Sage Journals: Discover world-class research

Abstract

Crohn’s disease affects those individuals with polygenic risk factors. The identified risk loci indicate that the genetic architecture of Crohn’s disease involves both innate and adaptive immunity and the response to the intestinal environment including the microbiome. Genetic risk alone, however, predicts only 25% of disease, indicating that other factors, including the intestinal environment, can shape the epigenome and also confer heritable risk to patients. Patients with Crohn’s disease can have purely inflammatory disease, penetrating disease or fibrostenosis. Analysis of the genetic risk combined with epigenetic marks of Crohn’s disease and other disease associated with organ fibrosis reveals common events are affecting the genes and pathways key to development of fibrosis. This review will focus on what is known about the mechanisms by which genetic and epigenetic risk factors determine development of fibrosis in Crohn’s disease and contrast that with other fibrotic conditions.

Keywords

Inflammatory bowel disease intestinal fibrosis epigenetics genetics Crohn’s disease

Introduction

Disease pathogenesis results from the heritable risk that accrues from alterations in DNA sequence, risk polymorphisms, and from alterations in the epigenome that control gene expression when exposed to environmental change. Epigenetic control of gene expression is exerted through modification of DNA regulatory elements or enhancers that induce transition of condensed heterochromatin, where gene transcription is inhibited by histone modifications and DNA methylation, to euchromatin, where genes are accessible for transcription. Gene expression is also controlled by small non-coding interfering RNAs, microRNAs, which post-transcriptionally regulate gene expression. Crohn’s disease is a polygenic disorder with more than 200 risk loci identified by genome-wide association studies (GWAS). However, understanding the risk of disease development or expression of a specific phenotype of Crohn’s disease in a patient is not predicted or understood solely by genetic risk. Examination of the epigenetic changes associated with development of fibrosis in Crohn’s disease and fibrosis in other organs, including the lungs, heart, liver, and kidneys reveals patterns that are common to all. This review will focus on what is known about the mechanisms by which genetic and epigenetic risk factors determine development of fibrosis in Crohn’s disease and contrast that with other fibrotic conditions.

Genetics

Inflammatory bowel diseases, Crohn’s disease, and ulcerative colitis are polygenic diseases for which ∼200 risk loci have been identified.^1,2 The mostly highly significant genetic associations are with the intracellular bacterial sensor NOD2, defective autophagic responses with ATG16L1 and IRGM, and with the IL-23R, indicating the genetic architecture of Crohn’s disease involves both defective innate and adaptive immune responses to intestinal microbiota.¹ To date, a deeper analysis of GWAS data has not fully revealed a genomic basis that accounts for individual Crohn’s disease phenotypes.^3,4 An approach using multi-locus genetic risk scores has improved the genetic risk assessment of inflammatory bowel disease (IBD) but also indicates that rather than established risk variants, other independent variables modulate disease.^5,6 Ethnic variations in the complement of associated risk loci does not account for ethnic variations in disease location or behavior in Crohn’s disease.^2,7 Purely genetic models of Crohn’s disease are prone to underestimate the interactions among risk loci, termed epistasis.⁸ Epistatic components need to be integrated into these models by estimating the contribution of non-genetic factors, termed missing heritability, which can be accounted for by epigenetics.^9,10

Examination of genetic risk loci by pathway analysis or gene ontogeny identifies groups of polymorphisms likely to play a role in pathogenesis of fibrostenosis. TGF-β is a key cytokine that is central to the development of fibrosis. The TGF-β pathway includes identified risk variants in Smad3 and Smad7, variants in the Janus-activated kinase (Jak)-Tyk2-STAT3 pathway that regulates TGF-β expression in these cells and the negative feedback of this pathway, SOCS3.^11,12 The events lead to development of fibrosis in mesenchymal cells: fibroblasts, myofibroblasts and smooth muscle, the cell types that, once activated in Crohn’s disease, produce autocrine TGF-β1 and are responsible for extracellular matrix production.¹³ The functional outcomes of mutations in these key GWAS risk loci that mechanistically result in TGF-β1-dependent fibrosis are distinct from the outcomes of mutations leading to initial and sustained inflammation in epithelial and immune cells in the intestine. In the case of TGF-β signaling, Smad7 is increased in epithelial and immune cells, inhibiting Treg responses, whereas Smad7 is diminished in subepithelial myofibroblasts and allows sustained TGF-β signaling and extracellular matrix production.^14–16

