背景 干眼患者泪液分泌功能的持续异常导致眼表和泪腺处于长期炎性细胞浸润状态,炎症反应刺激淋巴细胞增生以产生对泪腺的免疫攻击,且炎症本身亦干扰腺体的正常分泌,其中趋化因子及其受体对淋巴细胞趋化、聚集到炎症部位起着关键的作用. 目的 分析干眼患者眼表细胞中CC趋化因子受体5(CCR5)、CXC趋化因子受体3(CXCR3)及其配体的表达变化,探讨以CCR5、CXCR3为特征的Th1细胞诱导的迟发性变态反应在干眼发生和发展中的机制.方法 采用前瞻性研究方法.纳入2012年6-12月在新疆医科大学第一附属医院眼科确诊的干眼患者30例60眼及年龄和性别匹配的健康志愿者30人60眼,受试者均接受泪膜破裂试验(BUT)、基础泪液分泌试验Ⅰ(SⅠt)和角膜结膜荧光素染色检查.采用结膜印记细胞学的方法获取2个组受试者的结膜上皮细胞,应用流式细胞术分析2个组受试者结膜组织CCR5、CXCR3表达阳性细胞率;应用real-time PCR对受检者结膜中正常T细胞表达和分泌调节活性因子(RANTES)、巨噬细胞炎性蛋白(MIP)-1α、MIP-1 β、γ干扰素诱导的单核因子(MIG)、γ干扰素诱导蛋白10(IP10)及干扰素诱导T细胞α趋化因子(I-TAC) mRNA的相对表达量进行检测,比较干眼组和健康对照组受试者间CCR5、CXCR3及其配体表达的差异,分析CCR5、CXCR3阳性表达率与BUT、SⅠt及角膜结膜荧光素染色评分的关系.结果 干眼组BUT、SⅠt值分别为(2.90 ±1.37)s和(4.00 ±2.49) mm/5 min,明显低于健康对照组的(8.56 ±4.69)s和(11.31 ±5.23) mm/5 min,差异均有统计学意义(t=3.172、2.186,均P＜0.05);干眼组角膜结膜荧光素染色评分及CCR5和CXCR3的阳性表达率分别为0.90 ±0.57、(3.38±0.66)％和(2.64±0.47)％,均明显高于对照组的0.14 ±0.06、(2.12±0.21)％和(1.12±0.11)％,差异均有统计学意义(t=2.297、3.151、5.454,均P＜0.05).干眼组CCR5阳性细胞率与BUT、SⅠt值均呈负相关(r=-0.473、-0.385,均P＜0.05),CXCR3阳性细胞率与BUT、SⅠt值均呈负相关(r=-0.753、-0.684,均P＜0.05);CCR5与CXCR3阳性细胞率及角膜结膜荧光素染色评分均呈正相关(r=0.231、0.336,均P＜0.05).干眼组RANTES、MIP-1α、MIG和IP10 mRNA的相对表达量明显高于健康对照组,差异均有统计学意义(t=3.091、2.894、2.688、2.245,均P＜0.05),而2个组间MIP-1β和I-TAC mRNA相对表达量的差异均无统计学意义(t=0.512、1.979,均P＞0.05).CCR5阳性细胞率与结膜中RANTES和MIP-1αmRNA相对表达量均呈正相关(r=0.473、0.285,均P＜0.05),而CCR5与MIP-1β则无明显相关性(r=0.214,P＞0.05).CXCR3阳性细胞率与MIG和IP10 mRNA相对表达量均呈正相关(r=0.553、0.314,均P＜0.05);CXCR3与I-TAC mRNA的相对表达量间无明显相关(r=0.364,P＞0.05).结论 干眼的发生可能伴随着炎性细胞的长期浸润,以CCR5、CXCR3为标志的Th1型细胞及自然杀伤细胞诱导的迟发性变态反应可能是导致干眼眼表损伤的主要原因,CCR5和CXCR3及其配体可作为调节干眼组织免疫炎症反应的靶点.%Background Sustained abnormal tear secretion in dry eye patients may lead to ocular surface and lacrimal glands in the state of long-term inflammatory cell infiltration.Lacrimal gland suffers immune attack by lymphoproliferative,and inflammation interferes normal gland secretion,in which chemokines and their receptors on lymphocytes play a key role.Objective This study was to investigate the inflammation mechanism of delayed allergy induced by Th1 cell in the development of dry eye.Methods Sixty eyes of 30 patients with dry eye and matched 30 healthy volunteers were enrolled in Affiliated First Hospital of Xinjiang Medical University from June to December in 2012.Schirmer Ⅰ test (S Ⅰ t),break up time (BUT) of tear and corneal fluorescence stain (FLS) were performed on the subjects,and conjunctiva epithelial cells were obtained using cytological method of conjunctiva imprinting.Positive cell rates of CC chemokine receptor 5 (CCR5) and CXC chemokine receptor 3 (CXCR3) were detected by flow cytology,and the relative expression levels of regulated upon activation of normal T