Department of Human Genetics, Saarland University, Homburg, Germany.
Department of Internal Medicine, Nephrology and Hypertension, Saarland University Medical Center, Homburg, Germany.
Sci Rep. 2017 Jul 12;7(1):5162. doi: 10.1038/s41598-017-05070-0.
Although microRNAs are supposed to be stable in-vivo, degradation processes potentially blur our knowledge on the small oligonucleotides. We set to quantify the effect of degradation on microRNAs in mouse to identify causes for distorted microRNAs patterns. In liver, we found 298, 99 and 8 microRNAs whose expression significantly correlated to RNA integrity, storage time at room temperature and storage time at 4 °C, respectively. Expression levels of 226 microRNAs significantly differed between liver samples with high RNA integrity compared to liver samples with low RNA integrity by more than two-fold. Especially the 157 microRNAs with increased expression in tissue samples with low RNA integrity were most recently added to miRBase. Testing potentially confounding sources, e.g. in-vitro degraded RNA depleted of small RNAs, we detected signals for 350 microRNAs, suggesting cross-hybridization of fragmented RNAs. Therefore, we conclude that especially microRNAs added in the latest miRBase versions might be artefacts due to RNA degradation. The results facilitate differentiation between degradation-resilient microRNAs, degradation-sensitive microRNAs, and likely erroneously annotated microRNAs. The latter were largely identified by NGS but not experimentally validated and can severely bias microRNA biomarker research and impact the value of microRNAs as diagnostic, prognostic or therapeutic tools.
尽管 microRNAs 在体内应该是稳定的，但降解过程可能会模糊我们对这些小寡核苷酸的认识。我们着手量化降解对小鼠中 microRNAs 的影响，以确定导致 microRNAs 模式扭曲的原因。在肝脏中，我们发现了 298、99 和 8 个 microRNAs，它们的表达分别与 RNA 完整性、室温下储存时间和 4°C 下储存时间显著相关。与 RNA 完整性低的肝脏样本相比，具有高 RNA 完整性的肝脏样本中 226 个 microRNAs 的表达水平差异显著，超过两倍。特别是在 RNA 完整性低的组织样本中表达增加的 157 个 microRNAs 是最近添加到 miRBase 中的。测试潜在的混杂来源，例如体外降解的 RNA 耗尽了小 RNA，我们检测到 350 个 microRNAs 的信号，表明碎片 RNA 的交叉杂交。因此，我们得出结论，特别是最新版本 miRBase 中添加的 microRNAs 可能是由于 RNA 降解而产生的假象。这些结果有助于区分抗降解 microRNAs、敏感 microRNAs 和可能错误注释的 microRNAs。后者主要通过 NGS 鉴定，但未通过实验验证，可能严重偏倚 microRNA 生物标志物研究，并影响 microRNAs 作为诊断、预后或治疗工具的价值。
