雷帕霉素对大鼠实验性自身免疫性葡萄膜炎的治疗作用

Therapeutic effects of rapamycin on experimental autoimmune uveoretinitis

背景雷帕霉素不仅具有抗菌作用，而且是一种良好的免疫抑制剂，可用于多种自身免疫性疾病的治疗．对实验性自身免疫性葡萄膜炎（EAU）的治疗作用是目前研究的热点之一。目的研究雷帕霉素对EAU的治疗作用，并观察雷帕霉素对EAU各免疫细胞群炎性因子表达的影响。方法25只Lewis大鼠采用随机数字表法分为EAU组（20只）和正常对照组（5只）。光感受器间维生素A类结合蛋白（IRBP）R16肽段与完全氟氏佐剂充分乳化后于Lewis大鼠后足皮下注射以建立EAU模型，EAU模型鼠再按分层随机的原则分为模型对照组和雷帕霉素组，每组10只大鼠。雷帕霉素组造模后即应用O．2mg／（kg·d）雷帕霉素（0．4m1）腹腔内连续注射7d，模型对照组及正常对照组大鼠采用等体积的生理盐水进行腹腔内注射。造模后第4天开始每日裂隙灯下观察大鼠EAU的症状，造模后第14天制备大鼠视网膜切片，以苏木精-伊红染色法进行组织病理学观察，参照Caspi的标准对EAU症状及组织病理学分级进行评分。应用免疫组织化学染色法检测各组大鼠视网膜中炎性因子干扰素-γ（IFN-γ）、白细胞介素-17（IL-17）的表达情况。结果造模后6d模型对照组大鼠EAU炎症评分逐渐升高，12d达到高峰，然后逐渐下降。雷帕霉素组大鼠EAU炎症评分变化呈现相同的趋势，但各时间点EAU炎症评分均明显低于模型对照组，差异均有统计学意义（P〈0．01）。视网膜组织病理学研究表明，模型对照组大鼠视网膜结构紊乱，大量炎性细胞浸润，组织病理学评分为3．30±0．48，而雷帕霉素组视网膜结构接近正常，组织学评分为0．90±0．45，差异有统计学意义（t=16．541，P〈0．01）。雷帕霉素组IFN-γ、IL-17在大鼠视网膜中的表达量（A值）分别为21．16±4．23和49．86±6．59，明显低于模型对照组的62．14±7．32和124．85±6．33，差异均有统计学意义（q=33．334、q=56．923，P〈0．01）。结论雷帕霉素通过抑制EAU视网膜中IFN-γ、IL-17等炎性因子的表达而对EAU发挥治疗作用，其机制可能是通过抑制Th1、Th17细胞群来实现的。

Background Studies determined that rapamycin has not only the antibiosis but also suppressing the auto-immunology. The treating effect of rapamyein on experimental autoimmune uveoretinitis （EAU） is still concerned. Objective This work was to investigate the therapeutic effect of rapamycin on EAU and study the effect on the expression of inflammatory cytokines which were secreted by T lymphocyte subgroup in EAU. Methods EAU was induced in 20 SPF male Lewis rats by subcutaneous injection of interphotoreceptor retinoid binding protein （IRBP） RI6 peptide emulsified in adjuvant. The rats were randomized into model control group and rapamycin injection group and 10 rats for each group. The rapamycin of 0.2 mg/（ kg· d） 0.4 ml was intraperitoneally injected for the consecutive 7 days immediately after modeling, and the equal volume of normal saline solution was used at the same fashion in the model control group and 5 normal matched rats（ normal control group）. The ocular manifestation of the rats were examined under the slit lamp regularly. The retinal sections of the rats were prepared in the 14 days after modeling for the histopathological examination with hematoxylin and eosin staining. The ocular signs of inflammation and histopathological severity were scored based on the criteria of Caspi. Expression of interferon-γ （IFN-γ） and interleukin-17 （IL-17） in rat retinas were detected by immunohistochemistry. This experiment followed the Regulation for the Administration of Affair Concerning Experimental Animals by Tianjin Municipal Government. Results The scores of ocular signs elevated gradually from 6 through 12 days after modeling and slowly declined after that in the model control group. A same tendency was seen in the rapamyein injection group. The significant differences were seen in the scores of ocular signs between the two groups from 6 to 14 days（P〈0. 01 ）. The disorder of retinal structure and infiltration of inflammatory cells were seen in the model control group, but the retina layers were normal in the rapamycin injection group. The pathological score was evidently declined in the rapamycin injection group （0. 90± 0. 45 ） in comparison with model control group （3.30±0.48） （ t = 16. 541 , P〈0. 01 ）. The expressions of the IFN-＇,/and IL-17 in retina located in the outer nuclear layer,inner nuclear layer,internal plexiform layer and retinal ganglion cell layer with the weakened levels （A values） in rapamycin injection group compared with model control group, showing a considerably difference between them （ IFN-γ： 21. 16 ± 4.23 vs 62. 14± 7.32 ; IL- 17 ： 49.86 ± 6.59 vs 124. 85_± 6.33 ） （q=33. 334,q=56. 923 ,P〈0. 01 ）. Conclusions Rapamycin down-regulates the expressions of IFN-γ and IL-17 in retina and further eliminates the inflammatory response in the rat with EAU by the suppression of Thl and Thl7 cells function in EAU.