Epigenetics in Inflammatory Bowel Disease. Contribution of DNA methylation to Ulcerative Colitis pathogenesis

Ulcerative colitis (UC) is a chronic, inflammatory disease of the gastrointestinal tract, primarily affecting the colon. Along with Crohn’s disease (CD) they constitute the most common subtypes of inflammatory bowel disease (IBD). Genetic variants can only explain approximately 20 % of all IBD cases, indicating a more complex pathogenesis which is not fully understood. It has been implied that an interplay between environmental factors, intestinal microbiome, nutrition, and genetic variation contributes to UC development. It is also suggested that epigenetic modifications might have a role in IBD disease development. Epigenetic modifications like DNA methylation can regulate gene expression via structural modifications of DNA. The main objective of this work is to investigate if DNA methylation contributes to UC pathogenesis. Newly diagnosed treatment-naïve UC patients with different disease phenotype and healthy controls were included in this study. Next generation sequencing (NGS) technology was applied to obtain transcriptomic and DNA methylation profiles of UC. Our research shows that DNA methylation profiles differ according to disease severity and gender. The transcriptomic profiles for mild to moderate UC revealed genes regulating tissue-specific pathophysiological properties of tight junctions in the mucosa. Additionally, a gender-dependent pathogenesis of UC could be noted. Genes related to the preservation of mucosal integrity and detoxification of microbial-derived metabolites showed an increased expression in females. Genes related to anti-microbial, and cytotoxicity were found in males, indicating a higher risk for developing colorectal cancer (CRC). Correlations of the transcriptomic and DNA methylation profiles revealed a prominent promoter hypermethylation of genes related to homeostasis and defence, and promoter hypomethylation for genes related to immune response in mild to moderate UC. Surprisingly, the DNA methylation profile for severe UC revealed that hypomethylation were prominent in genes related to anti-inflammatory responses, indicating that hypomethylation might mitigate inflammation during severe UC. These findings can be potentially useful for developing epigenetic drugs and allow new treatment strategies for UC patients in the future

Epigenetics in Inflammatory Bowel Disease. Contribution of DNA methylation to Ulcerative Colitis pathogenesis

Ulcerative colitis (UC) is a chronic, inflammatory disease of the gastrointestinal tract, primarily affecting the colon. Along with Crohn’s disease (CD) they constitute the most common subtypes of inflammatory bowel disease (IBD). Genetic variants can only explain approximately 20 % of all IBD cases, indicating a more complex pathogenesis which is not fully understood. It has been implied that an interplay between environmental factors, intestinal microbiome, nutrition, and genetic variation contributes to UC development. It is also suggested that epigenetic modifications might have a role in IBD disease development. Epigenetic modifications like DNA methylation can regulate gene expression via structural modifications of DNA. The main objective of this work is to investigate if DNA methylation contributes to UC pathogenesis. Newly diagnosed treatment-naïve UC patients with different disease phenotype and healthy controls were included in this study. Next generation sequencing (NGS) technology was applied to obtain transcriptomic and DNA methylation profiles of UC. Our research shows that DNA methylation profiles differ according to disease severity and gender. The transcriptomic profiles for mild to moderate UC revealed genes regulating tissue-specific pathophysiological properties of tight junctions in the mucosa. Additionally, a gender-dependent pathogenesis of UC could be noted. Genes related to the preservation of mucosal integrity and detoxification of microbial-derived metabolites showed an increased expression in females. Genes related to anti-microbial, and cytotoxicity were found in males, indicating a higher risk for developing colorectal cancer (CRC). Correlations of the transcriptomic and DNA methylation profiles revealed a prominent promoter hypermethylation of genes related to homeostasis and defence, and promoter hypomethylation for genes related to immune response in mild to moderate UC. Surprisingly, the DNA methylation profile for severe UC revealed that hypomethylation were prominent in genes related to anti-inflammatory responses, indicating that hypomethylation might mitigate inflammation during severe UC. These findings can be potentially useful for developing epigenetic drugs and allow new treatment strategies for UC patients in the future

