Extracellular RNA in circulation mediates intercellular communication in normal and pathological processes. One mode of circulating miRNA transport in bodily fluids is within 30–150 nm small extracellular vesicles (sEVs) or exosomes. Uptake of sEVs can regulate gene expression in recipient cells enabling circulating miRNAs to exert paracrine and systemic effects. Complex regional pain syndrome (CRPS) is a debilitating pain disorder characterized by chronic inflammation. Our previous investigations identified a significant decrease of hsa-miR-939 in whole blood from CRPS patients compared to control; we also observed that overexpression of miR-939 can negatively regulate several proinflammatory genes in vitro. Though downregulated in whole blood, miR-939 was significantly upregulated in sEVs isolated from patient serum. Here we investigated miR-939 packaging into sEVs in vitro under inflammation induced by monocyte chemoattractant protein-1 (MCP-1), a chemokine that is upregulated in CRPS patients. Stimulation of THP-1 monocytes by MCP-1 led to elevated levels of miR-939 in sEVs, which was abrogated using inhibitors of exosome secretion. miRNAs loaded into exosomes largely contain short miRNA sequence motifs called EXOmotifs. Mutation analysis of miR-939 showed that EXOmotif is one of the possible cellular mechanisms responsible for packaging miR-939 into sEVs. We confirmed gene expression changes in recipient cells following the uptake of sEVs enriched in miR-939 using RNA sequencing. Additionally, our data from primary immune cell-derived sEVs of CRPS patients and controls demonstrate that while the relative expression of miR-939 is higher in sEVs derived from B cells, T cells and NK cells relative to monocyte-derived sEVs in controls, only the B cell-derived sEVs showed a significantly higher level of miR-939 in CRPS patients. Differential miRNA sorting into exosomes and its functional impact on recipient cells may contribute to the underlying pathophysiology of CRPS.

循环中的细胞外RNA在正常和病理过程中介导细胞间通讯。在体液中循环miRNA转运的一种模式是在30-150 nm的小细胞外囊泡（sEVs）或外泌体中。sEV的摄取可以调节受体细胞中的基因表达，从而使循环的miRNA发挥旁分泌和全身作用。复杂区域性疼痛综合征（CRPS）是一种以慢性炎症为特征的虚弱性疼痛疾病。我们以前的研究发现，CRPS患者的全血中的hsa-miR-939与对照组相比有显着降低。我们还观察到，miR-939的过度表达可在体外负面调节多个促炎基因。尽管在全血中下调，但miR-939在从患者血清中分离出的sEV中显着上调。在这里，我们研究了miR-939体外包装到sEV中的情况在单核细胞趋化蛋白-1（MCP-1）诱导的炎症中，趋化因子在CRPS患者中被上调。MCP-1对THP-1单核细胞的刺激导致sEV中miR-939的水平升高，这被外泌体分泌抑制剂所消除。装入外泌体的miRNA大部分包含称为EXOmotifs的短miRNA序列基序。miR-939的突变分析表明，EXOmotif是负责将miR-939包装到sEV中的可能细胞机制之一。我们证实了使用RNA测序技术摄取富含miR-939的sEV后受体细胞中的基因表达发生了变化。此外，我们从CRPS患者和对照的原发性免疫细胞衍生的sEV中获得的数据表明，尽管miR-939的相对表达在B细胞衍生的sEV中较高，相对于对照中单核细胞衍生的sEV，T细胞和NK细胞仅在CRPS患者中显示出B细胞衍生的sEV显着更高的miR-939水平。分化成外泌体的差异性miRNA及其对受体细胞的功能影响可能有助于CRPS的潜在病理生理。