MicroRNA-126基因敲减小鼠腹腔巨噬细胞功能的变化及意义  

Functional changes of mouse peritoneal macrophages in microRNA-126 knock-down mice

作　　者：郭萌萌[1] 赵娟娟[1] 陶弋婧 胡燕[1] 秦娜琳[1] 郑静[1] 田丹[1] 周涯[2] 徐林[1] 

机构地区：[1]遵义医学院免疫学教研室,贵州省免疫学研究生教育创新基地,遵义563000 [2]遵义医学院医学物理学教研室,遵义563000

出　　处：《现代免疫学》2015年第4期322-327,共6页Current Immunology

基　　金：国家自然科学基金(31370918;81260398);教育部新世纪优秀人才计划(NCET-12-0661);贵州省国际合作项目(10C315)

摘　　要：研究microRNA-126(miR-126)基因敲减小鼠腹腔巨噬细胞的功能变化,初步探讨其意义。采用real-time PCR检测LPS刺激前后野生型(wild type,WT)小鼠腹腔巨噬细胞miR-126表达变化,CCK8法检测其增殖情况;观察miR-126基因敲减(knock down,KD)小鼠腹腔巨噬细胞的形态变化,并用real-time PCR检测miR-126表达变化;CCK8法检测LPS刺激下miR-126KD小鼠腹腔巨噬细胞增殖变化;FACS检测巨噬细胞表面MHCⅡ类分子和CD86分子的表达变化;最后,real-time PCR检测巨噬细胞表达炎症因子IL-6、TGF-β和Ⅰ型精氨酸酶(arginase 1,Arg-1)等的变化情况。结果显示,WT小鼠腹腔巨噬细胞在LPS刺激后miR-126表达水平下调,显著低于未刺激组(P<0.05),而增殖能力明显增强(P<0.05);与WT小鼠相比,miR-126KD小鼠腹腔巨噬细胞的miR-126表达量明显下调(P<0.05);形态上,WT小鼠腹腔巨噬细胞有较长伪足,多呈梭形,而miR-126KD小鼠腹腔巨噬细胞多呈圆形,细胞边缘光滑较少见伪足;LPS作用48h后,miR-126KD小鼠腹腔巨噬细胞的增殖能力明显强于WT小鼠(P<0.05);且其膜MHCⅡ类分子和CD86分子表达也较WT小鼠显著上调(P<0.05);LPS刺激下,miR-126KD小鼠腹腔巨噬细胞CCL-1表达显著增加,而IL-6、TGF-β和Arg-1的表达水平显著减少(P<0.05)。这些结果提示,miR-126基因敲减可显著影响小鼠腹腔巨噬细胞的增殖能力和功能相关分子的表达,提示miR-126对小鼠腹腔巨噬细胞的功能具有重要的调控作用。This study aimed to investigate the function of peritoneal macrophages in microRNA-126 knock down mice, and to explore the associated consequences. The relative expression of miR-126 in the peritoneal microphages of the wild type （WT） mice pre- and post-stimulation with LPS was examined by real-time PCR and proliferation of peritoneal macrophages was detec- ted by CCK8 assay. For the miR-126 gene knock down （Knock down, KD） mice morphological change of peritoneal macropha- ges was examined, and the relative expression of miR-126 was estimated by real time-PCR; After LPS stimulation, the prolif- eration of peritoneal macrophages in the miR-126KD mice was detected by CCK8 assays changes of surface expression levels of MHC class II and CD86 molecules were analyzed by FACS. Finally, changes in the expression of inflammatory molecules inclu- ding IL-6, TGF-β and I type arginase （Ang-1） in peritoneal macrophages were detected using RT-PCR. The result showed that relative expression of miR-126 in peritoneal macrophages in WT mouse was reduced significantly after LPS stimulation （P〈0. 05）. Conversely, the proliferation of peritoneal macrophages was increased significantly （P〈0.05）; Compared with the WT mice, miR-126 expression levels of peritoneal macrophages in miR-126KD mouse was significantly decreased （P〈0.05）. Morphologically, peritoneal macrophages in WT mice were mostly fusiform in shape, with long pseudopodia, while those in miR- 126KD were round with smooth edges and rare pseudopodia. Forty-eight hours after LPS stimulation, the proliferation of peri- toneal macrophages in miR-126KD mouse was significantly elevated than that in the WT mice （P〈0.05） and the expression levels of MHCⅡ and CD86 molecules were also significantly increased （P〈0.05）. Finally, peritoneal macrophages in miR- 126KD mice expressed higher levels of CCL-I and lower levels of IL-6, TGF-β and Arg-1 （P〈0.05）. Our results thus indicate that miR-126 gene knock down could significantly influence

关 键 词：MIR-126 基因敲减 巨噬细胞 LPS IL-6 

分 类 号：R392.11[医药卫生—免疫学]