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尿液中肾小管损伤微小RNA生物标志物的鉴定:新一代测序与基于定量聚合酶链反应的分析平台的比较

Identification of tubular injury microRNA biomarkers in urine: comparison of next-generation sequencing and qPCR-based profiling platforms.

作者信息

Nassirpour Rounak, Mathur Sachin, Gosink Mark M, Li Yizheng, Shoieb Ahmed M, Wood Joanna, O'Neil Shawn P, Homer Bruce L, Whiteley Laurence O

机构信息

Drug Safety, Pfizer Worldwide Research and Development, 1 Burtt Rd, Andover, MA 01810, USA.

出版信息

BMC Genomics. 2014 Jun 18;15(1):485. doi: 10.1186/1471-2164-15-485.

Abstract

BACKGROUND

MicroRNAs (miRNAs) are small, non-coding RNAs that regulate protein levels post-transcriptionally. miRNAs play important regulatory roles in many cellular processes and have been implicated in several diseases. Recent studies have reported significant levels of miRNAs in a variety of body fluids, raising the possibility that miRNAs could serve as useful biomarkers. Next-generation sequencing (NGS) is increasingly employed in biomedical investigations. Although concordance between this platform and qRT-PCR based assays has been reported in high quality specimens, information is lacking on comparisons in biofluids especially urine. Here we describe the changes in miRNA expression patterns in a rodent model of renal tubular injury (gentamicin). Our aim is to compare RNA sequencing and qPCR based miRNA profiling in urine specimen from control and rats with confirmed tubular injury.

RESULTS

Our preliminary examination of the concordance between miRNA-seq and qRT-PCR in urine specimen suggests minimal agreement between platforms probably due to the differences in sensitivity. Our results suggest that although miRNA-seq has superior specificity, it may not detect low abundant miRNAs in urine samples. Specifically, miRNA-seq did not detect some sequences which were identified by qRT-PCR. On the other hand, the qRT-PCR analysis was not able to detect the miRNA isoforms, which made up the majority of miRNA changes detected by NGS.

CONCLUSIONS

To our knowledge, this is the first time that miRNA profiling platforms including NGS have been compared in urine specimen. miRNAs identified by both platforms, let-7d, miR-203, and miR-320, may potentially serve as promising novel urinary biomarkers for drug induced renal tubular epithelial injury.

摘要

背景

微小RNA(miRNA)是一类小的非编码RNA,可在转录后调节蛋白质水平。miRNA在许多细胞过程中发挥重要的调节作用,并与多种疾病有关。最近的研究报道了多种体液中存在大量miRNA,这增加了miRNA可作为有用生物标志物的可能性。下一代测序(NGS)在生物医学研究中的应用越来越广泛。尽管在高质量样本中已报道了该平台与基于qRT-PCR的检测方法之间的一致性,但在生物体液尤其是尿液中的比较信息仍很缺乏。在此,我们描述了肾小管损伤(庆大霉素)啮齿动物模型中miRNA表达模式的变化。我们的目的是比较对照组和确诊肾小管损伤大鼠尿液样本中基于RNA测序和qPCR的miRNA谱。

结果

我们对尿液样本中miRNA测序和qRT-PCR之间一致性的初步检查表明,由于灵敏度的差异,两个平台之间的一致性很小。我们的结果表明,尽管miRNA测序具有更高的特异性,但它可能无法检测尿液样本中低丰度的miRNA。具体而言,miRNA测序未检测到一些qRT-PCR鉴定出的序列。另一方面,qRT-PCR分析无法检测到miRNA异构体,而这些异构体构成了NGS检测到的miRNA变化的大部分。

结论

据我们所知,这是首次在尿液样本中比较包括NGS在内的miRNA谱分析平台。两个平台都鉴定出的miRNA,即let-7d、miR-203和miR-320,可能有潜力作为药物诱导的肾小管上皮损伤的新型尿液生物标志物。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0e94/4079956/4edb4a3055a8/12864_2013_6184_Fig1_HTML.jpg

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