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目的诱导小鼠骨髓来源的巨噬细胞极化为M1型巨噬细胞和M2型巨噬细胞,检测干扰素调节因子5(IRF5)在M1型和M2型巨噬细胞中的表达差异,并用IRF5小干扰RNA(IRF5 siRNA)沉默巨噬细胞中IRF5基因表达,观察其极化状态的改变。方法用γ干扰素(IFN-γ)和脂多糖(LPS)诱导小鼠骨髓来源的巨噬细胞向M1型巨噬细胞分化,白细胞介素4(IL-4)诱导其向M2型巨噬细胞分化,实时定量PCR检测IRF5、IL-12、肿瘤坏死因子α(TNF-α)、诱导型一氧化氮合酶(i NOS)、精氨酸酶1(Arg1)和巨噬细胞甘露糖受体(MMR)mRNA在M1型和M2型巨噬细胞中的表达。用IRF5 siRNA转染巨噬细胞,实时定量PCR检测M1型巨噬细胞标志物IRF5、IL-12、TNF-α、i NOS和M2型巨噬细胞标志物Arg1、MMR mRNA的表达,评估其极化状态的改变。结果流式细胞术检测巨噬细胞分化率达81.7%,实时定量PCR检测显示M1型巨噬细胞中IRF5、IL-12、i NOS mRNA的表达量明显高于M2型巨噬细胞,TNF-α亦高于M2型巨噬细胞;M2型巨噬细胞中Arg1、MMR mRNA表达量同M1型巨噬细胞比较显著增高。IRF5 siRNA转染巨噬细胞后,IRF5、IL-12、TNF-α、i NOS mRNA的表达量显著降低,Arg1、MMR mRNA的表达量明显增加。Western blot法检测结果显示IRF5、IL-12、TNF-α、i NOS蛋白的表达量显著降低,Arg1、MMR蛋白的表达量明显增加,极化状态向M2型巨噬细胞偏移。结论 IRF5对巨噬细胞的极化调控有重要作用,可作为鉴别M1型巨噬细胞和M2型巨噬细胞的重要标志物。
Objective To induce murine bone marrow-derived macrophages to polarize into type M1 macrophages and type M2 macrophages and to detect the expression differences of interferon regulatory factor 5 (IRF5) in type M1 and type M2 macrophages. The expression of IRF5 Small interfering RNA (IRF5 siRNA) silenced IRF5 gene expression in macrophages and observed the change of its polarization state. Methods Mice bone marrow-derived macrophages were induced to differentiate into type M1 macrophages with IFN-γ and lipopolysaccharide (LPS). Interleukin-4 (IL-4) The expression of IRF5, IL-12, TNF-α, iNOS, Arg1 and macrophage mannose receptor were detected by real-time quantitative PCR. (MMR) mRNA expression in M1 and M2 macrophages. The macrophages were transfected with IRF5 siRNA and the expression of Arg1 and MMR mRNA, markers of M1 macrophage markers IRF5, IL-12, TNF-α, iNOS and M2, were detected by real-time PCR. Change of state Results The rate of macrophage differentiation was 81.7% by flow cytometry. Real-time quantitative PCR showed that the expression of IRF5, IL-12 and iNOS mRNA in M1 macrophages was significantly higher than that in M2 macrophages and TNF-α Also higher than type M2 macrophages; M2 type macrophages in Arg1, MMR mRNA expression was significantly higher than M1 macrophages. IRF5 siRNA transfection of macrophages, IRF5, IL-12, TNF-α, iNOS mRNA expression was significantly reduced, Arg1, MMR mRNA expression increased significantly. The results of Western blot showed that the expression of IRF5, IL-12, TNF-α and iNOS protein was significantly decreased, the expression of Arg1 and MMR protein was significantly increased, and the polarization state shifted to M2 macrophages. Conclusion IRF5 plays an important role in regulating the polarization of macrophages and can be used as an important marker to identify M1 macrophages and M2 macrophages.