subjects.Surficial tissues were binocularly scraped using disinfected scraper from upper eyelid conjunctiva, and 4 μl of tear fluid was obtained with capillary tube.The expressions of IL-4, IL-5 and IL-13 protein and mRNA in the conjunctival epithelial cells were detected by immunohistochemistry and real-time fluorescence quantitative PCR.The IL-4, IL-5 and IL-13 concentrations in tear fluid were assayed by Luminex method.This study complied with Declaration of Helsinki and the research protocol was approved by the Shanxi Eye Hospital Ethics Committee.Written informed consent was obtained from each subject prior to entering the cohort.Results IL-4, IL-5 and IL-13 were positively expressed in cytoplasm of conjunctival epithelial cells in the VKC group,SAC group and PAC group,but the expressions of IL-4,IL-5 and IL-13 were absent in the normal control group.The relative expression levels of IL-4 mRNA were 4.11±1.24,2.71±0.71 and 2.00±0.80;the relative expression levels of IL-5 mRNA were 4.02±0.43,2.07±0.45 and 1.47±0.50;and the relative expression levels of IL-13 mRNA expression levels were 6.44±0.66,4.35±1.26 and 2.39±0.86 in the VKC group,SAC group and PAC group, showing significant differences among the 4 groups (F =51.32,220.18,162.49, all at P＜0.01).The relative expression levels of IL-4,IL-5 and IL-13 mRNA were significantly higher in the VKC group than those in the SAC group and PAC group;and those in the SAC group were significantly elevated in comparison with the PAC group (all at P＜0.05).No IL-4, IL-5 and IL-13 were detected in the tear fluid in the normal control group;while the concentrations of IL-4,IL-5 and IL-13 in the tear fluid were (14.06±3.50), (10.88±1.82) and (34.28±8.42) pg/ml in the VKC group,and (7.71 ±0.65), (5.10± 1.33), (23.77±6.29) pg/ml in the SAC group as well as (3.30± 1.50) pg/ml, (2.43± 1.28) pg/ml and (17.67 ± 4.28) pg/ml in the PAC group, showing significant differences among the 3 groups (F =200.29,260.49,128.23, all at P＜0.01).IL-4, IL-5 and IL-13 concentrations in the tear fluid were significantly higher in the VKC group than those in the SAC group and PAC group,and those in the SAC group were significantly raised in comparison with the PAC group (all at P＜0.01).Conclusions IL-4,IL-5 and IL-13 participate in the pathogenesis of multiple allergic conjunctivitis,but their expressions in the ocular surficial tissue are discriminatory in different types of allergic conjunctivitis.