Other risk loci have been identified that confer risk of fibrostenotic disease that involve other pathways leading to fibrosis in the intestine. The 5T5T polymorphism at the matrix metalloprotein-3 (MMP3) gene increased the risk of developing fibrostenotic complications.¹⁷ The MMPs and tissue metalloproteinases (TIMPs) are key regulators of the balance between extracellular matrix deposition and degradation. Homozygosity for the rs1363670 G-allele near IL-12B is an independent risk factor for development of fibrostenosis, and for a shorter time to critical stricture formation in the ileum.¹⁸ Other risk alleles have been identified in patients with penetrating disease. The Montreal classification is hierarchical, whereby patients may present with penetrating disease that is the result of underlying fibrostenosis. Identifying Montreal Class B2 fibrostenotic Crohn’s disease, however, as distinct from patients with Montreal Class B1 inflammatory and Montreal Class B3 penetrating Crohn’s disease, is difficult but of crucial importance in understanding risk loci and susceptibility of a particular phenotype.¹⁹

Epigenetics

The identified genetic factors and susceptibility loci account for only 13.6% of disease variability and no more than 25% of the genetic risk in Crohn’s disease.^1,2 Epigenetic processes translate environmental events associated with genetic risk into regulation of chromatin, which shapes the expression of genes, and thereby the activity of specific cell types that participate in disease pathophysiology. Epigenetic mechanisms are emerging as key mediators of the effects of both genetics and the environment on gene expression and disease.²⁰ In addition to a set of inherited epigenetic marks, there are likely non-heritable epigenetic marks that are more dynamic and change in response to environmental stimuli.²¹ In Crohn’s disease interaction of the environment, including the intestinal microbiome and metabolome, with the susceptible patient’s genome and immune system shape the epigenome. These non-genetic effects that alter gene expression and function are implied by the results of multi-locus genetic risk analyses and represent the missing heritability in GWAS.^5,6

Epigenetics is defined as a “stably inherited phenotype that results from mechanisms other than changes in DNA sequence”.¹¹ Although initially an individual’s epigenome was not thought to be heritable, there is now increasing evidence that epigenetic inheritance can persist for multiple generations.²² Evidence from a number of lines of investigation demonstrates epigenetic heritability from cell to cell during mitosis, from generation to generation during meiosis, and includes true transgenerational inheritance,²³ which means transmittance of information from one generation to the next that affects the traits of offspring without alteration of the sequence of DNA. Such mechanisms have been shown to include incomplete erasure of DNA methylation, parental effects, transmission of distinct RNA types (e.g. mRNA, non-coding RNA, miRNA), and persistence of subsets of histone marks.²³ Epimutations, that is, epigenetic changes that are sustained in the germ line, can be transmitted in a true intergenerational fashion by surviving the developmental reprogramming that erases epigenomic changes present in the parent. This mechanism has been shown to be operative in animal models of liver fibrosis. Remodeling of DNA methylation and histone acetylation in offspring of mice harboring epigenetic changes altering TGF-β1 expression that results in liver fibrosis is lowered in male F₁ and F₂ generations through a process termed suppressive adaptation.²⁴ Humans with milder non-alcoholic fatty liver disease have hypomethylation of the anti-fibrogenic factor PPAR-γ promoter compared with patients with more severe fibrosis, lending support to this notion. All these aforementioned findings suggest transmission of an epigenetic suppressive adaptation that can help offspring better adapt to future hepatic insults that might result in fibrosis. Suppressive adaptation, however, was not seen in the setting of renal fibrosis.²⁴

Even though all cells within the intestine or an organism share a common genome, gene expression in an individual cell type is regulated by the unique epigenetic events that affect that cell type, and may be distinct from neighboring cell types. This can account for the sometimes contradictory epigenetic mechanisms that are identified as regulating gene expression in different cell types such as epithelial, immune, and mesenchymal cells. Thus understanding the mechanisms regulating gene expression in a cell type critical to a disease process, for example mesenchymal cells and fibrosis, based on an epigenetic analysis of DNA obtained from heterogeneous cell populations can be difficult.

