Jiang Nan, Zhao Gui-Qiu, Lin Jing, Hu Li-Ting, Che Cheng-Ye, Li Cui, Wang Qian, Xu Qiang, Zhang Jie, Peng Xu-Dong
Department of Ophthalmology, the Affiliated Hospital of Qingdao University, Qingdao 266003, Shandong Province, China.
Int J Ophthalmol. 2016 Apr 18;9(4):491-6. doi: 10.18240/ijo.2016.04.03. eCollection 2016.
To observe the presence and expression of indoleamine 2,3-dioxygenase (IDO) during the corneal immunity to Aspergillus fumigatus (A. fumigatus) in the murine models.
The murine model of fungal keratitis was established by smearing with colonies of A. fumigatus after scraping central epithelium of cornea and covering with contact lenses in C57BL/6 mice. The mice were randomly divided into control group, sham group and A. fumigatus keratitis group. The cornea was monitored daily using a slit lamp and recorded disease score after infection. Corneal lesion was detected by immunofluorescence staining. IDO mRNA and protein were also detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blot.
The disease score and slit lamp photography indicated that disease severity was consistent with corneal inflammation in the murine models, and the disease scores in A. fumigatus keratitis group were obviously higher than those in the sham group. By immunofluorescence staining, IDO was mainly localized in corneal epithelium and stroma in the murine corneal tissues with A. fumigatus keratitis. Compared with the sham group, IDO mRNA expression was significantly enhanced in corneal epithelium infected by A. fumigatus. Furthermore, IDO protein expression detected by Western blot was in accord with transcript levels of IDO mRNA measured by qRT-PCR. IDO protein expression was enhanced after A. fumigatus infection compared with the sham group.
IDO is detected in corneal epithelium and stroma locally, which indicates IDO takes part in the pathogenesis of A. fumigatus keratitis and plays a key role in immune regulation at the early stage.
观察吲哚胺2,3-双加氧酶(IDO)在小鼠烟曲霉角膜免疫模型中的存在及表达情况。
刮除C57BL/6小鼠角膜中央上皮后涂抹烟曲霉菌落并覆盖隐形眼镜,建立真菌性角膜炎小鼠模型。将小鼠随机分为对照组、假手术组和烟曲霉角膜炎组。每日用裂隙灯监测角膜情况,并记录感染后的疾病评分。通过免疫荧光染色检测角膜病变。采用定量逆转录-聚合酶链反应(qRT-PCR)和蛋白质印迹法检测IDO的mRNA和蛋白。
疾病评分和裂隙灯照相显示,小鼠模型中的疾病严重程度与角膜炎症一致,烟曲霉角膜炎组的疾病评分明显高于假手术组。免疫荧光染色显示,在烟曲霉角膜炎小鼠角膜组织中,IDO主要定位于角膜上皮和基质。与假手术组相比,烟曲霉感染的角膜上皮中IDO mRNA表达显著增强。此外,蛋白质印迹法检测的IDO蛋白表达与qRT-PCR检测的IDO mRNA转录水平一致。与假手术组相比,烟曲霉感染后IDO蛋白表达增强。
在角膜上皮和基质中局部检测到IDO,这表明IDO参与了烟曲霉角膜炎的发病机制,并在早期免疫调节中起关键作用。