Transcriptional profiling of ulcerative colitis in remission

Background - This study addresses whether existing transcriptional profiles can improve and support the current definition of UC in remission apart from the today existing endoscopic, histological and laboratory scoring systems. Methods - Mucosal biopsies from treatment-naïve UC patients (n = 14), healthy controls (n = 16), and UC patients in remission (n = 14) underwent RNA-Seq using the Next Seq550 instrument from Illumina. The algorithm package STAR-2.5.2b was used for down-stream analysis. Principal component analysis (PCA), Limma, and p-value adjustment methods were used to obtain a dataset of significantly differentially expressed genes (DEGs). Gene annotations were performed by using the PANTHER classification system (http://pantherdb.org/), and KEGG (www.genome.jp/kegg/). For functional enrichment the clusterProfiler package and REACTOME database (https://reactome.org/) was used. Fractions of specific cell populations in samples were estimated by applying the R/Bioconductor CellMix manual (http://web.cbio.uct.ac.za/~renaud/CRAN/web/CellMix/). TNF-α levels in biopsies were estimated by qPCR and values <7000 copies/µg protein are considered as non-inflamed tissues. Results - Analyses revealed 927 significantly DEGs in remission when compared with UC and normal samples. PCA showed a clear distinction between remission-, normal and UC samples along the first principal component 1 (PC1) with 45.7%, and second principal component (PC2) with 9.3% of the total variance. Cell fractions of monocytes, T cells, neutrophils, B cells/ lymphoid cells and myeloid cells decreased during remission, while the fraction of epithelial cells increased when compared with UC. This is in concordance with the observed inverse regulation of the common up-regulated inflammatory UC genes during remission. A circumvent situation is also observed for down-regulated UC genes with genes involved in TGFβ signalling, transport and drug metabolism. Aside from DEGs involved in innate—and adaptive immune responses, genes like neuropeptide YY (PYY) and neurotrophic receptor tyrosine kinases (NTRK1 and NTRK2) showed increased expression during remission. Conclusions - Apart from reduced major key inflammatory activities seen for UC, we propose that a gut–brain communication network is involved during remission beside the partial restoration of immunological functions and recovery of local homeostasis in the epithelial mucus layer and lamina propria. In addition, a certain role for the composition of bacterial gut flora may be implied. These results can be useful for the development of treatment strategies for remission and might be useful molecular targets for further investigations aiming to predict relapse of UC patients in the future

Transcriptional signatures that define ulcerative colitis in remission

Background - This study addresses whether existing specific transcriptional profiles can improve and support the current status of the definition of ulcerative colitis (UC) remission apart from the existing endoscopic, histologic, and laboratory scoring systems. For that purpose, a well-stratified UC patient population in remission was compared to active UC and control patients and was investigated by applying the next-generation technology RNA-Seq. Methods - Mucosal biopsies from patients in remission (n = 14), patients with active UC (n = 14), and healthy control patientss (n = 16) underwent whole-transcriptome RNA-Seq. Principal component analysis, cell deconvolution methods, gene profile enrichment, and pathway enrichment methods were applied to define a specific transcriptional signature of UC in remission. Results - Analyses revealed specific transcriptional signatures for UC in remission with increased expression of genes involved in O-glycosylation (MUC17, MUC3A, MUC5AC, MUC12, SPON1, B3GNT3), ephrin-mediated repulsion of cells (EFNB2E, EFNA3, EPHA10, EPHA1), GAP junction trafficking (TUBA1C, TUBA4A, TUBB4B, GJB3, CLTB), and decreased expression of several toll-like receptors (TLR1, TLR3, TLR5, ). Conclusions - This study reveals specific transcriptional signatures for remission. Partial restoration and improvement of homeostasis in the epithelial mucus layer and revival of immunological functions were observed. A clear role for bacterial gut flora composition can be implied. The results can be useful for the development of treatment strategies for UC in remission and may be useful targets for further investigations aiming to predict the outcome of UC in the future

DNA hypo-methylation facilitates anti-inflammatory responses in severe ulcerative colitis.

