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为探讨GEFH1与树突细胞(dendritic cell,DC)TLR4两条下游信号途径的关系,从野生型和TRIF基因、IFNα/β受体基因敲除小鼠中分离和培养DC,LPS或IL-6刺激后收集细胞制备总cDNA,通过实时定量PCR检测GEFH1的mRNA表达。再用GEFH1的siRNA转染DC,LPS刺激后检测IL-6和IL-12a的mRNA表达。结果,在LPS刺激后,GEFH1的mRNA表达在野生型小鼠的DC中显著增加,在TRIF基因、IFN-α/β受体基因敲除小鼠的DC中则未被上调。此外,GEFH1siRNA处理后,IL-6和IL-12a的mRNA表达均上升显著(P>0.05),而在IL-6刺激的野生型小鼠的DC中,GEFH1的mRNA没有明显改变。以上结果提示在转录水平,TRIF-IFNβ信号通路、而非IL-6可诱导GEFH1基因表达。GEFH1可能对MyD88途径中细胞因子IL-6和IL-12a的表达有负调节作用。
To investigate the relationship between GEFH1 and two downstream signaling pathways of dendritic cell (DC) TLR4, DC, LPS or IL-6 were isolated and cultured from wild-type and TRIF genes and IFNα / β receptor knockout mice After stimulation, cells were harvested to prepare total cDNA, and GEFH1 mRNA expression was detected by real-time quantitative PCR. DCs were transfected with siRNA of GEFH1, and the mRNA expression of IL-6 and IL-12a was detected after LPS stimulation. As a result, mRNA expression of GEFH1 significantly increased in DCs of wild-type mice after LPS stimulation and not in DCs of TRIF gene and IFN-α / β-receptor knockout mice. In addition, mRNA expression of both IL-6 and IL-12a increased significantly (P> 0.05) after GEFH1 siRNA treatment, whereas mRNA of GEFH1 did not change significantly in DCs stimulated by IL-6. The above results suggest that TRIF-IFNβ signaling, but not IL-6, can induce GEFH1 gene expression at the transcriptional level. GEFH1 may negatively regulate the expression of cytokines IL-6 and IL-12a in the MyD88 pathway.