ABSTRACT

Major efforts are made to characterize the presence of microRNA (miRNA) and messenger RNA in blood plasma to discover novel disease-associated biomarkers. MiRNAs in plasma are associated to several types of macromolecular structures, including extracellular vesicles (EV), lipoprotein particles (LPP) and ribonucleoprotein particles (RNP). RNAs in these complexes are recovered at variable efficiency by commonly used EV- and RNA isolation methods, which causes biases and inconsistencies in miRNA quantitation. Besides miRNAs, various other non-coding RNA species are contained in EV and present within the pool of plasma extracellular RNA. Members of the Y-RNA family have been detected in EV from various cell types and are among the most abundant non-coding RNA types in plasma. We previously showed that shuttling of full-length Y-RNA into EV released by immune cells is modulated by microbial stimulation. This indicated that Y-RNAs could contribute to the functional properties of EV in immune cell communication and that EV-associated Y-RNAs could have biomarker potential in immune-related diseases. Here, we investigated which macromolecular structures in plasma contain full length Y-RNA and whether the levels of three Y-RNA subtypes in plasma (Y1, Y3 and Y4) change during systemic inflammation. Our data indicate that the majority of full length Y-RNA in plasma is stably associated to EV. Moreover, we discovered that EV from different blood-related cell types contain cell-type-specific Y-RNA subtype ratios. Using a human model for systemic inflammation, we show that the neutrophil-specific Y4/Y3 ratios and PBMC-specific Y3/Y1 ratios were significantly altered after induction of inflammation. The plasma Y-RNA ratios strongly correlated with the number and type of immune cells during systemic inflammation. Cell-type-specific “Y-RNA signatures” in plasma EV can be determined without prior enrichment for EV, and may be further explored as simple and fast test for diagnosis of inflammatory responses or other immune-related diseases.

摘要

做出重大努力来表征血浆中microRNA（miRNA）和信使RNA的存在，以发现与疾病相关的新型生物标记。血浆中的MiRNA与多种类型的大分子结构相关，包括细胞外囊泡（EV），脂蛋白颗粒（LPP）和核糖核蛋白颗粒（RNP）。这些复合物中的RNA通过常用的EV-和RNA分离方法以可变的效率回收，这会导致miRNA定量的偏差和不一致。除miRNA外，EV中还包含其他各种非编码RNA种类，并存在于血浆细胞外RNA池中。Y-RNA家族的成员已经在EV中从各种细胞类型中检测到，并且属于血浆中最丰富的非编码RNA类型。我们先前显示，全长Y-RNA穿梭到免疫细胞释放的EV中是受微生物刺激调节的。这表明Y-RNA可能有助于EV在免疫细胞通讯中的功能特性，并且与EV相关的Y-RNA在免疫相关疾病中可能具有生物标志物的潜力。在这里，我们调查了血浆中哪些大分子结构包含全长Y-RNA，以及系统性炎症期间血浆中三种Y-RNA亚型（Y1，Y3和Y4）的水平是否发生了变化。我们的数据表明血浆中大多数全长Y-RNA与EV稳定相关。此外，我们发现来自不同血液相关细胞类型的EV包含特定于细胞类型的Y-RNA亚型比率。使用人体模型进行全身性炎症，我们显示炎症诱导后中性粒细胞特异性Y4 / Y3比值和PBMC特异性Y3 / Y1比值显着改变。在全身性炎症期间，血浆Y-RNA比率与免疫细胞的数量和类型密切相关。血浆EV中特定于细胞类型的“ Y-RNA信号”可以在无需事先富集EV的情况下进行测定，并且可以作为诊断炎症反应或其他免疫相关疾病的简单快速检测方法进行进一步探索。