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目的研究叉头盒O1(FoxO1)对RAW264.7巨噬细胞白细胞相关免疫球蛋白样受体1(LAIR-1)启动子转录活性的调控作用。方法采用100 ng/m L脂多糖(LPS)刺激RAW264.7巨噬细胞6、12、24 h,流式细胞术观察RAW264.7细胞活化过程中LAIR-1、CD11b以及细胞吞噬功能的变化。软件预测FoxO1与LAIR-1启动子之间存在结合位点。构建FoxO1fl/fl/Lys M-Cre小鼠,观察FoxO1敲除后,RAW264.7巨噬细胞LAIR-1的水平变化。构建LAIR-1基因启动子荧光素酶报告基因载体,转染FoxO1观察对其转录活性的影响。结果 LPS刺激后RAW264.7细胞活化,CD11b表达升高,吞噬能力增强,LAIR-1水平下调。成功构建了髓系细胞FoxO1基因敲除小鼠FoxO1fl/fl/Lys M-Cre,该小鼠LAIR-1 mRNA水平显著降低。软件预测发现在LAIR-1基因启动子区域存在潜在的FoxO1转录因子结合位点,荧光素酶报告基因证实FoxO1可激活LAIR-1启动子转录活性。结论 FoxO1可激活巨噬细胞LAIR-1启动子转录活性。
Objective To investigate the regulatory effect of boxhead O1 (FoxO1) on the transcriptional activity of LAIR-1 promoter in RAW264.7 macrophages. Methods RAW264.7 macrophages were stimulated with 100 ng / mL lipopolysaccharide (LPS) for 6, 12, 24 h. The changes of LAIR-1, CD11b and phagocytosis of RAW264.7 cells were observed by flow cytometry. The software predicts the presence of a binding site between FoxO1 and the LAIR-1 promoter. The FoxO1fl / fl / Lys M-Cre mice were constructed to observe the changes of LAIR-1 level in RAW264.7 macrophages after FoxO1 knockdown. The LAIR-1 gene promoter luciferase reporter gene vector was constructed and transfected into FoxO1 to observe its transcriptional activity. Results LPS stimulation activated RAW264.7 cells, CD11b expression increased phagocytic capacity, LAIR-1 levels were down-regulated. The myeloid FoxO1 knockout mouse FoxO1fl / fl / Lys M-Cre was successfully constructed and the level of LAIR-1 mRNA in this mouse was significantly reduced. The software predicts that there is a potential FoxO1 transcription factor binding site in the promoter region of LAIR-1 gene, and luciferase reporter gene confirmed that FoxO1 can activate LAIR-1 promoter transcriptional activity. Conclusion FoxO1 activates the transcriptional activity of LAIR-1 promoter in macrophages.