LPS耐受单核细胞中p50抑制IKKα结合于IL-1β启动子区

Inhibition of IKKα binding to IL-1β promoter by pSO in LPS tolerant THP-1 cells

目的研究LPS耐受细胞中IL-1β启动子区p50对IKKα的作用，揭示pSO抑制IL-1β mRNA转录的机制。方法运用人单核细胞系THP-1模拟LPS耐受，使用染色体免疫沉淀（ChIP）和real—time PCR技术定量IL-1β启动子区pSO与IKKα的结合情况，并应用基因沉默技术研究p50和／或IKKα沉默后对IL-1β mRNA转录情况的影响。结果耐受细胞IL-1β启动子区p50结合并不减少，而IKKα结合降低；p50沉默后，IKKα的结合增加，同时IL-1β mRNA转录增加；pSO和IKKα双沉默后，IL-1β mRNA转录又降低。结论在耐受的THP-1细胞中，IL-1β mRNA转录的降低至少部分原因是由于pSO抑制了IKKα对IL-1β启动子区的结合而造成的。

Objective To study the effect of p50 on IKKα at IL-1β promoter in LPS tolerant cells and to reveal the mechanism of the inhibition of IL-1β mRNA by p50. Methods THP-1 human promonocyte model of endotoxin tolerance that simulates the sepsis leukocyte phenotype was used. Chromatin immunoprecipitation assay（CHIP） and real-time PCR were applied to quantify the binding of p50 and IKKα to IL-1β promoter. IL-1β mRNA transcription was studied after knocking-down of p50 and/or IKKα. Results With LPS stimulation, p50 binding did not reduce but somewhat increased at IL-1β promoter in tolerant THP-1 cells. Knocking-down of p50 increased the transcription of IL-1β mRNA, which revealed the inhibitory effect of p50 in tolerant cells. In contrast, the accumulation of IKKα to IL-1β promoter decreased with LPS stimulation in tolerant cells; However, IKKα binding increased after p50 gene knock-down. In the meantime, IL-1β mRNA transcription increased; At last, IL-1β mRNA decreased again after double-knocking down of p50 and IKKα. Conclusion p50 is an inhibitory protein at IL-1β promoter in tolerant THP-1 cells. The unresponsiveness of IL-1β mRNA transcription to LPS at least partly results from the inhibition of IKKα binding to IL-1β promoter by p50.