cells exp ressed and secreted (RANTES),macrophage inflammatory protein (MIP)-lα and MIP-1β,monokine induced by interferon-γ (IFN-γ) (MIG),interferon-γ inducible protein 10 (IP10) and interferon-inducible T-cell α chemoattractant (I-TAC) mRNA were quantitatively assayed by real-time PCR.Differences of the positive rates of CCR5,CXCR3 and lignds were compared between dry eye group and normal control group.Relationship between the positive rates of CCR5,CXCR3 and BUT,S Ⅰ t,FLS scores was analyzed.Results The values of BUT,S Ⅰ t were (2.90±1.37) seconds and (4.00±2.49) mm/5 minutes in the dry eye group,which were significantly lower than (8.56±4.69) seconds and (11.31 ±5.23) mm/5 minutes in the normal control group (t =3.172,2.186,both at P＜0.05).FLS scores,positive rates of CCR5 and CXCR3 were0.90±0.57,(3.38±0.66) ％ and (2.64±0.47)％ in the dry eye group,showing significant elevations in comparison with 0.14±0.06,(2.12±0.21) ％ and (1.12±0.11) ％ in the normal control group (t=2.297,3.151,5.454,all at P＜0.05).In the dry eye group,the masculine rate of CCR5 was negatively correlated with BUT and S Ⅰ t (r=-0.473,-0.385,both at P＜0.05).The masculine rate of CXCR3 was negatively correlated with BUT and S Ⅰ t (r =-0.753,-0.684,both at P＜0.05).No considerable correlations between the positive rate of CCR5 with the positive rate of CXCR3 or FLS scores (r =0.231,0.336,both at P＜0.05).The relative expression levels of RANTES,MIP-1α,MIG and IP10 mRNA in the dry eye group were significantly higher than those in the normal control group (t =3.091,2.894,2.688,2.245,all at P＜0.05),but there were no significant differences in the relative expression levels of MIP-1β and I-TAC mRNA between the two groups (t =0.512,1.979,both at P＞0.05).The positive correlations were seen between the masculine rate of CCR5 with the relative expression of RANTES or MIP-1α mRNA (r=0.473,0.285,both at P＜0.05),but there was no obvious correlation between the masculine rate of CCR5 and the MIP-1 β mRNA expression(r=0.214,P＞0.05).In addition,the masculine rate of CXCR3 was positive correlated with the expressions of MIG and IP10 mRNA (r=0.553,0.314,both at P＜0.05),whereas the masculine rate of CXCR3 was not related to the expression of I-TAC mRNA (r=0.364,P＞0.05).Conclusions Dry eye is probably along with the long-term infiltration of inflammatory cells.The delayed allergy induced by Th1 cells and the nature killed cells is probably the primary cause to xerophthalmia.CCR5,CXCR3 and their ligands might be the regulative targets in the inflammation mechanism of dry eye.
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