NOD2-and disease-specific gene expression profiles of peripheral blood mononuclear cells from Crohn's disease patients

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 nucleotidebinding 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 realtime 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,CLEC5 A(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;CLEC5 A,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 CLEC5 A and LYZ as CD-and NOD2-associated genes of PBMCs and encourage further studies on their pathomechanistic roles.

Genetic association analysis of CLEC5A and CLEC7A gene single-nucleotide polymorphisms and Crohn’s disease

BACKGROUND Crohn’s disease(CD)is characterized by a multifactorial etiology and a significant impact of genetic traits.While NOD2 mutations represent well established risk factors of CD,the role of other genes is incompletely understood.AIM To challenge the hypothesis that single nucleotide polymorphisms(SNPs)in the genes CLEC5 A and CLEC7 A,two members of the C-type lectin domain family of pattern recognition receptors,may be associated with CD.METHODS SNPs in CLEC5 A,CLEC7 A and the known CD risk gene NOD2 were studied using real time PCR-based SNP assays.Therefore,DNA samples from 175 patients and 157 healthy donors were employed.Genotyping data were correlated with clinical characteristics of the patients and the results of gene expression data analyses.RESULTS In accordance with previous studies,rs2066844 and rs2066847 in NOD2 were found to be significantly associated with CD(allelic P values=0.0368 and 0.0474,respectively).Intriguingly,for genotype AA of rs1285933 in CLEC5 A,a potential association with CD(recessive P=0.0523;odds ratio=1.90)was observed.There were no associations between CD and SNPs rs2078178 and rs16910631 in CLEC7 A.Variants of rs1285933 had no impact on CLEC5 A gene expression.In contrast,genotype-dependent differences of CXCL5 expression in peripheral blood mononuclear cells were observed.There is no statistical interactionbetween the tested SNPs of NOD2 and CLEC5 A,suggesting of a novel pathway contributing to the disease.CONCLUSION Our data encourage enlarged follow-up studies to further address an association of SNP rs1285933 in CLEC5 A with CD.The C-type lectin domain family member also deserves attention regarding a potential role in the pathophysiology of CD.