MicroRNA Profiling in Mucosal Biopsies of Eosinophilic Esophagitis Patients Pre and Post Treatment with Steroids and Relationship with mRNA Targets Shaolei Lu 1 , Vincent A. Mukkada 2 , Shamlal Mangray 1 , Kelly Cleveland 1 , Nick Shillingford 1 , Christoph Schorl 3 , Alexander S. Brodsky 3 , Murray B. Resnick 1 * 1 Department of Pathology and Laboratory Medicine, Rhode Island Hospital, the Alpert School of Medicine, Brown University, Providence, Rhode Island, United States of America, 2 Department of Pediatrics, Hasbro Children’s Hospital, Providence, Rhode Island, United States of America, 3 Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island, United States of America Abstract Background: The characterization of miRNAs and their target mRNAs involved in regulation of the immune process is an area of intense research and relatively little is known governing these processes in allergic inflammation. Here we present novel findings defining the miRNA and mRNA transcriptome in eosinophilic esophagitis (EoE), an increasing recognized allergic disorder. Methods: Esophageal epithelial miRNA and mRNA from five paired biopsies pre- and post-treatment with glucocorticosteroids were profiled using Taqman and Affymetrix arrays. Validation was performed on additional paired biopsies, untreated EoE specimens and normal controls. Differentially regulated miRNAs and mRNAs were generated, within which miRNA-mRNA target pairs with high predicted confidence were identified. Results: Compared to the post-glucocorticoid treated esophageal mucosa, of all the 377 miRNA sequences examined, 32 miRNAs were significantly upregulated and four downregulated in the pre-treated biopsies. MiR-214 was the most upregulated (150 fold) and miR-146b-5b, 146a, 145, 142-3p and 21 were upregulated by at least 10 fold. Out of 12 miRNAs chosen for validation by qRT-PCR, five (miR-214, 146b-5p, 146a, 142-3p and 21) were confirmed and 11 shared the same trend. When the expression of the 12 miRNAs in the EoE mucosa was compared to unrelated normal mucosa, six (miR-214, 146b-5p, 146a, 21, 203, and 489) showed similar significant changes as in the paired samples and 10 of them shared the same trend. In the same five pairs of samples used to profile miRNA, 311 mRNAs were down-regulated and 35 were up- regulated in pre-treated EoE mucosa. Among them, 164 mRNAs were identified as potential targets of differentially regulated miRNAs. Further analysis revealed that immune-related genes, targeted and non-targeted by miRNAs, were among the most important genes involved in the pathogenesis of EoE. Conclusions: Our findings add to the accumulating body of data defining a regulatory role for miRNA in immune and allergic processes. Citation: Lu S, Mukkada VA, Mangray S, Cleveland K, Shillingford N, et al. (2012) MicroRNA Profiling in Mucosal Biopsies of Eosinophilic Esophagitis Patients Pre and Post Treatment with Steroids and Relationship with mRNA Targets. PLoS ONE 7(7): e40676. doi:10.1371/journal.pone.0040676 Editor: Sunil K. Ahuja, South Texas Veterans Health Care System and University Health Science Center San Antonio, United States of America Received March 13, 2012; Accepted June 12, 2012; Published July 16, 2012 Copyright: ß 2012 Lu et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Molecular Pathology Core of the COBRE Center for Cancer Research Development, funded by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number P20GM103421. The previous segment of this project was supported by the National Center for Research Resources (NCRR) under P20RR017695. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: mresnick@lifespan.org Introduction Eosinophilic esophagitis (EoE) is an increasingly recognized antigen-driven disorder of the esophagus occurring in children and adults [1]. Diagnosis of the disorder is based on clinicopathological correlation between the patient’s clinical manifestations and histologic findings on mucosal biopsies. There is increasing evidence to support that EoE can be an aero-and food allergen driven process in which Th2 derived cytokines and chemokines play important roles [2]. Recently published studies have focused on the EoE transcriptome and a number of dysregulated genes influencing key cellular players such as eosinophils, lymphocytes, mast cells, esophageal epithelial cells and subepithelial myofibro- blasts have been characterized [3,4]. MicroRNAs (miRNAs) are small, non-coding 19–25 nucleotide long RNAs that constitute the most abundant class of regulators of gene expression [5]. Their primary mechanism of action is post- transcriptional regulation via RNA interference leading to mRNA strand degradation or translational inhibition. MiRNAs partici- pate in the regulation of a wide range of physiological and pathological processes including development, neoplasia, stress response and inflammation. The interplay between miRNA and PLoS ONE | www.plosone.org 1 July 2012 | Volume 7 | Issue 7 | e40676