RCS大鼠视网膜色素变性过程中视网膜免疫微环境的改变  

Changes of the immune-microenvironment in retinal degeneration process of RCS rat

作　　者：唐环宇 卢紫阳 何军材 郜原 阴正勤 Tang Huanyu;Lu Ziyang;He Juncai;Gao Yuan;Yin Zhengqin(Department of Ophthalmology,Key Laboratory of Visual Damage and Regeneration&Restoration of Chongqing,Southwest Hospital,Army Medical University,Chongqing 400038,China)

机构地区：[1]陆军军医大学西南医院,全军眼科中心,视觉损伤与修复重庆市重点实验室,重庆400038

出　　处：《中华实验眼科杂志》2018年第11期839-846,共8页Chinese Journal Of Experimental Ophthalmology

基　　金：国家重点基础研究发展计划(973计划)项目(2013CB967002);重庆市基础科学与前沿技术研究(重点)项目(cstc2017jcyjBX0058)。

摘　　要：目的研究在英国皇家外科学院(RCS)大鼠视网膜色素变性(RP)发展过程中,介导细胞免疫的T淋巴细胞和自然杀伤(NK)细胞及其分泌的细胞因子γ干扰素(IFN-γ)的变化。方法选取出生后20日龄(P20)、P40、P60 RCS-rdy--P+大鼠(简称RCS大鼠)和相应日龄RCS-rdy+-P+大鼠(简称对照大鼠)各12只,细胞因子液相悬浮芯片检测大鼠视网膜匀浆中多种炎性细胞因子的质量浓度。实时荧光定量聚合酶链反应(real-time PCR)检测各日龄大鼠视网膜中白细胞介素-2(IL-2)、CC类趋化因子配体2(CCL2)、CXC趋化因子配体9(CXCL9)、CXCL10和IFN-γmRNA的表达水平;免疫组织化学技术检测大鼠视网膜中T淋巴细胞(CD4+、CD8+)和NK细胞(CD161+)的分布;流式细胞术检测大鼠视网膜中T淋巴细胞(CD4+、CD8+)和NK细胞(CD161+)的比例,酶联免疫吸附试验(ELISA)检测大鼠视网膜组织匀浆中IFN-γ的质量浓度。结果RCS组大鼠视网膜中淋巴细胞相关细胞因子和趋化因子mRNA表达水平均表现出随日龄增加而表达上调的趋势,P60 RCS大鼠视网膜中IL-2、CCL2、CXCL9、CXCL10、CXCL11和IFN-γmRNA表达水平均较P20 RCS大鼠和对照组大鼠表达水平明显升高,差异均有统计学意义(均P<0.05)。P60 RCS大鼠视网膜中CD4、CD8、CD161细胞阳性率分别为(9.09±0.89)%、(18.77±0.38)%和(9.41±0.38)%。P60 RCS大鼠视网膜中IFN-γ阳性细胞比例为(8.29±0.27)%,较对照大鼠的(0.28±0.02)%显著升高,差异有统计学意义(t=29.03,P=0.00)。P60 RCS大鼠视网膜中CD4+、CD8+和CD161+淋巴细胞主要分布在内层视网膜,且IFN-γ阳性标记与淋巴细胞表面标记呈共定位。不同日龄RCS大鼠和对照大鼠视网膜中IFN-γ质量浓度比较,差异均有统计学意义(F组别=16.49,P<0.01;F时间=21.05,P<0.01),其中P60 RCS大鼠视网膜中IFN-γ质量浓度较P20 RCS大鼠、P40 RCS大鼠和对照组大鼠明显升高,差异均有统计学意义(均P<0.05)。结论RCS大鼠RP改变了视网膜内相对免�Objective To explore the immune-microenvironment of the retinas at different stages of retinal degeneration in Royal College of Surgeon(RCS)rats.Methods RCS-rdy--P+(RCS)rats at early stage(P20),middle stage(P40)and late stage(P60)were involved,12 rats at each post-natal day,RCS-rdy+-P+rats severed as control.Relative concentrations of rat cytokines in rat retina homogenate were detected by using Bio-Plex Suspension Array System.Relative expressions of interleukin-2(IL-2),C-C motif ligand 2(CCL2),chemokine(C-X-C motif)ligand 9(CXCL9),CXCL10,CXCL11 and interferon-γ(IFN-γ)mRNA in rat retina were analyzed by real-time PCR.Expressions of IFN-γand immune cells surface marker CD4,CD8 and CD161 in the retinas were detected by immunohistochemical staining.Percentage of IFN-γpositive T lymphocytes and natural killer(NK)cells in rat retina were analyzed by flow cytometry.The concentrations of IFN-γin rat retina homogenate were evaluated by enzyme-linked immunosorbent assay(ELISA).The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results Lymphocytes related cytokines and chemokines mRNA expression levels in the RCS rat retinas showed increase trends with the extension of time.The expression levels of IL-2,CCL2,CXCL9,CXCL10,CXCL11 and IFN-γmRNA in P60 RCS rat retinas were significantly increased than those in the P20 RCS rat retinas and the control rat retinas(all at P<0.05).The positive rates of CD4,CD8 and CD161 cells in the retinas of P60 RCS rats was(9.09±0.89)%,(18.77±0.38)%and(9.41±0.38)%,respectively.The proportion of IFN-γpositive cells in the retinas of P60 RCS rats was(8.29±0.27)%,which was significantly higher than that of the control rats([0.28±0.02]%),with a significant difference between them(t=29.03,P=0.00).CD4+,CD8+and CD161+lymphocytes were mainly distributed in the retinas of P60 RCS rats,and the expressions of IFN-γwere co-located with lymphocyte surface markers.There were signif

关 键 词：视网膜变性疾病 淋巴细胞 Γ干扰素 

分 类 号：R774.1[医药卫生—眼科]