益气化痰活血方对NOD.H-2^h4小鼠甲状腺Th17细胞分化影响的研究

Influence of Yiqi Huatan Huoxeu Fang on differatiation of Th17 cells in NOD.H-2^(h4) mice

目的探讨益气化痰活血方对NOD.H-2^(h4)小鼠甲状腺白细胞介素-23(IL-23)/白细胞介素-17(IL-17)炎症轴的影响。方法选雌性8周龄NOD.H-2^(h4)小鼠,随机分成正常组、模型组、益气化痰活血方低剂量组(20.8 g/kg)和高剂量组(62.4 g/kg)。正常组饲以双蒸水,其余各组饲以含0.05%碘化钠水,8周后制成自身免疫性甲状腺炎(AIT)动物模型,再处理因素8周后麻醉处死小鼠。放射免疫分析法(RIA)检测血清总T3(TT3)、总T4(TT4),固相化学发光酶免疫法(IRMA法)检测血清促甲状腺激素(TSII);光镜下观察甲状腺细胞形态;免疫组化方法测定甲状腺间质细胞CD68及小鼠甲状腺IL-17、IL-23蛋白表达;RT-PCR法检测小鼠甲状腺IL-17、IL-23 mRNA表达;流式细胞仪分析小鼠脾细胞Th17细胞比例。结果各组间血清IT3、TT4及TSH值比较无明显差异(P>0.05);与正常组比较,模型组AIT的发生率、CD68、IL-17、IL-23表达及Th17细胞比例显著升高(P<0.01);与模型组比较,益气化痰活血方不同剂量组CD68、IL-17、IL-23表达显著下降(P<0.01),Th17细胞比例降低(P<0.01或P<0.05)。结论益气化痰活血方治疗AIT的机制可能与其抑制IL-23/IL-17炎症轴,下调Th17细胞比例有关。

Objective To investigate the influence of Yiqi Huatan Huoxeu Fang on inflammatory axis of thyroid interlukin-23 （IL-23）/interlukin-17 （IL-17） in NOD. H-2h4 mice. Methods Female NOD. H-2h4 mice （8 weeks old ） were chosen and divided into normal group, model group, low-dose Yiqi Huatan Huoxeu Fang group （low-dose group, 20.8 g/kg） and high-dose Yiqi Huatan Huoxeu Fang group （high-dose group, 62.4 g/kg）. Normal group was given double distilled water and other groups, Nal water （0.05 % ）. After 8 weeks, the model of autoimmune thyroiditis （AIT） was established and intervened for 8 weeks. The levels of serum total T3 （TT3） and total T4 （TF4） were detected by using radioimmunoassay （ RIA）, and level of serum thyroid stimulating hormone （TSH） was detected by using solid-phase chemi-luminescent enzyme immunoassay. The morphological changes of thyroid cells were observed with light microscope. The protein expressions of thyroid mesenchymal CD68, and thyroid IL-17 and IL-23 were detected by using immunohistochemistry technique, mRNA expressions of IL-17 and IL-23 were detected by using RT-PCR, and percentage of splenocyte Thl7 was analyzed by using flow eytometry. Results The levels of TT3, TT4 and TSH had no significant difference compared among all groups （P 〉 0.05）. The incidence of AIT, expressions of CD68, IL-17 and IL-23 and percentage of Thl7 all increased significantly in model group compared with normal group （P 〈 0.01 ）. The expressions of CD68, IL-17 and IL-23 decreased significantly （P 〈0.01 ） and percentage of Th17 decreased （P 〈 0.01 or P 〈0.05） in high-dose group and low-dose group compared with model group. Conclusion The mechanism of Yiqi Huatan Huoxeu Fang for treating AIT may be related to that it can inhibit the inflammatory axis of IL-23/IL-17 and down-regulate percentage of Th17.