四川大学学报(医学版)
四川大學學報(醫學版)
사천대학학보(의학판)
JOURNAL OF WEST CHINA UNIVERSITY OF MEDICAL SCIENCES
2010年
1期
35-38
,共4页
雷松%岳利民%毛咏秋%杨莉
雷鬆%嶽利民%毛詠鞦%楊莉
뢰송%악이민%모영추%양리
MHC-Ig/肽复合体%实验性自身免疫性葡萄膜炎%细胞毒T细胞
MHC-Ig/肽複閤體%實驗性自身免疫性葡萄膜炎%細胞毒T細胞
MHC-Ig/태복합체%실험성자신면역성포도막염%세포독T세포
MHC-Ig/peptide polymer%EAU%CTLs
目的 探讨MHC-Ig/肽复合体用于小鼠实验性自身免疫性葡萄膜炎(EAU)特异性细胞毒性T淋巴细胞(CTLs)研究的应用价值.方法 用人类光感受器间维生素A类结合蛋白(IRBP)合成多肽IRBP161-180免疫B10RⅢ小鼠制备EAU动物模型.将源于IRBP161-180多肽序列的10种不同短肽片段与MHC-Ig多聚体形成复合体,用于检测BIORⅢ小鼠EAU中特异性CTLs.比较各种MHC-Ig/IRBP短肽复合体与CTLs的结合能力,以及刺激CTLs的增殖和lFN-γ的表达水平,并选用对CTLs作用最强的复合体,检测小鼠在EAU不同发病期外周血CTLs的表达情况.结果 各MHC-Ig/IRBP短肽复合体能够检测出抗原特异CTLs,其中MHC-Ig/IRBP168-177短肽复合体检测出CTLs的阳性率达12.3%;并且其刺激CTLs的增殖和IFN-γ的表达水平明显高于其它短肽组(P＜0.01),可初步推断短肽序列IRBP168-177为该EAU模型特异CTLs主要的抗原识别表位.用MHC-Ig/IRBP168-177短肽复合体检测EAU疾病过程中外周血CTLs的表达发现,EAU在炎症急性期特异性CTLs的阳性表达最高(4.9%±1.1%).结论 MHC-Ig/肽复合体技术,是一种新的定量分析抗原特异性CTLs的方法 ,能检测抗原特异性CTLs的表达情况,分析CTLs的抗原识别表位,为动物模型EAU的研究提供了高效、特异、敏感的检测手段.
目的 探討MHC-Ig/肽複閤體用于小鼠實驗性自身免疫性葡萄膜炎(EAU)特異性細胞毒性T淋巴細胞(CTLs)研究的應用價值.方法 用人類光感受器間維生素A類結閤蛋白(IRBP)閤成多肽IRBP161-180免疫B10RⅢ小鼠製備EAU動物模型.將源于IRBP161-180多肽序列的10種不同短肽片段與MHC-Ig多聚體形成複閤體,用于檢測BIORⅢ小鼠EAU中特異性CTLs.比較各種MHC-Ig/IRBP短肽複閤體與CTLs的結閤能力,以及刺激CTLs的增殖和lFN-γ的錶達水平,併選用對CTLs作用最彊的複閤體,檢測小鼠在EAU不同髮病期外週血CTLs的錶達情況.結果 各MHC-Ig/IRBP短肽複閤體能夠檢測齣抗原特異CTLs,其中MHC-Ig/IRBP168-177短肽複閤體檢測齣CTLs的暘性率達12.3%;併且其刺激CTLs的增殖和IFN-γ的錶達水平明顯高于其它短肽組(P＜0.01),可初步推斷短肽序列IRBP168-177為該EAU模型特異CTLs主要的抗原識彆錶位.用MHC-Ig/IRBP168-177短肽複閤體檢測EAU疾病過程中外週血CTLs的錶達髮現,EAU在炎癥急性期特異性CTLs的暘性錶達最高(4.9%±1.1%).結論 MHC-Ig/肽複閤體技術,是一種新的定量分析抗原特異性CTLs的方法 ,能檢測抗原特異性CTLs的錶達情況,分析CTLs的抗原識彆錶位,為動物模型EAU的研究提供瞭高效、特異、敏感的檢測手段.
목적 탐토MHC-Ig/태복합체용우소서실험성자신면역성포도막염(EAU)특이성세포독성T림파세포(CTLs)연구적응용개치.방법 용인류광감수기간유생소A류결합단백(IRBP)합성다태IRBP161-180면역B10RⅢ소서제비EAU동물모형.장원우IRBP161-180다태서렬적10충불동단태편단여MHC-Ig다취체형성복합체,용우검측BIORⅢ소서EAU중특이성CTLs.비교각충MHC-Ig/IRBP단태복합체여CTLs적결합능력,이급자격CTLs적증식화lFN-γ적표체수평,병선용대CTLs작용최강적복합체,검측소서재EAU불동발병기외주혈CTLs적표체정황.결과 각MHC-Ig/IRBP단태복합체능구검측출항원특이CTLs,기중MHC-Ig/IRBP168-177단태복합체검측출CTLs적양성솔체12.3%;병차기자격CTLs적증식화IFN-γ적표체수평명현고우기타단태조(P＜0.01),가초보추단단태서렬IRBP168-177위해EAU모형특이CTLs주요적항원식별표위.용MHC-Ig/IRBP168-177단태복합체검측EAU질병과정중외주혈CTLs적표체발현,EAU재염증급성기특이성CTLs적양성표체최고(4.9%±1.1%).결론 MHC-Ig/태복합체기술,시일충신적정량분석항원특이성CTLs적방법 ,능검측항원특이성CTLs적표체정황,분석CTLs적항원식별표위,위동물모형EAU적연구제공료고효、특이、민감적검측수단.
Objective To explore the application of MHC-Ig/peptide polymer technique for detecting antigenspecific cytotoxic T lymphocytes(CTLs)in mice with experimental autoimmune uveitis(EAU).Methods B10RⅢ mice were immunized with an interphotoreceptor retinal-binding protein(IRBP)synthetic peptide(IRBP161-180).The in vivo primed T cells were separated and stained with MHC-Ig polymer combined with a panel of truncated peptides derived from IRBP161-180.The level of IRBP-specific CTLs cells was determined by FACS analysis.The CD8~+T cells were isolated from the primed T cells and stimulated with complex polymers containing MHC-Ig and various IRBP-derived peptides.The proliferation of CD8~+ T cells was measured by ~3H thymidine incorporation.The production of interferon-(IFN-)in the cell suspensions was measured by ELISA.Results The IRBP-specific CTLs were detected by MHC-Ig/peptide polymers.The MHC-Ig/IRBP168-177 peptide polymer dtected 12.3%specific CTLs,showing greater ability in stimulating proliferation of CTLs and production of IFN-than the other MHC-Ig/peptide polymers(P＜0.01).The truncated 10-mer peptide,IRBP168-177,was the major antigenic epitope for the IRBP-specific CTLs.The MHC-Ig/IRBP168-177 peptide polymer detected the highest level(4.9%± 1.1%)of specific CTLs from peripheral blood mononuclear cells(PBMCs)at the acute stage of EAU.Conclusion The MHC-Ig polymer technique is an effective instrument for detecting antigen-specific CTLs,with good sensitivity and specificity in EAU studies.
