Hoentjen Frank, Sartor R Balfour, Ozaki Michitaka, Jobin Christian
Center for Gastrointestinal Biology and Diseases, University of North Carolina at Chapel Hill, USA.
Blood. 2005 Jan 15;105(2):689-96. doi: 10.1182/blood-2004-04-1309. Epub 2004 Jul 13.
Interleukin-10-deficient (IL-10(-/-)) mice develop an IL-12-mediated intestinal inflammation in the absence of endogenous IL-10. The molecular mechanisms of the dysregulated IL-12 responses in IL-10(-/-) mice are poorly understood. In this study, we investigated the role of nuclear factor-kappa B (NF-kappaB) and signal transducers and activators of transcription 3 (STAT3) in lipopolysaccharide (LPS)-induced IL-12p40 gene expression in bone marrow derived-dendritic cells (BMDCs) isolated from wild-type (WT) and IL-10(-/-) mice. We report higher IL-12p40 mRNA accumulation and protein secretion in LPS-stimulated BMDCs isolated from IL-10(-/-) compared with WT mice. LPS-induced NF-kappaB signaling is similar in IL-10(-/-) and WT BMDCs as measured by IkappaBalpha phosphorylation and degradation, RelA phosphorylation and nuclear translocation, and NF-kappaB transcriptional activity, with no down-regulatory effects of exogenous IL-10. Chromatin immunoprecipitation demonstrated enhanced NF-kappaB (cRel, RelA) binding to the IL-12p40 promoter in IL-10(-/-) but not WT BMDCs. Interestingly, LPS induced STAT3 phosphorylation in WT but not IL-10(-/-) BMDCs, a process blocked by IL-10 receptor blocking antibody. Adenoviral gene delivery of a constitutively active STAT3 but not control green fluorescence protein (GFP) virus blocked LPS-induced IL-12p40 gene expression and cRel recruitment to the IL-12p40 promoter. In conclusion, dysregulated LPS-induced IL-12p40 gene expression in IL-10(-/-) mice is due to enhanced NF-kappaB recruitment to the IL-12p40 promoter in the absence of activated STAT3.
白细胞介素-10缺陷（IL-10(-/-)）小鼠在缺乏内源性IL-10的情况下会发生IL-12介导的肠道炎症。目前对IL-10(-/-)小鼠中IL-12反应失调的分子机制了解甚少。在本研究中，我们调查了核因子-κB（NF-κB）和信号转导子与转录激活子3（STAT3）在从野生型（WT）和IL-10(-/-)小鼠分离的骨髓来源树突状细胞（BMDCs）中脂多糖（LPS）诱导的IL-12p40基因表达中的作用。我们报告，与WT小鼠相比，从IL-10(-/-)小鼠分离的LPS刺激的BMDCs中IL-12p40 mRNA积累和蛋白质分泌更高。通过IkappaBalpha磷酸化和降解、RelA磷酸化和核转位以及NF-κB转录活性测量，LPS诱导的NF-κB信号在IL-10(-/-)和WT BMDCs中相似，外源性IL-10没有下调作用。染色质免疫沉淀显示，在IL-10(-/-)而非WT BMDCs中，NF-κB（cRel、RelA）与IL-12p40启动子的结合增强。有趣的是，LPS诱导WT而非IL-10(-/-) BMDCs中的STAT3磷酸化，这一过程被IL-10受体阻断抗体阻断。组成型活性STAT3而非对照绿色荧光蛋白（GFP）病毒的腺病毒基因递送阻断了LPS诱导的IL-12p40基因表达和cRel募集到IL-12p40启动子。总之，IL-10(-/-)小鼠中LPS诱导的IL-12p40基因表达失调是由于在缺乏活化STAT3的情况下，NF-κB募集到IL-12p40启动子增强所致。