Epigenetic changes that regulate gene expression and function are grouped into four main types: DNA methylation, histone modifications, nucleosome positioning, and small or non-coding interfering RNAs. No information on nucleosome positioning as it relates to fibrosis in Crohn’s disease exists to date, and therefore this will not be discussed further here. The other processes are discussed in greater detail as they relate to the development of fibrosis in general, and to what is known about the development of fibrosis in patients with Crohn’s disease (Table 1).

Table 1.

Genes that can be regulated by epigenetic changes in the development of organ fibrosis.

	Organs involved in fibrosis					Epigenetic mechanism
Gene	Lungs	Skin	Liver	Kidney	Intestine	Methylation	Histone modification	miRNA	References
Smad3	↑	↑	↓	↑	↑	N/A	HDAC1	miR-21, miR-154, miR-29	64, 69, 70, 75–77
Smad7	↓	↓	↓	↓	↓	DNMT1	N/A	miR-21, miR-17-5 p	44, 78, 79
SOCS3	↓	N/A	↓	↓	↓	DNMT1	N/A	miR-19 b	11, 12, 40
MMP	↑	↑	↑	↑	↑	Promoter	HDAC	miR-17, miR-18 a, b & miR-19 a, b,	50, 51, 95
α-SMA	↑	↑	↑	↑	↑	CpG, DNMT1, DNMT3b	H3K4me1	N/A	39, 41
COL	↑	↑	↑	↑	↑	DNMT1, DNMT3b	H4 acetylation, Fli-1 acetylation, H3K4me1	miR-18 a, b & miR-19 a, b, miR-29	38, 41, 54, 66, 67, 71, 72
VMP1	N/A	N/A	N/A	N/A	↑	N/A	N/A	miR-21	42
TGF-β1	↑	↑	↑	↑	↑	Smad7 methylation, Smad4 hypermethylation	H3K4me3↑, H2A.Z↑, ↓H3K9me2 and H3K9me3 on the promoters of ECM genes	miR-21, miR-17 & miR-19 a, b	43, 44
COX2	↓	↑	↑/↓	↑	↓	Promoter hypermethylation or hypomethylation	↓histone H3 and H4 acetylation, ↑ in H3K9me3, H3K27me3, and DNA methylation	Reported mostly in cancer research, miR-101, miR-26 b, miR-146 a, miR-16 and miR-122	52, 38, 80–88
CXCL10	↓	↑	↑	↓	↑	N/A	H3	unknown	53, 91–94
TIMP1	↑/↓	↓	↑	↑	↑	DNMT1	H3K4me1	miR-17, miR-29, miR-1293	54, 95, 96
Fli-1	↓	N/A	N/A	N/A	N/A	Promoter hypermethylation	P300-induced acetylation	N/A	33, 89, 90
Spry-1	↓	↓	↓	↓	↓	Promoter hypermethylation	HDAC↑ Spry-1 gene expression	miR-29, miR-21	66, 100
PTEN	↓	↓	↓	↓	↓	DNMT1-induced hypermethylation	Its interaction with histone H1 to keep chromatin condensation	miR-21	43, 97–99
PPAR-α	↓	↓	↓	↓	↓	Promoter hypermethylation	N/A	miR-21, miR-10 b, miR-33 a	62–64, 101–105
STAT3	↑	↑	↑	↑	↑	DNMT1,	HDACs, SET1, LSD1, EZH2	miR-21, miR-17, miR-29, miR-98	43, 66–70, 106–109
Thy-1	↓	↑	↑	↓/↑	↑	DNMT1, Hypermethylation	H3, HDAC inhibitor	N/A	52, 53, 110–113
IL-27	↓/↑	↑	↓	↓/↑	↓	N/A	N/A	N/A	29, 114–117
NOD2	↓/↑	N/A	↑	N/A	↓	DNA methylation	H3K4Me2 and H4Ac, H3K27Me3 histone modifications	miR-29, miR-192	29, 124–130
TNF-α	↑	↑	↑	↑	↑	DNA demethylation	Histone acetylation, H3K9 and H3K4 methylation	miR-23 a, miR-155, miR-346	29, 118–123
SMT1	↑	N/A	↑	↑	↓	DNA methylation	N/A	N/A	29, 131
IL-19	↑	↑	N/A	↑	↓	N/A	N/A	N/A	29

