小儿脓毒症Toll样受体信号途径转导分子及调节因子的变化研究

Role of transduce molecules and modulatory factors of signal pathways of Toll-like receptor in inflammatory response of children with sepsis

目的探讨Toll样受体（TLRs）信号途径转导分子及调节因子在小儿脓毒症异常炎症免疫反应发病机制中的可能作用。方法选择脓毒症患儿10例、严重脓毒症患儿13例及同期健康体检儿童17例。采用实时荧光定量聚合酶链反应（real—time PCR）检测前炎症细胞因子[肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）]，以及TLRs信号途径转导分子和调节因子的mRNA表达；采用酶联免疫吸附法（ELISA）检测前炎症细胞因子蛋白水平。结果与健康对照组比较，脓毒症组TNF-α、IL-1β和IL-6的mRNA和蛋白表达均增加，TLRs信号途径转导分子TLR2、TLR4、髓样细胞分化蛋白-88（MyD88）、TNF相关因子6（TRAF6）、IL-1受体相关激酶4（IRAK4）、转化生长因子-β活化激酶1（TAK1）、TAK1结合蛋白2（TAB2）的mRNA表达以及TLRs信号途径正性调节因子TLR4相关蛋白（PRAT4B）、信号转换接头蛋白2（STAP2）的mRNA表达均明显增高（P均〈0．01），且严重脓毒症组较脓毒症组升高更显著（P均〈0．01）。脓毒症组TLRs信号途径负性调节因子IL-1受体相关激酶3（IRAK—M）、核转录因子-κB抑制性锌指蛋白（Triad3A）的mRNA表达均明显增高（P均〈0．01），严重脓毒症组表达则明显低于脓毒症组（P均〈0．01）。结论TLRs信号途径转导分子／调节因子异常表达可能是脓毒症时全身炎症反应的原因之一。

Objective To investigate the role of transduce molecules and modulatory factors of signal pathways of Toll-like receptors （TLRs） in aberrant inflammatory response in children with sepsis. Methods Peripheral blood mononuclear cell （PBMC） and serum were obtained from 10 children with sepsis, 13 children with severe sepsis and 17 age-matched healthy children as controls. Real-time polymerase chain reaction （real-time PCR） were used to evaluate the mRNA expression levels of TLR2, TLR4, myeloid differentiation primary response gene 88 （MyD88）, interleukin-1 receptor-associated kinase 4 （IRAK-4）, tumor necrosis factor receptor-associated factor 6 （TRAF6）, TAK1-binding protein 2 （TAB2） transforming growth factor-β-activating kinase 1 （TAK1）, interleukin-1 receptor-associated kinase 3 （IRAK-M）, zinc finger protein inhibiting nuclear factor-κB （Triad3A）, protein associated with Tlr4 （PRAT4B） and signal-transducing adaptor protein-2 （STAP2）, and the levels of mRNA expression and production of pro-inflammatory cytokines [tumor necrosis factor-α （TNF-α）, interleukin-1β （IL-1β） and IL-6] were measured by real-time PCR and enzyme-linked immunosorbnent assay （ELISA）. Results Compared with the healthy control group, the levels of gene expression and protein of pro-inflammatory cytokines （TNF-α, IL-1β and IL-6）, the levels of gene expression of signal pathway molecules of TLRs （TLR2, TLR4, MyD88, TRAF6, IRAK4, TAK1 and TAB2） and the levels of gene expression of positive modulators of TLRs pathways （PRAT4B and STAP2） were significantly increased in sepsis group and severe sepsis group （all P〈0.01）, and the levels in severe sepsis group were even higher than those in sepsis group （all P〈 0. 01）. The mRNA expression levels of negative modulators of TLRs pathways （IRAK-M and Triad3A） were significantly elevated in sepsis group compared with those in healthy control group （all P〈0.01）, but the mRNA expression levels of the negative modulators in severe sepsis group were significantly lowered compared with those in sepsis group （all P 〈 0. 01）. Conclusion Aberrant expression of signal pathway molecules and the modulators in TLRs signaling might play an important role on production and development of abnormal inflammatory response in children with sepsis.