【摘要】 目的：观察正常人和配戴软性角膜接触镜(soft contact lens, SCL)人群结膜上皮细胞表面亚临床炎症因子HLADR和细胞内黏性分子1型ICAM1(CD54)的表达改变法：采用结膜印迹细胞学(conjunctival impression cytology,CIC)收集结膜上皮细胞，流式细胞学检查方法测定正常人及配戴软性角膜接触镜者HLADR和CD54的表达水平,计算阳性细胞率。结果：长期配戴软性角膜接触镜(>5a)组及(<5a)组中ICAM1和HLADR的表达较正常人组相比明显增加(PHLADR1< 0.01, PICAM1,1<0.01;PHLADR2<0.01,PICAM1,2<0.01),，前二者比较CD54有明显变化(P<0.01)而HLADR无明显变化(P>0.05)。结论:长期配戴软性角膜接触镜者结膜亚临床炎症因子ICAM1和HLADR明显增加，眼表的亚临床炎症增加。
【关键词】 印迹细胞学;软性角膜接触镜;ICAM1;HLADR;眼表组织
Changes of ICAM1 and HLADR induced by longterm soft contact lens wearMaoZhu Zhao , Yang Zheng, YiPing QiaoFoundation item: Key Fund Project of Sichuan Provincial Education Department (No.329)
1East Ward Eye Department, Sichuan Provincal Peoples Hospital, Chengdu 610110, Sichuan Province, China; 2Department of Ophthalmology,Affiliated Hospital of Luzhou Medical College, Luzhou 646000, Sichuan Province, ChinaAbstractAIM: To observe the change of inflammation agent production of the conjunctiva HLADR and intercellular adhesion molecule type 1 (ICAM1) (CD54) in normal subjects and in soft contact lens (SCL) wear subjects. METHODS: Conjunctival impression cytology(CIC) were used to collect conjunctival epithelium, and the expression levels of HLADR and CD54 in normal subjects and in SCL wear subjects were determined by flow cytometry. The percentage of positive cells was calculated and compared between each group. RESULTS: A significant increase of inflammation markers (HLADR and ICAM1) was observed in the longterm(>5a) and shortterm(<5a) SCL group compared with the control group (PHLADR1<0.01,PICAM1,1<0.01;PHLADR2<0.01,PICAM1,2<0.01). For the two SCL groups, ICAM1 has significant change(P<0.01), but HLADR has no significant change(P>0.05 ).CONCLUSION: This study confirms the difference in expression of inflammation markers (HLADR and ICAM1) in SCL and the healthy adult. It demonstrates a significant increase of subclinical conjunctival inflammation in SCL compared with the normal.
KEYWORDS: conjunctival impression cytology; soft contact lens; ICAM1; HLADR; ocular surface
0引言
配戴软性角膜接触镜会引起急性和慢性临床并发症(如角膜溃疡，巨乳头状结膜炎或角膜新生血管等的形成)。角膜接触镜作为表面异物可能会导致眼表的亚临床改变,Pisella等[1]报道，配戴角膜接触镜会引起结膜的亚临床炎症。结膜上皮细胞的亚临床炎症通常是一种慢性侵蚀的过程，这种病变属于眼表的慢性改变，比如干眼症,巨乳头状结膜炎等。我们采用结膜印迹细胞学(conjunctival impression cytology,CIC)，通过流式细胞学方法检测结膜上皮表达HLADR和ICAM1的变化，评估眼表炎症反应及配戴软性角膜接触镜人群中结膜的可能改变，探讨长期配戴软性角膜接触镜与眼表亚临床炎症的相关性。
1对象和方法
1.1对象
对200704/200804进行LASIK术前检查，年龄18～40岁之间健康成年人，男女比例随机。其中正常对照组26例，男14例，女12例，年龄18～25(平均19.0)岁;配戴SCL 1d～4a，平均1a者32例，男18例，女14例，年龄18～25(平均19.5)岁;配戴5.5～14a，平均6a者27例，男7例，女20例，年龄21～39(平均25)岁。受试者均身体健康，无吸烟史，无长期操作电脑史，近2wk内未滴任何眼液并经检查排除实质性角结膜干燥症和Sjgren综合征及由其他原因引起的眼部疾病。
1.2方法
醋酸纤维膜8cm×10cm(浙江台州四青塑料厂);HLA DRPEmAb、CD54FITCmAb，Mouse IgG1FITC，IgG1PE mAb(阴性对照)来自于Beckman Coulter 公司;EpicsXL型号流式细胞仪来自Beckman Coulter 公司。醋酸纤维膜剪成5mm×7mm大小，环氧乙烷消毒备用。取材时用眼科无齿镊夹持纤维膜，将粗糙面置于受试者颞侧球结膜表面，玻璃棒轻压约10s取出，将滤纸条含1g/L甲醛PBS液的试管中(PH 7.2～7.4)。采集样本后放置于快速旋涡混匀器上处理30min，尽可能将细胞从纤维膜上洗涤入1g/L甲醛PBS液中变为悬浮细胞;样本3000转/min离心5min，去上清; 每管样本分别加入HLA DRPEmAb，CD54FITCmAb各20μL，阴性对照组加Mouse IgG1FITC，IgG1PEmAb各20μL，室温避光孵育20min;每管加入PBS液500μL，上流式细胞仪，分析阳性细胞百分率。以细胞膜上表达CD54 和HLADR抗体为阳性细胞：CD54阳性细胞%= CD54阳性细胞数÷(CD54阳性细胞数+CD54阴性细胞数)×100%-非特异性结合细胞数;HLADR阳性细胞%= HLADR阳性细胞数÷(HLADR阳性细胞数+HLADR阴性细胞数)×100%-非特异性结合细胞数;
统计学分析：采用SPSS 13.0软件进行统计学处理，对流式细胞学数据检测3组间比较采用KruskalWallis Test方法，各组之间CD54和HLADR 之间的比较采用MannWhitney方法，相关性采用 Test Spearman 相关分析。检验水准P<0.05。
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