小鼠巨噬细胞功能极化可塑性的初步探讨

A preliminary study on macrophage plasticity

目的通过鉴定小鼠骨髓来源巨噬细胞(bone marrow-derived macrophages,BMDM)的功能表型,从而研究巨噬细胞功能极化的可塑性及其内在机制。方法采用流式细胞术检测体外诱导分化的小鼠骨髓来源巨噬细胞纯度;采用荧光定量PCR技术检测由RAW264.7极化的M1、M2巨噬细胞中M1、M2特定标记基因的表达来鉴定RAW264.7的功能状态;小鼠BMDM极化为M1、M2巨噬细胞时,荧光定量PCR技术检测M1、M2中特定标记基因TNF-α、IL-6、Arginase、Fizz-1 mRNA水平的表达,组蛋白去乙酰化酶抑制剂Trichostatin A(TSA)、DNA去甲基化酶抑制剂5-氮杂-2-脱氧胞(Aza)刺激M1、M2巨噬细胞后,检测TNF-α、IL-6、Arginase、Fizz-1 mRNA水平表达的变化。结果 RAW264.7细胞经LPS刺激8 h极化为M1巨噬细胞;RAW264.7细胞经IL-4体外刺激24 h极化为M2巨噬细胞。小鼠BMDM在LPS、IL-4刺激下,分别极化为M1、M2巨噬细胞,改变体外刺激条件,M1巨噬细胞可重分化为M2样巨噬细胞,M2巨噬细胞可重分化为M1样巨噬细胞,M1样巨噬细胞和M2巨噬细胞是介于M1、M2中间状态的2种巨噬细胞;药物TSA、Aza的加入使M1、M2中特定标记基因TNF-α、IL-6、Arginase、Fizz-1 mRNA水平表达增加。结论巨噬细胞的功能极化具有可塑性,巨噬细胞的极化可能与染色质状态的改变有关。

We aimed to study plasticity of functional differentiation of macrophage and its intrinsic mechanism by identifying functional genotype(M1,M2) of mouse bone marrow-derived macrophages(BMDM).Purity of mouse BMDM induced differently in vitro was detected by flow cytometry;when RAW264.7 polarized to M1 and M2 macrophages,fluorescent quantitation PCR was used to identify the functional status of RAW264.7 by detecting expressions of marked genes in M1 and M2;fluorescent quantitation PCR was used to detect the mRNA expression levels of TNF-α,IL-6,Arginase and Fizz-1 when mouse BMDM polarized to M1 and M2 macrophages and treated by Trichostatin A(TSA,an histone deacetylase inhibitor) and Aza(DNA demethylase inhibitor).RAW264.7 cells polarized to M1 macrophage after induction by LPS for 8 hours or to M2 macrophage after induction by IL-4 for 24 hours.And mouse BMDM polarized to M1 and M2 macrophages after induction by LPS and IL-4,respectively.M1 macrophage was able to redifferentiated to M2-like macrophage while M2 macrophage could redifferentiated to M1-like macrophage when induced conditions in vitro changed.The mRNA expression levels of TNF-α,IL-6,Arginase,and Fizz-1 in M1 and M2 increased after treatment of TSA and Aza.The functional differentiation of macrophage showed that the plasticity and polarization of macrophage may be associated with changes of chromatin status.