Aberrant MicroRNA Expression in Endometrial Carcinoma Using Formalin-Fixed Paraffin-Embedded (FFPE) Tissues

Taek Sang Lee,Hye Won Jeon,Yong Beom Kim,Young A Kim,Soon Beom Kang,Min A Kim,Wei Yan

doi:10.1371/journal.pone.0081421

Abstract

This study aimed to identify the candidate miRNAs in the carcinogenesis of endometrial carcinoma, and to explore whether FFPE material would be suitable for miRNA profiling. We identified the differences between miRNA expression profiles using human miRNA microarray in endometrioid endometrial adenocarcinomas (EECs) and normal endometria. Of those tested, miR-200a*, miR-200b*, miR-141, miR-182, and miR-205 were greatly enriched. The expressions of these five miRNAs were validated using quantitative real-time reverse transcription-PCR (qRT-PCR). We then performed qRT-PCR miR expression profiling in 30 FFPE specimens (20 EECs, 10 normal endometria) and re-confirmed the results of differential expression between cancer and normal tissue. Following this, we tested whether the specific inhibition of overexpressed miRNAs would alter chemosensitivity. In the in vitro cell viability assay, anti-miR200b* showed a trend toward enhanced cytotoxicity slightly in cisplatin compared to the negative control (p = 0.07). This information provided the candidate miRNAs for further confirmation of the role of miRNAs in the carcinogenesis of EECs, potentially serving as a diagnostic or therapeutic tool. FFPE specimens of endometrial tissues are suitable as a source for miRNA microarray profiling.

Highlights

MicroRNAs are small non-coding RNAs of 19–24 nucleotides that regulate gene expression post-transcriptionally by imperfect base-pairing to messenger RNAs [1]
We present the results of miRNA deregulation in a set of endometrioid endometrial cancer (EEC) and normal endometrial tissues from both FF and formalin-fixed paraffin-embedded (FFPE) samples using real-time quantitative PCR array
In line with the microarray results, the expression levels of the five miRNAs were much higher in endometrial cancer tissues and endometrial cancer cell lines (Hec1A, Ishikawa) than in non-tumor tissues. (Fig. 3) To generalize the above results and examine whether the miRNA sequencing could be applied to FFPE-derived samples, we investigated the expressions of three miRNAs using 20 microdissected FFPE endometrial cancer samples and 10 non-tumor endometrial FFPE tissues

Summary

Introduction

MicroRNAs (miRNAs) are small non-coding RNAs of 19–24 nucleotides that regulate gene expression post-transcriptionally by imperfect base-pairing to messenger RNAs [1]. Hundreds of miRNAs have been identified in various animal genomes. Each miRNA is believed to target as many as two hundred transcripts and approximately 30% of human genes are regulated by miRNAs [2]. The biological functions of most miRNAs are not fully understood. It has been suggested that miRNAs are involved in various biological processes, including cell proliferation, apoptosis, differentiation, and metabolism [3,4]. Since the discovery in 2002 that miRNA dysregulation is linked to cancer, [5] many studies have sought to describe the relationship between miRNAs, cancer progression, and metastasis [6]

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