Severe ulcerative colitis (UC) is a potentially life-threatening disease with a potential colorectal cancer (CRC) risk. The aim of this study was to explore the relationship between transcriptomic and genome-wide DNA methylation profiles in a well-stratified, treatment-naïve severe UC patient population in order to define specific epigenetic changes that could be responsible for the grade of disease severity. Mucosal biopsies from treatment-naïve severe UC patients (n = 8), treatment-naïve mild UC (n = 8), and healthy controls (n = 8) underwent both whole transcriptome RNA-Seq and genome-wide DNA bisulfite- sequencing, and principal component analysis (PCA), cell deconvolutions and diverse statistical methods were applied to obtain a dataset of significantly differentially expressed genes (DEGs) with correlation to DNA methylation for severe UC. DNA hypo-methylation correlated with approximately 80% of all DEGs in severe UC when compared to mild UC. Enriched pathways of annotated hypo-methylated genes revealed neutrophil degranulation, and immuno-regulatory interactions of the lymphoid system. Specifically, hypo-methylated anti-inflammatory genes found for severe UC were IL10, SIGLEC5, CD86, CLMP and members of inflammasomes NLRP3 and NLRC4. Hypo-methylation of anti-inflammatory genes during severe UC implies an interplay between the epithelium and lamina propria in order to mitigate inflammation in the gut. The specifically DNA hypo-methylated genes found for severe UC can potentially be useful biomarkers for determining disease severity and in the development of new targeted treatment strategies for severe UC patients

Genome-Wide DNA Methylation in Treatment-Naïve Ulcerative Colitis

Background and Aims: The aim of this study was to investigate the genome-wide DNA methylation status in treatment-naïve ulcerative colitis [UC], and to explore the relationship between DNA methylation patterns and gene expression levels in tissue biopsies from a well-stratified treatment-naïve UC patient group. Methods; Mucosal biopsies from treatment-naïve patients [n = 10], and a healthy control group [n = 11] underwent genome-wide DNA bisulfite sequencing. Principal component analysis [PCA] and diverse statistical methods were applied to obtain a dataset of differentially methylated genes. DNA methylation annotation was investigated using the UCSC Genome Browser. Gene set enrichments were obtained using the Kyoto Encyclopaedia of Genes and Genomes [KEGG] and PANTHER. Results: Of all significantly differentially expressed genes [DEGs], 25% correlated with DNA methylation patterns; 30% of these genes were methylated at CpG sites near their transcription start site [TSS]. Hyper-methylation was observed for genes involved in homeostasis and defence, whereas hypo-methylation was observed for genes playing a role in immune response [i.e. chemokines and interleukins]. Of the differentially DNA methylated genes, 25 were identified as inflammatory bowel disease [IBD] susceptibility genes. Four genes [DEFFA6, REG1B, BTNL3, OLFM4] showed DNA methylation in the absence of known CpG islands. Conclusions: Genome-wide DNA methylation analysis revealed distinctive functional patterns for hyper-and hypo-methylation in treatment-naïve UC. These distinct patterns could be of importance in the development and pathogenesis of UC. Further investigation of DNA methylation patterns may be useful in the development of the targeting of epigenetic processes, and may allow new treatment and target strategies for UC patients

Transcriptomic Landscape of Treatment-Naïve Ulcerative Colitis

Background and Aims: Ulcerative colitis [UC] is a chronic inflammatory disease that effects the gastrointestinal tract and is considered one of the most prominent and common forms of inflammatory bowel disease [IBD]. This study aimed to define and describe the entire transcriptomic landscape in a well-stratified, treatment-naïve UC patient population compared with control patients by using next-generation technology, RNA-Seq. Methods: Mucosal biopsies from treatment-naïve UC patients [ n = 14], and healthy controls [ n = 16] underwent RNA-Seq. Principal component analysis [PCA], cell deconvolution methods, and diverse statistical methods were applied to obtain and characterise a dataset of significantly differentially expressed genes [DEGs]. Results: Analyses revealed 1480 significantly DEGs in treatment-naïve UC when compared with controls. Cell populations of monocytes, T cells, neutrophils, B cells/ lymphoid cells, and myeloid cells were increased during inflammation, whereas the fraction of epithelial cells were reduced in UC, which is reflected by the DEGs; 79 DEGs were identified as IBD susceptibility genes, and 58 DEGs were expressed in a gender-specific manner. MUC5B, REG3A, DEFA5, and IL33 might be considered as colorectal cancer [CRC] risk factors following UC in males. AQP9 together with CLDN2 may have a role regulating tissue-specific physiological properties in tight junctions in UC. An additional functional role for AQP9 in the synthesis and/or the function of mucus can be implied. Conclusions: This study reveals new potential players in UC pathogenesis in general, and provides evidence for a gender-dependent pathogenesis for UC. These results can be useful for the development of personalised treatment strategies for UC in the futur