富碘中药复方与碘过量对碘缺乏NOD.H-2^(h4)小鼠甲状腺Th17细胞分化影响的比较

A Comparative Study on the Effect of Iodine-rich Herbal Decoction and Iodine Excess on the Differentiation of Th17 cells in Iodine-deficiency NOD.H-2h4 Mice

目的比较富碘中药复方与碘过量对碘缺乏NOD.H-2h4小鼠甲状腺IL-23/IL-17炎症轴的影响。方法选雌性8周龄NOD.H-2h4小鼠,给低碘饲料,饮双蒸水,喂养90天后制成自身免疫甲状腺炎易感碘缺乏甲状腺肿模型,后随机分成4组:正常对照组(NC)、模型对照组(MC)、富碘中药复方组[(HIE),海藻15g,昆布15g,海带7.5g,碘含量1900.36μg/L]和碘过量组[(IE)含碘1900μg/L的去离子水,用碘酸钾配制]。再给处理因素90 d后处死小鼠。放射免疫分析法(RIA法)测小鼠血清总T3(TT3)、总T4(TT4)。IRMA法测定小鼠血清促甲状腺激素(TSH)。光镜下观察甲状腺滤泡形态。用免疫组化方法测定甲状腺间质细胞CD68及IL-17的表达。实时定量(RT-PCR)法检测NOD小鼠甲状腺IL-17mRNA和IL-23mRNA表达。Western blot蛋白印迹杂交法测定小鼠甲状腺IL-17和IL-23的蛋白表达。结果给处理因素90 d后,各组间血清TT3﹑TT4及TSH值比较均无明显差异(P﹥0.05)。NC,MC,IE及HIE组小鼠自身免疫甲状腺炎的发生率分别为:2/10,0/10,9/10,3/10。与IE组相比,HIE组的甲状腺间质细胞CD68的表达水平(整合光密度0.28±0.07 vs 0.13±0.06)明显降低(P﹤0.01);HIE组小鼠甲状腺组织中IL-17的表达水平(整合光密度0.21±0.07vs 0.19±0.05)明显降低(P﹤0.01)。HIE组IL-17mRNA及IL-23mRNA平均相对表达量(1.31±0.06;1.43±0.50),明显低于IE组(2.39±0.05;2.21±0.70),P均<0.05。HIE组的IL-17及IL-23的蛋白表达量(0.73±0.02;0.27±0.02),明显低于IE组(0.80±0.03;0.36±0.03),P均﹤0.05。结论与单纯碘过量相比,富碘中药复方能明显降低碘缺乏NOD.H-2h4小鼠碘致自身免疫甲状腺炎的发生率,其机理可能与富碘中药复方能明显抑制碘补充过程中甲状腺内IL-23、IL-17的高表达有关。

Objective To compare the effect of iodine - rich herbal decoction and iodine excess on the inflammatory axis of IL - 23/IL - 17 cells in NOD. H - 2h4 mice with deficiency iodine. Methods 8 - week - old NOD. H - 2h4 female mice were made common animal model of susceptible autoimmune thyroiditis and goiter caused by iodine deficiency by feeding a low iodine diet and double distilled water with 90 days, randomly divided into four groups after the success of modeling：the normal control group （ NC）, the model control group （ MC）, the iodine - rich herbal decoction group （HIE） （ The ingredients of HIE group include Haizao 15 g, Kunbu 15g, Haidai 7.5g with the iodine concentration of 1900. 361~g/L） and the iodine excess group （IE） （the IE group is made of potassium iodate with the iodine concentration of 19001.Lg/L）. After 90days of giving treatment factors ,the mice were killed. The levels of serum total triiodothyronine （rIT3） and total thyroxine （TY4） were measured by radi Y （RIA）. Meanwhile the mice serum thyroid stimulating hormone （TSH）were measured by radioimmunoassay （IRMA）. The mor- phology of thyroid cells were observed under optical microscopy. The expression of CD68 in thyroid stroma cells and IL - 17 were detected with immunohistochemical method. The expressions of IL - 17 mRNA and IL - 23 mRNA in the thyroid of NOD. H - 2114 mice were determined by RT - PCR method. The expression of IL - 17 and IL - 23 in the thyroid of NOD. H - 2h4 mice were determined by the Western blot method. Results 90 days after treatment by different factors, there was not obvious difference among every group on serum TT3 ,T Y4 and TSH （P 〉 0.05）. The prevalence of autoimmune thyroiditis on the mice in the NC group, the IE group,the MC group and the HIE group is respectively 2/10,9/10,0/10 and 3/10. Compared with the IE group,the expression of CD68 in thyroid stroma cells in the HIE group was significantly decreased （the integrated optical density：0.28 ±0.07 vs 0.13±0.06 ） （ P 〈 0.01 ） and the expression of IL - 17 in thyroid tissue in the HIE group was obviously decreased（ the integrated optical density： 0.21 ±0.07 vs 0.19 ±0.05） （P 〈 0.01 ）. The mean relative expression of IL - 17 mRNA and IL - 23 mRNA in the HIE group （ 1.31 ± 0.06 ; 1.43 ± 0.50） is significantly less than in the IE group （2.39 ± 0.05 ;2.21 ± 0.70） （ P 〈 0.05 ）. The protein expression of IL- 17 and IL- 23 in the HIE group（0.73 ± 0.02 ;0.27 ± 0.02）is significantly less than in the IE group （0.80 ±0.03 ;0.36 ±0.03） （P 〈 0.05）. Conclusion Compared with the iodine excess,rich iodine Chinese herbal medicine decoction may obviously reduce the incidence of autoimmune thyroiditis in iodine deficiency NOD. H - 2h4 mice , and the mechanism may be related to obvious inhibition of high expression of IL - 23 and IL - 17 in thyroid.