IL-10基因转染对EAT大鼠脾细胞增殖及Th1细胞因子分泌的影响  

The effect of IL-10 gene transmission into thyroid tissues on the PTg-stimulated splenocytes proliferation and Th1 cytokines production in rats

作　　者：唐伟[1] 刘超[1] 贾悦[1] 刘翠萍[1] 覃又文[1] 吕国良[1] 王昕[1] 孙敏[1] 茅晓东[1] 

机构地区：[1]南京医科大学第一附属医院内分泌科,江苏南京210029

出　　处：《中国药理学通报》2005年第10期1227-1231,共5页Chinese Pharmacological Bulletin

基　　金：江苏省科技青年基金资助项目(NoBQ2000017)

摘　　要：目的在建立EAT动物模型的基础上,探索通过非病毒载体基因转染方法,以改善EAT大鼠免疫功能异常状态,并观察对大鼠脾细胞增殖及分泌Th1细胞因子的影响。方法选用♀W istar大鼠建立EAT动物模型。成模大鼠分为4组:A组(PBS+PLL),B组(pORF质粒+PLL)、C组(pOR-Fm IL10质粒+PLL)及D组(pORFm IL10质粒+MEM)。使用R IA方法测定TgAb、TmAb滴度。脾脏淋巴细胞培养,采用[3H]-TdR标记方法测定cpm值并计算淋巴细胞增殖指数,ELISA方法检测细胞因子IFN-γ、TNF-α及IL-2。结果C组与A、B、D组比较,第4、6、8周TgAb、TmAb滴度降低,(P<0.01,F=42.66,F=32.65,F=9.66;F=22.25,F=81.35,F=14.84)。此外,淋巴细胞增殖试验结果显示,C组PTg刺激的淋巴细胞增殖指数低于A,B组,(P<0.05)。PTg刺激的脾细胞培养上清中细胞因子IFN-γ、TNF-α及IL-2检测结果,A、B组高于C组(P<0.01,P<0.05)。结论通过大鼠甲状腺组织内局部注射pORFm IL10质粒使甲状腺滤泡上皮细胞表达IL-10,能够降低自身抗体水平和针对抗原反应的T淋巴细胞增殖反应及PTg刺激的脾细胞产生的Th1细胞因子。多聚赖氨酸可以延长目的基因表达时间,更有效地改善EAT大鼠的免疫功能状态。Aim The autoimmune response in experimental autoimmune thyroiditis （EAT） is characterized by lymphocytes infiltration of the thyroid gland and evaluated by circulating autoantihodies specifically to thyroid antigen. Since IL-10 seems to be very effective in immunomodulation of EAT, we investigated local whether expression of IL-10 in the thyroid could alter the immune disorder condition. Methods Wistar female rats weighed 60 -80 g were immunized with PTg 100μg emulcificated with CFA subcutaneously. Enhanced immunization with the same dose of antigen emulcificated with IFA at 2 wk,3 wk and 4 wk. EAT rats were divided into four groups：group A （PBS ＋ PLL）, five rats, group B （pORF ＋ PLL） five rats, group C （pORFm IL10＋ PLL） ten rats, group D （pORFm IL10 ＋ MEM） five rats. The plasmid was mixed with lipofectamine, then injected into the thyroid tissues of rats on day 18 after-plasmid immunization. The rats were sacrificed at 8wk, thyroids were mored and fixed in 10% neutral formalin solution and processed according to standard procedures. The specimens were stained with hematoxylin- eosin-safran. The level of serum TgAb and TmAb were detected at 0, 2, 4, 6, 8 wk. In vitro proliferative responses to ConA and different concentrations of PTg were measured by culturing 4 × 10^5 spleen cells pulsed with 18.5 KBq of [^3H ] thymidine for the final 12h and then harvested for liquid scintillation counting. Thl cytokines IFN-γ, TNF-α and IL-2 were detected using ELISA. Results A dramatic lymphocytic infiltration was observed in thyroids of experimental autoimmune thyroiditis rats. The serum TgAb and TmAb of EAT rats increased at 2 wk after first immunization, but no statistical difference was found among the groups. The rats in group C had lower level of TgAb and TmAb than those of the other groups at 4,6,8 wk, （P 〈0.01,F =42.66, F =32.65, F=9.66; F=22.25, F=81.35,F=14.84, respectively）. The proliferative response to TG was suppressed in group C than that in groups A and B （P〈0.05）

关 键 词：IL-10 自身免疫 淋巴细胞增殖 基因治疔 

分 类 号：R-332[医药卫生] R329.24