↑: upregulation; ↓: downregulation; ↓/↑: conflicting evidence; N/A: study not done.

DNA methylation

Methylation of cytosine by replacement of the hydrogen in position 5 (5MeC) in the context of CpG dinucleotides that are clustered in CpG islands is a common DNA modification. Of the 28 CpG dinucleotides present in the human genome, 60–80% are methylated.²⁵ Methylation typically, but not always, represses gene expression by either interfering with the binding of transcription factors to their DNA binding sites or recruiting methyl-CpG-binding proteins that attract histone and chromatin-modifying enzymes. DNA methyltransferases (DNMT)-1 and DNMT-3a and 3b are the primary enzymes responsible for methylation of CpG islands.²⁶ DNMT-1 is a maintenance methyltransferase, whereas DNMT-3a and 3b are de novo methyltransferases. Methylation is reversed by two processes, active and passive demethylation. The ten-eleven translocation methylcytosine dioxygenase (TET) family of enzymes function to catalyze active demethylation via 5MeC hydroxymethylation (5HMeC) which attracts DNA excision and repair machinery, restoring DNA to a demethylated status.²⁷ Passive demethylation occurs when maintenance methylation is absent and progressive dilution of 5MeC occurs during DNA replication.²⁸

DNA methylation and fibrosis

Alterations of DNA methylation have been examined in a number of disease processes that result in tissue fibrosis including systemic sclerosis, pulmonary and cardiac fibrosis, hepatic fibrosis, and intestinal fibrosis in Crohn’s disease.^21,29–35 Hypermethylation of specific genes as well as global changes in DNA methylation have been identified in these organ systems. Two genomic studies in patients with idiopathic pulmonary fibrosis (IPF) demonstrated extensive DNA methylation changes in the control of IPF gene expression.^36–38 Different levels of CpG island methylation are present in specific genes regulating a fibroproliferative phenotype in IPF, and myeloproliferative diseases, via miR-17∼92, involve an increased DNMT-1-mediated feedback loop involving both microRNAs and DNA methylation.^39,40 Notably altered CpG island methylation in the α-smooth muscle actin (α-SMA) promoter was present in pulmonary fibroblasts and myofibroblasts in patients with IPF.⁴¹ A core set of genes known to be related to fibrosis, including several collagens, were differentially methylated in patients with progressive renal fibrosis compared with controls.⁴² Recently, in a rat model of hypoxia-induced cardiac fibrosis, global hypermethylation of gene expression was observed along with upregulation of both DNMT-1 and DMNT-3b that was associated with upregulation of collagen and α-SMA in renal fibroblasts.⁴³

