Differential stability of cell-free circulating microRNAs: implications for their utilization as biomarkers.
Differential stability of cell-free circulating microRNAs: implications for their utilization as biomarkers.
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DOI：
10.1371/journal.pone.0075184
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Piiper A
中科院分区：
文献类型：
--
作者：
Köberle V;Pleli T;Schmithals C;Augusto Alonso E;Haupenthal J;Bönig H;Peveling-Oberhag J;Biondi RM;Zeuzem S;Kronenberger B;Waidmann O;Piiper A
血液中循环的microRNAs，通过与蛋白质的络合作用和/或另外通过包裹在脂泡中而稳定，目前正被评估为生物标志物。它们与脂类/囊泡的不同结合对它们的稳定性和作为生物标志物的使用的影响在很大程度上是未被探索的，并且是本研究的主题。从血清或血清制备的囊泡和非囊泡部分提取后，用定量逆转录聚合酶链式反应检测一组选定的microRNAs的水平。研究了这些microRNA在RNase A或RNase A家族酶抑制剂RNase A或RNase抑制剂作用下的稳定性。在5h的血清孵育过程中，microRNA-1和microRNA-122的水平显著下降，而microRNA-16、microRNA-21和microRNA-142-3p的水平没有显著下降。核糖核酸酶抑制剂可防止血清中microRNAs的丢失和全血中microRNA-122的降解，microRNA-122是一种不在血细胞中表达的微小RNA。MicroRNA-122的稳定也是通过溶血实现的。血清的长时间孵育导致小泡相关的microRNAs相对非小泡相关的microRNAs的丰富。与囊泡相关的microRNAs对RNase A处理的抵抗力高于与囊泡无关的相应microRNAs。血清microRNAs在长时间孵育时表现出分化的稳定性。核糖核酸酶抑制剂可能有助于有力地保护无细胞循环中的microRNAs的模式。对于不在血细胞中表达的microRNA，这也可以通过溶血来实现。与囊泡相关的microRNAs似乎比那些与囊泡无关的microRNAs更稳定，这可能有助于揭示miRNAs的其他生物标记物属性。
MicroRNAs circulating in the blood, stabilized by complexation with proteins and/or additionally by encapsulation in lipid vesicles, are currently being evaluated as biomarkers. The consequences of their differential association with lipids/vesicles for their stability and use as biomarkers are largely unexplored and are subject of the present study. The levels of a set of selected microRNAs were determined by quantitative reverse-transcription PCR after extraction from sera or vesicle- and non-vesicle fractions prepared from sera. The stability of these microRNAs after incubation with RNase A or RNase inhibitor, an inhibitor of RNase A family enzymes was studied. The levels of microRNA-1 and microRNA-122, but not those of microRNA-16, microRNA-21 and microRNA-142-3p, declined significantly during a 5-h incubation of the sera. RNase inhibitor prevented the loss of microRNAs in serum as well as the degradation of microRNA-122, a microRNA not expressed in blood cells, in whole blood. Stabilization of microRNA-122 was also achieved by hemolysis. Prolonged incubation of the sera led to enrichment of vesicle-associated relative to non-vesicle-associated microRNAs. Vesicle-associated microRNAs were more resistant to RNase A treatment than the respective microRNAs not associated with vesicles. Serum microRNAs showed differential stability upon prolonged incubation. RNase inhibitor might be useful to robustly preserve the pattern of cell-free circulating microRNAs. In the case of microRNAs not expressed in blood cells this can also be achieved by hemolysis. Vesicle-associated microRNAs appeared to be more stable than those not associated with vesicles, which might be useful to disclose additional biomarker properties of miRNAs.
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