Cunha Carolina, Gomes Cátia, Vaz Ana Rita, Brites Dora
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal; Department of Biochemistry and Human Biology, Faculty of Pharmacy, Universidade de Lisboa, Lisbon, Portugal.
Mediators Inflamm. 2016;2016:6986175. doi: 10.1155/2016/6986175. Epub 2016 Dec 21.
Identification of mediators triggering microglia activation and transference of noncoding microRNA (miRNA) into exosomes are critical to dissect the mechanisms underlying neurodegeneration. We used lipopolysaccharide- (LPS-) induced N9 microglia activation to explore new biomarkers/signaling pathways and to identify inflammatory miRNA (inflamma-miR) in cells and their derived exosomes. Upregulation of iNOS and MHC-II (M1-markers) and downregulation of arginase 1, FIZZ1 (M2-markers), and CX3CR1 (M0/M2 polarization) confirmed the switch of N9 LPS-treated cells into the M1 phenotype, as described for macrophages/microglia. Cells showed increased proliferation, activated TLR4/TLR2/NF-B pathway, and enhanced phagocytosis, further corroborated by upregulated MFG-E8. We found NLRP3-inflammasome activation in these cells, probably accounting for the increased extracellular content of the cytokine HMGB1 and of the MMP-9 we have observed. We demonstrate for the first time that the inflamma-miR profiling (upregulated miR-155 and miR-146a plus downregulated miR-124) in M1 polarized N9 cells, noticed by others in activated macrophages/microglia, was replicated in their derived exosomes, likely regulating the inflammatory response of recipient cells and dissemination processes. Data show that LPS-treated N9 cells behave like M1 polarized microglia/macrophages, while providing new targets for drug discovery. In particular, the study yields novel insights into the exosomal circulating miRNA during neuroinflammation important for emerging therapeutic approaches targeting microglia activation.
识别触发小胶质细胞激活的介质以及非编码微小RNA（miRNA）向细胞外囊泡的转移，对于剖析神经退行性变的潜在机制至关重要。我们利用脂多糖（LPS）诱导N9小胶质细胞激活，以探索新的生物标志物/信号通路，并鉴定细胞及其衍生细胞外囊泡中的炎性miRNA（炎症miR）。如巨噬细胞/小胶质细胞那样，诱导型一氧化氮合酶（iNOS）和主要组织相容性复合体II类分子（MHC-II，M1标志物）上调，精氨酸酶1、抵抗素样分子α（FIZZ1，M2标志物）和CX3C趋化因子受体1（CX3CR1，M0/M2极化）下调，证实经LPS处理的N9细胞转变为M1表型。细胞增殖增加，Toll样受体4/2（TLR4/TLR2）/核因子κB（NF-κB）通路激活，吞噬作用增强，乳脂肪球表皮生长因子8（MFG-E8）上调进一步证实了这一点。我们发现这些细胞中NLRP3炎性小体激活，这可能是我们观察到的细胞因子高迁移率族蛋白B1（HMGB1）和基质金属蛋白酶9（MMP-9）细胞外含量增加的原因。我们首次证明，在M1极化的N9细胞中炎性miR谱（miR-155和miR-146a上调以及miR-124下调）在其衍生的细胞外囊泡中也有体现，其他人在活化的巨噬细胞/小胶质细胞中也观察到了这一现象，这可能调节受体细胞的炎症反应和传播过程。数据表明，经LPS处理的N9细胞表现得如同M1极化的小胶质细胞/巨噬细胞，同时为药物研发提供了新靶点。特别是，该研究对神经炎症期间细胞外囊泡循环miRNA有了新的认识，这对于针对小胶质细胞激活的新兴治疗方法很重要。