Genome-wide methylation profiling in patients with IBD has identified numerous sites that are differentially methylated between cases and controls.³⁰ The most highly statistically significant include genes controlling altered immune activation, responses to luminal bacteria, and regulation of the Th17 pathway.²⁹ A significant enrichment in DNA methylation was seen within 50 kb of several Crohn’s disease GWAS risk loci including IL-27, IL-19, tumor necrosis factor (TNF), Soluble latent membrane-type 1 (SMT1) and NOD2. In this study by Nimmo and colleagues, methylation status was predictive of disease activity.²⁹ In pediatric Crohn’s disease, Adams et al. provided evidence that four of the most differentially methylated regions resided in proximity to the vacuole membrane protein-1 (VMP1) GWAS locus.⁴⁴ VMP1 is a putative transmembrane protein that has been reported to be involved in different biological events including autophagy, cell adhesion, and membrane translocation.⁴⁵ The microRNA (miR)-21 gene lies within the VMP1 gene. They share a common transcription start site and promoter region, but pri-miR-21 possesses its own unique promoter, thus VMP-1 and pri-miR-21 can be differentially transcribed. Primary miRNA (pri-miRNA) with about 100 nucleotides is transcribed from miRNA genes in the nucleus by RNA polymerase II and further processed into pre-miRNA by a microprocessor complex. Our own recent work has demonstrated that the increased transcription of miR-21 in muscle cells and myofibroblasts of patients with fibrostenotic Crohn’s disease determines the sustained TGF-β1 signaling that results in excess collagen and extracellular matrix production and fibrosis.⁴⁵ This process uniquely characterizes patients with Montreal Class B2 fibrostenotic Crohn’s disease as distinct from patients with Montreal Class B1 inflammatory and Montreal Class B3 penetrating Crohn’s disease.^19,45,46

Histone modifications of DNA and post-translational modification of proteins

Histones are also key players in epigenetics. The four core histones, H2a, H2B, H3, and H4 associate as two H2A–H2B dimers and a H3–H4 tetramer that comprise the nucleosome.⁴⁷ Adjacent nucleosome octamers are separated by ∼50 kb of DNA with the linker histone, H1 interposed between. All histones are also subject to post-translational modifications on their tail regions including phosphorylation, acetylation, methylation, ubiquitination, SUMOlyation, and ADPribosylation. These post-transcriptional modifications of histone contribute to the transcriptional state of the genomic DNA. Generally euchromatin, open or lightly packed chromatin with accessible DNA and actively transcribed genes, and heterochromatin, condensed or tightly packed inaccessible chromatin, are each characterized by different levels of specific histone acetylation and/or methylation and position along the genome in promoter regions or intron/exon regions.^48,49 Histone-modifying enzymes catalyze the post-translational modification of histones and non-histone proteins. This large group of enzymes includes histone acetyltransferases (e.g. p300/CBP) and histone deacetylases (e.g. HDACs), and lysine methyltransferases (e.g. LSD).⁵⁰ The transcription of a gene, therefore, is regulated by the cumulative influence of multiple histone modifications that results from the activity of histone-modifying enzymes. Data from the ENCODE project has identified key histones and their modifications that have become the most highly studied for their ability to control accessibility of chromatin and thereby regulation of gene expression (Table 1).⁵¹

Histone modification and fibrosis

Both histone acetylation and deacetylation are linked to the development of pulmonary fibrosis. It is worth noting that H3 hyperacetylation through decreased expression of histone deactylase is consistently associated with pulmonary fibrosis.^52,53 This regulation of HDAC expression in the lungs results in TGF-β-induced myofibroblast differentiation, and excess collagen and matrix metalloproteinase-1 production.⁵³ Acetylation levels of H3 also regulate the expression of cyclooxygenase-2, IFN-gamma-inducible protein 10 (CXCL10), and Thy-1 cell surface antigen, all of which are integral to the development fibrosis in the lung.^54,55 A similar process in hepatic stellate cells regulates expression of profibrotic genes including α-SMA, collagen I, tissue inhibitor of metalloproteinases1, and TGF-β1 via Histone3 lysine4 methyltransferase I.⁵⁶ In systemic sclerosis, increased p300 acetyl transferase activity induces acetylation of Fli-1 proto-oncogene, thereby relieving the transcriptional repression of collagens IαI and Iα2, the major collagen species in fibrosis.³³

