MicroRNA-146 a在脓毒症髓源抑制细胞中的表达及其意义

摘要

Objective To compare the phenotype of myeloid derived suppressor cells (MDSCs) separated from the bone marrow of mice 3 d and 7 d after cecal ligation and puncture ( CLP) and to elucidate its potential role in the accumulation and immuno-function of MDSCs by determining the expression of microRNA-146a(miR-146a)in order to explore the effect of miR-146 a on immonosuppression of MDSCs in sepsis .Methods A septic model was prepareol by CLP in adult male C57BL/6J mice.MDSCs(expressing cell-surface CD11b and GR-1 antigens )from bone marrow were harvested 3 and 7 days after CLP and were separated with magnetic bead sorting technique .Then,cytokines secretion and arginase-I activity were detected and the T cell proliferation in vitro and the expression of miR-146a of MDSCs (3 d and 7 d after CLP)were observed.Results MDSCs secreted mostly such promoting inflammatory factors as TNF-α, IL-6 3 days after CLP, but 7 days after CLP , they primarily secreted IL-10 and TGF-βwhich were anti-inflammatory factors . MDSCs had potent immunosuppressive properties by increasing T cell suppression in a late anti-inflammatory phase ( CLP3 d vs CLP7 d, P<0.05).In the meantime,miR-146a of the MDSCs in bone marrow was overexpressed in septic mice at 7 days(P<0.05). Moreover,the expression of miR-146a of the MDSCs in bone marrow of septic mice was higher at 7 days than at 3 days after CLP(P<0.05).Conclusion The data indicate that the phenotype of MDSCs evolves through early pro -inflammatory phase into the late anti-inflammatory phase .MDSCs have potent immunosuppressive properties in the late phase of sepsis . miR-146 a might play a crucial role in the regulation of immunosuppressive activity of MDSCs in late sepsis .%目的：比较脓毒症3和7 d的髓源抑制细胞（ MDSC）表型差异，了解其与MDSC增殖活性及免疫功能变化间的关系，通过检测MDSC中微小RNA-146a（microRNA-146a，miR-146a）的表达，探讨miR-146a对脓毒症MDSC免疫抑制活性的影响。方法采用盲肠穿孔结扎技术制备C57BL／6J小鼠脓毒症模型，运用磁珠分选方法收集分离CLP后3和7 d时小鼠骨髓MDSC（CD11b＋Gr1＋细胞），然后进行体外培养并给予脂多糖（LPS）刺激，收集细胞培养上清液，并检测3和7 d时分泌的细胞因子（ TNF-α、TGF-β、IL-6、IL-10）和精氨酸酶活性，以及检测对T细胞的抑制作用与miR-146a的表达。结果 CLP后3 d时MDSC以分泌促炎因子（TNF-α、IL-6）为主，CLP后7 d时以分泌抗炎因子（ IL-10、TGF-β）为主。 CLP后7 d比3 d时表现出更强的T细胞免疫抑制活性。 qRT-PCR检测小鼠骨髓MDSC中miR-146a表达，发现CLP 7 d时比3 d时的miR-146a表达量显著升高（ P＜0．05）。结论在脓毒症的进展过程中，MDSC表型随之发生演变，脓毒症晚期MDSC免疫抑制活性较早期显著增强。 miR-146 a表达上调可能是脓毒症后期MDSC免疫抑制活性增强的重要机制。