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目的探讨不同浓度丹参酮ⅡA(TanⅡA)对急性期川崎病(KD)患儿外周血单个核细胞(PBMC)核转录因子-κB(NF-κB)活化及肿瘤坏死因子-α(TNF-α)、白介素-8(IL-8)表达的影响。同时探讨KD发病机制和了解TanⅡA抗炎作用效果。方法研究对象为2010年12月至2011年5月成都市妇女儿童中心医院住院的KD患儿20例(KD组),同期门诊健康体检儿童20例为健康对照组,采用Ficoll密度梯度离心法分离静脉血PBMC,并分组进行培养,即自然培养组(空白)、12-肉豆蔻酰-13-乙酸佛波酯(PMA)、PMA+不同浓度TanⅡA(TanⅡA终浓度分别为1、3、10mg/L),采用免疫细胞化学法观察培养后各组细胞中NF-κB的活化率,通过双抗体酶联免疫吸附法(ELISA)检测各组培养上清液中TNF-α、IL-8的表达量。结果 KD组不同培养条件各指标均高于健康对照组(P<0.05)。加入PMA培养后,KD组和健康对照组各指标均明显高于空白条件,差异有统计学意义(P均<0.05)。加入PMA+3mg/LTanⅡA培养后,健康对照组TNF-α的表达量无明显下降(P>0.05);NF-κB的活化率与TNF-α和IL-8的表达量呈正直线相关关系(r=0.817,P<0.01;r=0.782,P<0.01)。TNF-α与IL-8的表达改变相一致,亦呈正直线相关关系(r=0.709,P<0.01)。结论急性期KD患儿PBMCs中NF-κB活化增强及TNF-α、IL-8表达量增加,参与了急性期KD免疫炎性反应,可能介导了急性期KD免疫血管损伤;TanⅡA在急性期KD患儿PBMCs中的抗炎作用具有剂量依赖性,其抗炎作用机制可能是通过抑制NF-κB的活化,进而抑制其下游TNF-α、IL-8的表达而实现。
Objective To investigate the effects of different concentrations of TanⅡA on activation of nuclear factor-κB (NF-κB) and expression of tumor necrosis factor-α (TNF-α) in peripheral blood mononuclear cells in children with Kawasaki disease (KD) Interleukin-8 (IL-8) expression. At the same time to explore the pathogenesis of KD and understand Tan Ⅱ A anti-inflammatory effect. Methods Twenty patients (KD group) with KD admitted to Chengdu Women and Children Hospital from December 2010 to May 2011 were enrolled in this study. Twenty healthy children were enrolled as healthy control group in the same period and were separated by Ficoll density gradient centrifugation (PMA), PMA + different concentrations of Tan Ⅱ A (Tan Ⅱ A final concentrations were 1, 3, 10 mg / L ). The rate of activation of NF-κB in each group was observed by immunocytochemistry. The expression of TNF-α and IL-8 in supernatants of each group was detected by double antibody enzyme-linked immunosorbent assay (ELISA) . Results The indexes of KD group under different culture conditions were higher than those of healthy control group (P <0.05). After adding PMA, the indexes in KD group and healthy control group were significantly higher than those in blank control group (P <0.05). After adding PMA + 3mg / LTanⅡA, the expression of TNF-α in the healthy control group had no significant difference (P> 0.05); The activation rate of NF-κB was positively correlated with the expression of TNF-α and IL- = 0.817, P <0.01; r = 0.782, P <0.01). TNF-α was consistent with the change of IL-8 expression, also showed a positive correlation (r = 0.709, P <0.01). Conclusions The activation of NF-κB and the increase of TNF-α and IL-8 in PBMCs of acute KD children are involved in the acute KD immune-inflammatory reaction, which may mediate the acute KD immune vascular injury. In the acute stage, The anti-inflammatory effect of PBMCs in KD patients is dose-dependent, and its anti-inflammatory mechanism may be through inhibiting the activation of NF-κB, and then inhibiting the expression of TNF-α and IL-8 downstream of PBMCs.