Differential patterns of histone H3 and H4 acetylation have been identified in Crohn’s disease.^57,58 Mokry et al. recently provided evidence that many of the GWAS risk loci overlap with DNA regulatory elements in the intestine, particularly histone H3 lysine 27 (H3K27ac) but also p300, which is responsible for H3K27 acetylation and H3K4me1.⁵⁹ Sadler et al. have demonstrated that collagen Iα2 expression induced by the cytokines interleukin-1β, TNF-α, and TGF-β is regulated by hyperacetylation of histone H4.⁶⁰

Small or non-coding RNA interference

RNA interference of gene expression by microRNA (miR), small ∼18–24 nucleotide non-coding single-stranded RNA molecules, is implicated in the epigenetic regulation of fibrosis.^61,62 In general, miRs post-transcriptionally repress gene expression by targeting mRNA for degradation. miR genes are located throughout the genome. They can be found in introns of coding regions, in introns or exons of non-coding genes, or in intergenic regions. In some cases they are transcribed independently from their own specific promoters, as is the case with primary microRNA-21 (pri-miR-21).⁶³

MicroRNA and fibrosis

A number of miRs have been identified that have a similar role in the regulation of fibrosis in the lung, liver, heart, kidney, or skin in addition to the intestine. While these miRs can have organ and tissue-specific regulation and effects, two are consistently associated with fibrosis and with the expression of TGF-β, miR-21, and miR-29. miR-21 is pro-fibrotic and is implicated in the transcriptional regulation of Sprouty homolog 1 (Spry-1), phosphatase and tensin homolog (PTEN), peroxisome proliferator-activated receptor-α (PPAR-α), signal transducer and activator of transcription-3 (STAT3), and Smad7.^45,64–66 It is worth noting that miR expression can itself be subject to epigenetic regulation. Transcription of miR-21, as noted above for example, is regulated by the methylation level of its promoter.⁶⁷ miR-29 a,b,c are anti-fibrotic and are implicated in the suppression of collagen expression, MMP, and Spry1 expression.^45,68–72 miR-29 expression is down-regulated by the TGF-β-dependent Smad3 transcription factor. The miR17∼92 cluster is also an important determinant of fibrosis. Transcribed from this cluster are miRs that can target key proteins in fibrosis including collagen IαI (miR-18 a,b and miR19a,b), TGF-β (miR-17 and miR-19 a,b), and MMPs (miR-17, miR-18 a,b and miR-19 a,b).^39,73,74

Summary

GWAS analysis of Crohn’s disease has identified numerous risk loci that account for up to 25% of the genetic risk. Recent investigation of the epigenome indicates differential changes in DNA methylation patterns, histone modifications, and differential expression of miRs can further contribute to the ‘heritable’ risk of developing fibrostenotic Crohn’s disease. Integration of genetic susceptibility with changes in the epigenome associated with the development of organ fibrosis may lead to a greater understanding of the heritable risk of Crohn’s disease and open the door to target therapeutically critical processes that prevent or reverse the development of fibrosis.

For progress to be made in Crohn’s disease, efforts to understand the epigenome and the changes that relate to the identified risk loci and their associated pathways, and thus the missing heritability of fibrosis, will be needed. This understanding will only come from exploration of strictly phenotyped and genotyped patients and in individual cell types relevant to fibrosis.

Footnotes

Conflict of interest

None declared.

Funding

This work was supported by DK49691 from NIH: National Institutes for Diabetes, Digestive and Kidney Diseases (JFK).

References

Jostins

Ripke

Weersma

. Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease. Nature 2012; 491: 119–124.

Liu

van Sommeren

Huang

. Association analyses identify 38 susceptibility loci for inflammatory bowel disease and highlight shared genetic risk across populations. Nat Genet 2015; 47: 979–986.

Rivas

Beaudoin

Gardet

. Deep resequencing of GWAS loci identifies independent rare variants associated with inflammatory bowel disease. Nat Genet 2011; 43: 1066–1073.

