敲除TLR2基因对同种异体小鼠角膜移植排斥反应的影响  

Variation of immune response after allogeneic corneal transplantation in TLR2 knock-out mice

作　　者：苏亚茹 白浪[1] 汤明芳[1] 于健[1] 刘晶[1] 孙宇飞[1] 孟婷[1] 

机构地区：[1]南方医科大学南方医院眼科,广东省广州市510000

出　　处：《眼科新进展》2016年第11期1006-1010,共5页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:81170887);南方医科大学南方医院横向课题匹配基金资助(编号:G201202)~~

摘　　要：目的探讨敲除Toll样受体2(toll-like receptor 2,TLR2)基因后是否抑制同种异体小鼠角膜排斥反应的发生。方法以BALB/c为供体,各取30只C57BL/6和同背景TLR2基因敲除鼠为受体行右眼角膜移植术,设为野生组(WT)和基因敲除组(KO);取19只C57BL/6行右眼自体角膜移植术,为自体组(ISO);各取9只C57BL/6和TLR2基因敲除鼠分别设为野生对照组(WT control)和基因敲除对照组(KO control)。术后每周两次观察植片并记录免疫排斥的发生时间;术后14 d,收集术眼同侧颈部淋巴结,行流式细胞术分析CD4+T细胞百分比;收集术眼角膜,行免疫组织化学染色和实时荧光定量PCR检测角膜干扰素(interferon,IFN)-γ、TLR2和My D88的表达。结果 WT组和KO组小鼠角膜移植术后中位生存时间KO组(35.0±3.8)d长于WT组(21.0±1.5)d(P<0.05)。流式细胞术分析显示,WT组同侧颈部淋巴结CD4+T细胞百分比较WT control组明显增高(P<0.05),而ISO和KO组相对于其对应的control组(WT/KO control)差异均无统计学意义(均为P>0.05);免疫组织化学结果显示,ISO和KO组间角膜IFN-γ和My D88分子表达无差异,但两者均低于WT组。KO组角膜几乎不表达TLR2分子,ISO组有微量表达,WT组表达明显增高;PCR检测显示,ISO和KO组角膜IFN-γ和TLR2 mRNA相对表达量差异均无统计学意义(均为P>0.05),但两者均低于WT组(均为P<0.05);KO组角膜My D88 mRNA相对表达量比ISO组高(P<0.05),但两者均低于WT组(均为P<0.05)。结论敲除TLR2基因可以一定程度抑制同种异体小鼠角膜排斥反应的发生。Objective To explore whether toll-like receptor 2 （TLR2） knock-out can attenuate immune rejection after allogeneic corneal transplantation. Methods BALB/c mice were used as donors, 30 C57BL/5 mice and 30 TLR2 knock-out mouse were chosen as receptors to establish allograft corneal transplantation models, which were grouped as the wide type （WT） mouse group and knock-out （KO） mouse group. Another 19 C57BL/5 mice were performed isograft corneal transplantation and were termed as isograft （ISO） group. Nine normal mice of wild-type C57BL/5 （WT control） or TLR2 knock-out （KO control） were respectively involved as negative controls. The edema and transparency were observed twice per week under slit lamp microscope after surgery and the rejection times were recorded according to Sonoda＇ s criteria. At 14 days after transplantation, the frequencies of CD4 ＋ T cells in ipsilateral cervical draining lymph nodes were analyzed by flow cytometry, and the expression of IFN-γ, TLR2 and MyD88 were detected by immunohistochemical staining and qPCR. Results The me- dian survival time （MST） in KO group was （35.0 ±3.8） days,while in WT group was （21.0 ± 1.5 ） days, there was significant difference between WT and KO group （P 〈 0. 05 ）. Lymphoid CD4 ＋ T cells percentage of WT group was significantly higher than that of WT control group （P 〈 0.05 ）. There was no statistical difference between ISO and WT control group, as well as KO and KO control group （ all P 〉 0.05 ）. Immunohistochemistry staining revealed that IFN-γ and MyD88 expressed almost alike in ISO and KO group, but there was a significant decreasing in ISO and KO group compared with WT group. No TLR2 could be detected in KO group, while expressed weakly in ISO group, and in- creased in WT group. The mRNA expression of IFN-γ and MyDg8 in ISO and KO group were nearly the same （ all P 〉 0.05 ）, but there was a marked decrease in ISO and KO group while compared with WT group（P 〈0.05）. The MyD88 mRNA expr

关 键 词：TOLL样受体2 基因 MYD88 角膜移植 免疫排斥 CD4+T细胞 干扰素-γ 

分 类 号：R772.2[医药卫生—眼科]