细胞因子信号传导抑制因子在实验性自身免疫性葡萄膜视网膜炎视网膜内的表达

目的 探讨细胞因子信号传导抑制因子(SOCS)在实验性自身免疫性葡萄膜视网膜炎(EAU)视网膜内的表达与意义.方法 随机对照实验设计方法.90只Lewis大鼠采用分层随机抽样进行分组,分别为未治疗组40只大鼠、治疗组40只大鼠及正常对照组10只大鼠.用光感受器间维生素A类结合蛋白(IRBP)诱导EAU动物模型,治疗组从免疫后1～28 d,给予环孢素(CsA)灌胃,每天20 mg/kg体重,而未治疗组给予等量生理盐水,正常对照组未给予免疫和治疗.分别于免疫后7、14、21、28 d按分层随机号抽取治疗组与未治疗组各10只大鼠进行相关枪测,观察大鼠眼部炎性变化,双眼取材,分别用实时荧光定量聚合酶链反应和蛋白免疫印迹法检测大鼠视网膜内SOCS mRNA和蛋白的表达,酶联免疫吸附试验方法检测血清细胞因子白细胞介素4(IL-4)、IL-12、干扰素γ(IFN-γ)的水平.利用单因素方差分析对治疗组与未治疗组大鼠血清中细胞因子的浓度和视网膜内SOCS蛋白表达量进行对比分析,多重比较采用最小显著差异法;用Mann-Whitney两样本比较秩和检验进行治疗组与未治疗组大鼠炎性反应分值比较.结果 免疫后14 d,未治疗组可见明显的虹膜睫状体炎,房水闪辉阳性,虹膜后粘连,瞳孔不规则甚至前房积脓,炎性反应分值是1.5±0.5;而治疗组眼部炎性反应不明显,炎性反应分值是0.5±0.3.未治疗组的IL-12、IFN-γ及IL-4免疫后14 d达到最高峰(均P＜0.05),免疫后28 d IL-12、IFN-γ下降到正常水平(均P＞0.05),IL-4仍高于正常对照组(P＜0.05);治疗组中IL-12、IFN-γ的动态变化不明显(均P＞0.05),IL-4在整个病程中呈上升趋势(均P＜0.05).SOCS1、含SH2结构的细胞因子诱导蛋白(CIS)mRNA免疫后14 d表达最高,未治疗组分别是正常对照组的3.41倍和3.36倍,治疗组分别为1.44倍和1.73倍;SOCS3和SOCS5mRNA在整个病程中逐渐升高,免疫后28 d表达最高,未治疗组中分别为正常对照组的1.95倍和3.16倍,治疗组中分别为正常对照组的1.59倍和3.58倍.未治疗组SOCS1和CIS蛋白在免疫后7、14、21 d时显著高于正常对照组(均P＜0.05),治疗组二者免疫后14 d显著高于正常对照组(均P＜0.05);两组中SOCS3和SOCS5蛋白在免疫后均显著高于正常对照组(均P＜0.05).结论 EAU发生过程中视网膜SOCS1升高与Th1型反应增强有关;在发病的不同阶段中CIS与SOCS3升高和Th2细胞反应增强以对抗Th1细胞反应,与缓解葡萄膜炎的发病强度有关;SOCS5水平升高在眼局部炎性反应发生过程中可能起保护作用.

Expression of suppressor of cytokine signaling in recta of experimental autoimmune uveoretinitis

Objective To investigate the expression and significance of suppressor of cytokine signaling(SOCS)mRNA and protein in retina of rats with experimental autoimmune uveoretinitis(EAU).Methods Research was designed with randomized controlled trials.Eighty Lewis rats were immunized with interphotoreceptor retinoid-binding protein(IRBP)to induce EAU.They were stratified randomly into two groups:nontreatment group(40 rats)and treatment group(40 rats).The treatment group wss administered saline buffer in the same way,ten normal Lewis rats without immunization and treatment as normal control group.The rats in both groups were stratified randomly into four sub-groups.Each sub-group had 10 rats and the rats were sacrificed at day 0,7,14,21,28 after immunization. All eyes were evaluated by slit-lamp microscopy.Retinas of rats were snbjected to analysis of SOCS mRNA and protein expression using quantitative RT-PCR and western blot,respectively.IL-4,IL-12 and IFN-γ in the serum were measured by ELISA.Results At day 14,the nontreatment group developed acute anterior uveitis and showed typical signs including aqueous flare,small or irregular pupil due to posterior synechiae and hypopyon,the score of inflammation was 1.5±0.5:but the treatment group did not suffer from severe anterior chamber inflammation,the score of inflammation was 0.5±0.3.The highest level of IL-4.IL-12 and IFN-γ production occurred at day 14(P＜0.05),followed by decline in the secretion of IL-12 and IFN-γ secretion to the baseline at day 28,while the concentration of IL-4 did not decrease significantly in the nontreatment group(P＞0.05).IL-12 and IFN-γ did not change significantly(P＞0.05)and IL-4 production reached the highest level at day 28 in the treatment group.Both SOCS1 and CIS mRNA increased to the highest level at day 14.Compared to normal controls,the copies of SOCS1 and CIS mRNA were 3.41 fold and 3.36 fold in the nontreatment group,and were only 1.44 fold and 1.73 fold in treatment group,respectively.Both SOCS3 and SOCS5 mRNA reached the highest level at day 28 after immunization in both groups.The copies of SOCS3 and SOCS5 mRNA were 1.95 fold and 3.16 fold in the nontreatment group,and were 1.59 fold and 3.58 fold in the treatment group,respectively.Marked increased expression of SOCS1 and CIS proteins in the nontreatment group as compared to normal controls was detected at 7,14,21 days;in the treatment group these proteins only increased significantly at day 14(P＜0.05).Expression of SOCS3 and SOCS5 proteins in both groups were higher than those in normal controls(P＞0.05).Conclusions Marked increased expression of SOCS1 of retina may involve in the development of a Th1 mediated immune response in EAU.CIS and SOCS3 of retina may play important roles in mitigating pathogenic effects of proinflammatory cytokines during different stages of EAU.Up-regulation of SOCS5 may have protective functions.