Cleynen

Mahachie John

Henckaerts

. Molecular reclassification of Crohn's Disease by cluster analysis of genetic variants. PLoS One 2010; 5: e12952–e12952.

Essers

Lee

Kugathasan

. Established genetic risk factors do not distinguish early and later onset Crohn’s Disease. Inflamm Bowel Dis 2009; 15: 1508–1514.

Muise

Xing

. Association between a multi-locus genetic risk score and inflammatory bowel disease. Bioinform Biol Insights 2013; 7: 143–152.

Huang

Haritunians

Okou

. Characterization of genetic loci that affect susceptibility to inflammatory bowel diseases in African Americans. Gastroenterology 2015; 149: 1575–1586.

Zuk

Hechter

Sunyaev

. The mystery of missing heritability: Genetic interactions create phantom heritability. Proc Natl Acad Sci USA 2012; 109: 1193–1198.

Koch

. Epigenetics: An epigenetic twist on the missing heritability of complex traits. Nat Rev Genet 2014; 15: 218–218.

10.

Loddo

Romano

. Inflammatory bowel disease: Genetics, epigenetics and pathogenesis. Front Immunol 2015; 6: 551–551.

11.

Kanehisa

Sato

Kawashima

. KEGG as a reference resource for gene and protein annotation. Nucleic Acids Res 2016; 44: D457–D462.

12.

Iness

Yoon

. Noncanonical STAT3 activation regulates excess TGF-β1 and Collagen I expression in muscle of stricturing Crohn’s Disease. J Immunol 2015; 194: 3422–3431.

13.

Flynn

Murthy

Grider

. Endogenous IGF-I and [alpha]V[beta]3 Integrin ligands regulate increased smooth muscle hyperplasia in stricturing Crohn's Disease. Gastroenterology 2010; 138: 285–293.

14.

Grider

Kuemmerle

. 361 Antagomir to MicroRNA-21 reverses the loss of negative TGF-signaling from inappropriately decreased Smad7 expression in Crohn's Disease, and decreases excess Collagen, CTGF, IGF-I and Fibrosis in TNBS-induced colitis. Gastroenterology 2012; 142: S–85.

15.

Monteleone

Del Vecchio Blanco

Monteleone

. Post-transcriptional regulation of Smad7 in the gut of patients with inflammatory bowel disease. Gastroenterology 2005; 129: 1420–1429.

16.

Monteleone

Kumberova

Croft

. Blocking Smad7 restores TGF-β1 signaling in chronic inflammatory bowel disease. J Clin Invest 2001; 108: 601–609.

17.

Meijer

MJW

Mieremet-Ooms

MAC

van Hogezand

. Role of matrix metalloproteinase, tissue inhibitor of metalloproteinase and tumor necrosis factor-α single nucleotide gene polymorphisms in inflammatory bowel disease. World J Gastroenterol 2007; 13: 2960–2966.

18.

Henckaerts

Van Steen

Verstreken

. Genetic risk profiling and prediction of disease course in Crohn's Disease patients. Clin Gastroenterol Hepatol 2009; 7: 972–980. e2.

19.

Satsangi

Silverberg

Vermeire

. The Montreal classification of inflammatory bowel disease: controversies, consensus, and implications. Gut 2006; 55: 749–753.

20.

Feinberg

. Phenotypic plasticity and the epigenetics of human disease. Nature 2007; 447: 433–440.

21.

Yang

Schwartz

. Epigenetics of idiopathic pulmonary fibrosis. Translat Res 2015; 165: 48–60.

22.

Petronis

. Epigenetics as a unifying principle in the aetiology of complex traits and diseases. Nature 2010; 465: 721–727.

23.

Trerotola

Relli

Simeone

. Epigenetic inheritance and the missing heritability. Hum Genomics 2015; 9: 17–17.

24.

Zeybel

Hardy

Wong

. Multigenerational epigenetic adaptation of the hepatic wound-healing response. Nat Med 2012; 18: 1369–1377.

25.

Rivera

Ren

. Mapping human epigenomes. Cell 2013; 155: 39–55.

