大鼠烟曲霉菌性角膜炎初期核苷酸结合寡聚域样受体在角膜中的表达及其免疫防御作用  

Expression and effect of nucleotide oligomerization domain-like receptors in the initial of Aspergillus fumigatus keratitis in rats

作　　者：渠晓黎[1] 赵桂秋[1] 许正杰[1] 高昂[1] 王楠[1] 刘莹[1] 林静[1] 

机构地区：[1]青岛大学医学院附属医院眼科,266003

出　　处：《中华实验眼科杂志》2014年第10期870-875,共6页Chinese Journal Of Experimental Ophthalmology

基　　金：国家自然科学基金项目（81170825）

摘　　要：背景核苷酸结合寡聚域（NOD）样受体（NLRs）在天然免疫过程中发挥重要作用，但其在真菌性角膜炎发生和发展过程中的作用研究少见。目的研究NOD，在大鼠烟曲霉菌性角膜炎（AFK）角膜组织中的表达及作用。方法72只成年清洁级Wistar大鼠按照随机数字表法随机分为正常对照组、单纯角膜上皮损伤组和AFK模型组，均以右眼作为实验眼。正常对照组12只大鼠中选取6只仅刮取角膜上皮用于逆转录PCR（RT—PCR）实验，另6只大鼠制备角膜标本用于组织病理学检查、免疫组织化学和免疫荧光染色实验。单纯角膜上皮损伤组共30只大鼠，仅刮除中央角膜上皮，AFK模型组大鼠共30只，刮除角膜上皮后接种烟曲霉菌标准株。分别于实验后4、8、16、24h选取各组6只大鼠刮取角膜上皮，采用RT—PCR法检测角膜中NOD2mRNA的表达，其他6只于实验后24h制备角膜标本，采用免疫组织化学法和免疫荧光染色法检测角膜中NOD2蛋白的表达，并将各组检测结果进行比较。实验后4、8、16、24h分别摘除各组大鼠右眼眼球，制作角膜标本进行常规组织病理学检查。结果裂隙灯显微镜下观察各组大鼠各时间点的角膜情况，均造模成功。RT—PCR结果显示，正常对照组角膜仅有微量NOD2mRNA的表达，单纯角膜上皮损伤组和AFK模型组大鼠角膜中NOD2mRNA的相对表达量明显增加，且在实验4、8、16、24h，AFK模型组大鼠角膜中NOD2mRNA的相对表达量均明显高于单纯角膜上皮损伤组，差异均有统计学意义（t=-0．409、-0．439、-0．534、-0．618，均P=0．000）。大鼠角膜组织病理学检查显示，正常对照组大鼠角膜结构完整；单纯角膜上皮损伤组可见到少部分角膜上皮缺损、前弹力层皱褶及角膜轻度水肿，有少量中性粒细胞浸润；AFK模型组大鼠可见角膜溃疡，角膜水肿增厚，浅基质层可见大量中性粒细胞浸润。Background Studies have determined that nueleotide binding oligomerization domain 2 （NOD2） plays a key role in innate immune response. However,whether NOD2 participates in the nature defense of fungal keratitis is unclear. Objective This study was to investigate the expression and significance of NOD2 on cornea in the initial of Aspergillus fumigatus keratitis （AFK） in rats. Methods Seventy-two adult clean Wistar rats were randomized into the normal control group, only corneal epithelial scraped group and AFK model group, and theAFK models were established by incubating Aspergillus fumigatus to cornea after corneal epithelium was scraped. All the operations were performed in the right eyes of rats. Reverse transcription PCR （RT-PCR） was carried out to detect the expression of NOD2 mRNA in corneal epithelium 4,8, 16,24 hours after operation. Twenty-four hours after operation,the expression of NOD2 protein in rat corneas was examined by immunochemistry and immnunofluorescence technology. Also,the rat corneas were obtained for regular histopathological examination. The use and care of the animals complied with Institutional Animal Care and Use Committee Guidebook by NIH. Results All the models were made successfully. RT-PCR revealed that a fewer NOD2 mRNA were expressed on cornea in the normal control group,but the expressing levels of NOD2 mRNA were increased in the only corneal epithelial scraped group and AFK model group. Compared with only corneal epithelial scraped group,the elevated values of NOD2 mRNA expression in the AFK model group were statistically significant at 4,8,16 and 24 hours after operation （ t = -0. 409, -0. 439, -0. 534,-0. 618, all at P = 0. 000）. The histopathological examination displayed that the cornel tissue had intact structure in the normal control group, and partly corneal epithelial deficiency, slight corneal swelling and fewer neutrophil granulocytes were seen in the only corneal epithelial scraped group. However,corneal ulcer,severe corneal edema and a lot of neutrop

关 键 词：烟曲霉菌 眼部感染 真菌 角膜炎 核苷酸结合寡聚域 天然免疫 大鼠 

分 类 号：R772.2[医药卫生—眼科]