Th17细胞因子在小鼠真菌性角膜炎中的表达

发布时间：2018-12-25 14:02

【摘要】：目的通过建立真菌性角膜炎小鼠模型，观察真菌性角膜炎的发生发展，探索TH17细胞因子在真菌性角膜炎中的表达情况，初步探讨IL-17及TH17细胞在真菌性角膜炎免疫学中的作用。方法清洁级BALB/c小鼠随机分成两组，其中一组为真菌感染组，小鼠右眼采用角膜表层镜片法建立茄病镰刀菌性角膜炎模型；另一组为损伤对照组，以PBS代替真菌菌液采取与感染组相同方法处理小鼠右眼。分别于感染后第1、3、5、7天，裂隙灯显微镜观察角膜炎特点并行临床评分；HE染色观察角膜病理变化；实时荧光定量PCR检测角膜中Th17细胞因子（IL-17）及其特异性转录因子RORγt的mRNA基因的表达变化；酶联免疫吸附试验检测IL-17因子蛋白的表达变化。结果1.病变特征所有小鼠真菌性角膜炎模型建立成功，真菌感染早期呈全角膜的弥漫性炎症反应，，浸润混浊进行性加重，感染后3天角膜灰白色雾状混浊，５天时病变典型，角膜周边新生血管长入，中央形成溃疡，第７天后病灶趋向好转，总病程持续约12天。临床评分各时间点有统计学差异（P＜0.05）。 2.病理学检查 HE染色可见早期角膜基质水肿，感染后第5天角膜基质紊乱多形粒细胞浸润明显增多，前房内可见渗出，晚期角膜基质中可见新生血管及管内红细胞，溃疡被瘢痕组织取代。 3.Real time PCR与ELISA检测结果 Thl7细胞因子（IL-17）和Th17细胞特异性转录因子（ROR γt）基因mRNA在真菌感染角膜后均出现表达，于第3、5、7天显著高于损伤对照组（P＜0.01），且组内比较5天时表达量最高（P＜0.01）； IL-17因子蛋白感染后各时间点表达量显著高于损伤对照组（P＜0.01）。结论新型CD4阳性细胞亚群Th17细胞在真菌性角膜炎中分化，参与其免疫反应，Th17细胞因子（IL-17）在真菌性角膜炎中发挥重要作用。
[Abstract]:Objective to investigate the expression of TH17 cytokines in fungal keratitis by establishing a mouse model of fungal keratitis. To explore the role of IL-17 and TH17 cells in the immunology of fungal keratitis. Methods BALB/c mice of clean grade were randomly divided into two groups. One group was fungal infection group. The model of Fusarium solanum keratitis was established by corneal surface lens method in the right eye of mice. The other group was treated with PBS instead of fungal solution in the same way as the infected group. The characteristics of keratitis were observed by slit lamp microscope and the pathological changes of cornea were observed by HE staining. The expression of Th17 cytokines (IL-17) and its specific transcription factor ROR 纬 t (ROR 纬 t) was detected by real-time fluorescence quantitative PCR and enzyme linked immunosorbent assay (Elisa). Result 1. All mice models of fungal keratitis were established successfully. The early stage of fungal infection was a diffuse inflammatory reaction of the whole cornea, the infiltration and opacity gradually aggravated, the cornea became gray and haze at 3 days after infection, and the pathological changes were typical at 5 days after infection. The neovascularization around the cornea resulted in central ulcer, and the lesion tended to improve after 7 days. The total course of disease lasted about 12 days. There was statistical difference in clinical score at different time points (P < 0.05). 2. HE staining showed early corneal stroma edema, increased corneal stromal disorder polymorphonuclear cell infiltration and exudation in anterior chamber on the 5th day after infection, neovascularization and erythrocyte in the late corneal stroma. The ulcer is replaced by scar tissue. 3.Real time PCR and ELISA analysis showed that Thl7 cytokines (IL-17) and Th17 cell specific transcription factor (ROR 纬 t) gene mRNA were expressed after fungal infection of cornea. On the 7th day, the expression level was significantly higher than that in the injured control group (P < 0.01), and the highest expression was observed at the 5th day after injury (P < 0.01). The expression of IL-17 factor protein at each time point after infection was significantly higher than that in the injured control group (P < 0. 01). Conclusion Th17 cells, a new type of CD4 positive cell subgroup, differentiate in fungal keratitis and participate in its immune response. Th17 cytokines (IL-17) play an important role in fungal keratitis.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R772.21

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