GSK-3β抑制剂对类风湿性关节炎小鼠脾脏树突状细胞亚群的影响

Effect of GSK-3β Inhibitor on Dendritic Cell Subsets in Spleen of Mice with Rheumatoid Arthritis

目的:观察糖原合成酶激酶-3β(GSK-3β)抑制剂对胶原诱导的关节炎(CIA)小鼠的治疗作用及其与树突状细胞(DC)亚群变化的相关性。方法:取6~8周龄近交系雄性DBA/l小鼠19只,分为正常组(n=5)、模型组(n=7)及治疗组(n=7),模型组与治疗组小鼠制作CIA模型,治疗组小鼠于造模后第22天开始每天予GSK-3β抑制剂(TDZD-8)5 mg/kg,模型组与正常组均予等量生理盐水,连续注射2周;观察和比较各组小鼠足趾肿胀程度及关节炎指数(AI)评分;第46天脱颈处死各组小鼠后切取右后肢踝关节,采用苏木素-伊红(HE)染色观察各组小鼠关节的组织学特征;采用免疫组织化学检测各组小鼠关节滑膜组织GSK-3β表达,采用流式细胞术检测各组小鼠脾脏树突状细胞(DC)中淋巴组织驻留型DC(cDC)及其CD8^+cDC、CD4^+CD8^-cDC、CD4^-CD8^-cDC各亚群变化。结果:与模型组比较,治疗组小鼠踝关节处红肿、出血点等现象明显好转;与模型组相比,治疗组小鼠AI评分在初次免疫后第33天开始下降,差异有统计学意义(P<0.05或P<0.01);HE染色结果显示为关节腔炎症状态明显改善;治疗组小鼠踝关节GSK-3β蛋白表达低于模型组,高于正常组,差异均有统计学意义(P<0.05);治疗组小鼠脾脏总cDC、CD4^+cDC比例高于模型组,低于正常组,差异均有统计学意义(P<0.05);治疗组小鼠脾脏CD4^-CD8c^-cDC比例低于模型组,高于正常组,差异均有统计学意义(P<0.05)。结论:GSK-3β抑制剂可通过调节CIA小鼠脾脏cDC数量及各亚群的比例,缓解其关节病变及其病理表现。

Objective:To observe the therapeutic effect of glycogen synthase kinase-3β(GSK-3β)inhibitor on collagen-induced arthritis(CIA)and its correlation with the changes of dendritic cell subsets.Methods:The male DBA/l mice of 6-8 week old were divided into the normal group(n=5),the model group(n=7)and GSK-3β treatment group(n=7).CIA models with reference to classical CIA model method for the model group and the treatment group were prepared.Then modeled mice were given 5 mg/kg TDZD-8 of reagents every day on the 22^nd day.The model group and the normal group were given the same amount of normal saline for 2 weeks.The scores of signs,toe swelling and arthritis index(AI)were observed and compared among the groups.On the 46^th day,the right hindlimb ankle joint was removed and the histological features of the joints were observed by hematoxylin-eosin(HE)staining,and the expression of synovial tissue GSK-3β was detected by immunohistochemistry,and the changes of lymphoid tissue retention DC(cDC)and its CD8^+cDC,CD4^+CD8^-cDC,CD4^-CD8^-cDC subsets in splenic dendritic cells were detected by flow cytometry.Results:Compared with the model group,GSK-3β treatment group mice diet,hair gloss,hair removal and joint redness,bleeding points and other phenomena were improved significantly.The AI scores of mice in the treatment group decreased more significantly on the 33^rd day after initial immunization than those in the model group,and the difference was statistically significant(P<0.05 or P<0.01).HE staining showed that the inflammatory state of articular cavity improved significantly.The expression of GSK-3β protein in the ankle joint in the treatment group was lower than that in the model group and higher than that in the normal group,and the difference was statistically significant(P<0.05).The proportion of total cDC,CD4^+cDC of spleen in the treatment group was higher than that in the model group and lower than that in the normal group,and the difference was statistically significant(P<0.05).The proportion of spleen CD4^-CD8c^-cDC in the treatment group was lower than that in the model group and higher than that in the normal group,and the difference was statistically significant(P<0.05).Conclusion:GSK-3β inhibitors can relieve joint lesions and pathological manifestations by regulating the number of cDC spleen and the proportion of each subgroup in CIA mice.