摘要
目的探讨核细胞核因子-κB(NF-κB)和基质金属蛋白酶9(MMP-9)在川崎病发生、发展中的作用及其机制。方法研究对象为川崎病患儿21例(川崎病组)、呼吸道感染患者18例(阳性对照组)、正常查体者17例(正常对照组)。采用密度梯度离心法无菌分离各组外周血PBMCs自然培养,采用RT-PCR法和Westen blot法测定细胞中NF-κB p65 mRNA和蛋白,采用ELISA方法测定细胞上清液中MMP-9蛋白。将川崎病组外周血PBMCs体外培养并分为川崎病1、2、3组,川崎病1组继续自然培养,川崎病2组加入终浓度为20 nmol/L的NF-κB激动剂PMA,川崎病3组同时加入PMA(浓度同上)及终浓度为100μmol/L的NF-κB抑制剂PDTC,孵育2 h后收集上清液和细胞,分别采用RT-PCR法和Westen blot法测定NF-κB p65 mRNA和蛋白表达,采用ELISA法测定细胞上清液中MMP-9蛋白水平。结果川崎病组PBMCs中NF-κB p65 mRNA、蛋白及上清液MMP-9表达均高于阳性对照组和健康对照组(P均<0.05)。川崎病1、3组PBMCs中NF-κB p65蛋白及上清液中MMP-9表达水平均低于川崎病2组(P均<0.05)。结论急性期川崎病患儿外周血PBMCs中NF-κB和细胞上清液中MMP-9表达增强;二者可能参与了川崎病的发生发展过程。。
Objective To explore the activation of nuclear factor-kappaB （ NF-κB ） and matrix metalloproteases 9 （MMP-9） in peripheral blood mononuclear cells （PBMCs）in children with Kawasaki disease and the relationship between NF-κB and MMP-9.Methods PBMCs were isolated and purified from blood of fifty-six children including 21children with KD, 18 children with general inflammatory and 17 normal ones by density gradient centrifugation ,and were cultured for one hour after cell concentration adjusted to 10^6/mL.Then both KD group and Control group were divided into three groups ,the first group was cultured naturally , the second one was stimulated by phorbol 12-myristate 13-acetate（PMA）, and the third one was stimulated by PMA and pyrrolidine dithiocarbamate （ PDTC） .PBMCs were cultured for 2 h.The mRNA and pro-tein levels of NF-κB p65 in PBMCs in each groups were measured by RT-PCR and Western blot , and the protein levels of MMP-9 were measured by ELISA .Results The mRNA and protein levels of NF-κB p65 and MMP-9 in KD group were much higher than that in other groups;Stimulating PBMCs by PMA ,the mRNA and protein levels of NF-κB p65 and MMP-9 were markedly increased in each group , and there was a significant difference in each group （P〈0.05）; Using PDTC can reduce the activation of NF-κB in PBMCs, and the protein levels of MMP-9 were significantly reduced by using PDTC in KD group.Conclusions The activity of NF-κB and MMP-9 in PBMCs in patients with acute KD is distinctly increased;The specific inhibitor of NF-κB can inhibit it＇s activation and the expression of MMP-9 obviously.It suggested that the NF-κB pathway can lead to the activation of MMP-9 in acute KD, and proper application of specific inhibitor of NF-κB may be a new choice for treating KD .
出处
《山东医药》
CAS
北大核心
2015年第6期11-13,共3页
Shandong Medical Journal
关键词
川崎病
核因子-ΚB
基质金属蛋白酶9
外周血单个核细胞
Kawasaki disease
nuclear factor-kappaB
matrix metalloproteases 9
peripheral blood mononuclear cells
