Basic Study
Copyright ©The Author(s) 2018. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Mar 21, 2018; 24(11): 1196-1205
Published online Mar 21, 2018. doi: 10.3748/wjg.v24.i11.1196
NOD2- and disease-specific gene expression profiles of peripheral blood mononuclear cells from Crohn’s disease patients
Holger Schäffler, Maria Rohde, Sarah Rohde, Astrid Huth, Nicole Gittel, Hannes Hollborn, Dirk Koczan, Änne Glass, Georg Lamprecht, Robert Jaster
Holger Schäffler, Maria Rohde, Sarah Rohde, Astrid Huth, Nicole Gittel, Hannes Hollborn, Georg Lamprecht, Robert Jaster, Department of Medicine II, Division of Gastroenterology, Rostock University Medical Center, Rostock 18057, Germany
Dirk Koczan, Institute of Immunology, Rostock University Medical Center, Rostock 18057, Germany
Änne Glass, Institute for Biostatistics and Informatics in Medicine and Ageing Research, Rostock 18057, Germany
Author contributions: Schäffler H, Rohde S and Jaster R designed the study; Huth A, Schäffler H and Lamprecht G took responsibility for patient care and follow-up; Rohde M, Rohde S, Hollborn H, Jaster R and Koczan D (microarray studies) performed the experiments; Gittel N, Huth A and Schäffler H collected the samples and performed the clinical characterization of the patients; Glass Ä performed the biostatistics; all authors analyzed the data; and Schäffler H and Jaster R wrote the manuscript.
Supported by a grant from the Damp-Foundation (2016-04) to Schäffler H and Rohde S.
Institutional review board statement: The study was approved by the ethics board of the Medical Faculty of the University of Rostock (A 2015-0042). Written informed consent was obtained from each participant prior to enrollment.
Conflict-of-interest statement: The authors declare that there is no conflict of interest.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Robert Jaster, MD, Academic Research, Professor, Senior Scientist, Department of Medicine II, Division of Gastroenterology, Rostock University Medical Center, E.-Heydemann-Str. 6, Rostock 18057, Germany. robert.jaster@med.uni-rostock.de
Telephone: +49-381-4947349 Fax: +49-381-4947482
Received: December 21, 2017
Peer-review started: December 21, 2017
First decision: January 18, 2018
Revised: January 29, 2018
Accepted: February 1, 2018
Article in press: February 1, 2018
Published online: March 21, 2018
Processing time: 84 Days and 22.4 Hours
Abstract
AIM

To investigate disease-specific gene expression profiles of peripheral blood mononuclear cells (PBMCs) from Crohn’s disease (CD) patients in clinical remission.

METHODS

Patients with CD in clinical remission or with very low disease activity according to the Crohn’s disease activity index were genotyped regarding nucleotide-binding oligomerization domain 2 (NOD2), and PBMCs from wild-type (WT)-NOD2 patients, patients with homozygous or heterozygous NOD2 mutations and healthy donors were isolated for further analysis. The cells were cultured with vitamin D, peptidoglycan (PGN) and lipopolysaccharide (LPS) for defined periods of time before RNA was isolated and subjected to microarray analysis using Clariom S assays and quantitative real-time PCR. NOD2- and disease-specific gene expression profiles were evaluated with repeated measure ANOVA by a general linear model.

RESULTS

Employing microarray assays, a total of 267 genes were identified that were significantly up- or downregulated in PBMCs of WT-NOD2 patients, compared to healthy donors after challenge with vitamin D and/or a combination of LPS and PGN (P < 0.05; threshold: ≥ 2-fold change). For further analysis by real-time PCR, genes with known impact on inflammation and immunity were selected that fulfilled predefined expression criteria. In a larger cohort of patients and controls, a disease-associated expression pattern, with higher transcript levels in vitamin D-treated PBMCs from patients, was observed for three of these genes, CLEC5A (P < 0.030), lysozyme (LYZ; P < 0.047) and TREM1 (P < 0.023). Six genes were found to be expressed in a NOD2-dependent manner (CD101, P < 0.002; CLEC5A, P < 0.020; CXCL5, P < 0.009; IL-24, P < 0.044; ITGB2, P < 0.041; LYZ, P < 0.042). Interestingly, the highest transcript levels were observed in patients with heterozygous NOD2 mutations.

CONCLUSION

Our data identify CLEC5A and LYZ as CD- and NOD2-associated genes of PBMCs and encourage further studies on their pathomechanistic roles.

Keywords: Peripheral blood mononuclear cells; Gene expression; NOD2; Lysozyme; Crohn’s disease; CLEC5A

Core tip: Peripheral blood mononuclear cells (PBMCs) are a useful tool to study peculiarities of the immune response in the context of Crohn’s disease (CD). Here, we investigated whether PBMCs from patients with CD, even at the stage of clinical remission, exhibit altered gene expression profiles after challenge with pathogen-associated molecular patterns and vitamin D. For TREM1, lysozyme and CLEC5A, disease-associated expression patterns, with higher transcript levels in patient-derived PBMCs, were observed. The two latter genes, along with four other transcripts, also showed NOD2-dependent expression profiles. TREM1 and CLEC5A may act with NOD2 in a regulatory network with a pathophysiological role in CD.