miR-34b-5p在骨髓来源巨噬细胞极化中作用的研究

The role of miR-34b-5p in bone marrow derived macrophage polarization

目的研究miR-34b-5p在骨髓来源巨噬细胞（bone marrow derived macrophages ，BMDM）极化中的作用。方法分离小鼠的骨髓细胞，在体外用巨噬细胞集落刺激因子诱导其分化为BMDM，进行PBS（对照）、M1[脂多糖（lipopolysaccharide，LPS）、干扰素（interferon，IFN）-γ]或M2 [白细胞介素4（interleukin-4，IL-4）]极化处理24 h，实时定量PCR检测BMDM的miR-34b-5p表达；BMDM分别转染miR-34b-5p模拟物、阴性对照模拟物、miR-34b-5p抑制物以及阴性对照抑制物，培养24 h后再进行对照、M1或M2极化处理24 h，实时定量PCR、流式细胞术和酶联免疫吸附试验检测诱导型一氧化氮合酶（inducible nitric oxide synthase，iNOS）、肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）和IL-6的表达变化。结果与对照组相比，M1极化的BMDMmiR-34b-5p表达水平升高（P〈0.05），M2极化的BMDM miR-34b-5p表达水平无明显变化（P〉0.05）；mRNA相对定量结果显示，在对照、M1和M2极化的BMDM中，与对照模拟物组相比，过表达miR-34b-5p组的iNOS、TNF-α和IL-6表达升高（均P〈0.05），与对照抑制物组相比，miR-34b-5p表达抑制组的iNOS、TNF-α和IL-6表达下降（均P〈0.05）；蛋白定量结果显示，与对照模拟物组相比，在PBS对照和M2极化的BMDM中，过表达miR-34b-5p组的iNOS、TNF-α和IL-6表达没有变化（均P〉0.05），PBS在M1极化的BMDM中，过表达miR-34b-5p组的iNOS、TNF-α和IL-6表达升高（均P〈0.05）。结论M1极化的BMDM高表达miR-34b-5p，且miR-34b-5p促进BMDM的M1型极化。

ObjectiveTo investigate the role of miR-34b-5p in the polarization of bone marrow derived macrophages（BMDM）.MethodsBone marrow cells were separated from mice, which were induced by macrophage colony-stimulating factor （M-CSF） to differentiate into BMDM.BMDM were stimulated by control treatments （PBS）, M1 [ lipopolysaccharide （ LPS） and interferon （IFN）-γ] or M2[（interleukin, IL）-4] polarization stimulus, respectively for 24 h. Real-time quantitative PCR was used to detect miR-34b-5p expression of BMDM.BMDM were transfected with miR-34b-5p mimic, negative control mimic （NC mimic）, miR-34b-5p inhibitor, and negative control inhibitor （NC inhibitor）. After 24 h, BMDM were treated with control or polarization stimulus, real-time quantitative PCR, flow cytometry and enzyme linked immunosorbent assay （ELISA） were used to detect the expression of inducible nitric oxide synthase （iNOS）, TNF-α and IL-6.ResultsCompared with control group, the expression of miR-34b-5p was elevated in M1 BMDM（P〈0.05）, the expression of miR-34b-5p was not changed in M2 BMDM（P〉0.05）. Compared to the cells treated with control mimic, over-expressed miR-34b-5p increased the expression of iNOS, TNF-α and IL-6 in M1 and M2 BMDM and control cells（P〈0.05）, while inhibited miR-34b-5p expression reduced the expression of iNOS, TNF-α, and IL-6（P〈0.05）. Compared with the cells treated with control mimic, over-expressed miR-34b-5p did not affect the expression of iNOS, TNF-α, and IL-6 in M1 and M2 BMDM, as well as in control cells（P〉0.05）, while over-expressed miR-34b-5p increased the expression of iNOS, TNF-α, and IL-6 the corresponding cells（P〈0.05）.ConclusionThe expression of miR-34b-5p in M1 BMDM is elevated.miR-34b-5p promotes M1 polarization of BMDM.