26.

Baubec

Colombo

Wirbelauer

. Genomic profiling of DNA methyltransferases reveals a role for DNMT3B in genic methylation. Nature 2015; 520: 243–247.

27.

Tahiliani

Koh

Shen

. Conversion of 5-Methylcytosine to 5-Hydroxymethylcytosine in mammalian DNA by MLL Partner TET1. Science 2009; 324: 930–935.

28.

Hackett

Surani

. Beyond DNA: Programming and inheritance of parental methylomes. Cell 2013; 153: 737–739.

29.

Nimmo

ERP

Prendergast

JGP

Aldhous

MCP

. Genome-wide methylation profiling in Crohn's disease identifies altered epigenetic regulation of key host defense mechanisms including the Th17 pathway. Inflamm Bowel Dis 2012; 18: 889–899.

30.

McDermott

Ryan

Tosetto

. DNA methylation profiling in inflammatory bowel disease provides new insights into disease pathogenesis. J Crohns Colitis 2016; 10: 77–86.

31.

Mann

. Epigenetics in liver disease. Hepatology 2014; 60: 1418–1425.

32.

Yang

Pedersen

Rabinovich

. Relationship of DNA methylation and gene expression in idiopathic pulmonary fibrosis. Am J Respir Crit Care Med 2014; 190: 1263–1272.

33.

Luo

Wang

Shu

. Epigenetic mechanisms: An emerging role in pathogenesis and its therapeutic potential in systemic sclerosis. Int J Biochem Cell Biol 2015; 67: 92–100.

34.

Neary

Watson

Baugh

. Epigenetics and the overhealing wound: The role of DNA methylation in fibrosis. Fibrogenesis Tissue Repair 2015; 8: 1–13.

35.

Tao

Yang

J-J

Shi

K-H

. DNA methylation in cardiac fibrosis: New advances and perspectives. Toxicology 2014; 323: 125–129.

36.

Rabinovich

Kapetanaki

Steinfeld

. Global methylation patterns in idiopathic pulmonary fibrosis. PLoS One 2012; 7: e33770–e33770.

37.

Sanders

Ambalavanan

Halloran

. Altered DNA methylation profile in idiopathic pulmonary fibrosis. Am J Respir Crit Care Med 2012; 186: 525–535.

38.

Evans

Barnes

Garner

. Epigenetic regulation of cyclooxygenase-2 by methylation of c8orf4 in pulmonary fibrosis. Clin Sci 2016; 130: 575–586.

39.

Dakhlallah

Batte

Wang

. Epigenetic regulation of miR-17∼92 contributes to the pathogenesis of pulmonary fibrosis. Am J Respir Crit Care Med 2013; 187: 397–405.

40.

Fourouclas

Gilby

. Methylation of the suppressor of cytokine signaling 3 gene (SOCS3) in myeloproliferative disorders. Haematologica 2008; 93: 1635–1644.

41.

Gharaee-Kermani

. Epigenetic regulation of myofibroblast differentiation by DNA methylation. Am J Pathol 2010; 177: 21–28.

42.

Mohtat

Suzuki

. Cytosine methylation changes in enhancer regions of core pro-fibrotic genes characterize kidney fibrosis development. Genome Biol 2013; 14: R108–R108.

43.

Watson

Horgan

Neary

. Epigenetic therapy for the treatment of hypertension-induced cardiac hypertrophy and fibrosis. J Cardiovasc Pharmacol Ther 2016; 21: 127–137.

44.

Adams

Kennedy

Hansen

. Two-stage genome-wide methylation profiling in childhood-onset Crohn's Disease implicates epigenetic alterations at the VMP1/MIR21 and HLA loci. Inflamm Bowel Dis 2014; 20: 1784–1793.

45.

Calvo-Garrido

Carilla-Latorre

Escalante

. Vacuole membrane protein 1, autophagy and much more. Autophagy 2008; 4: 835–837.

46.

Kuemmerle

. Increased pro-fibrotic miR-21 and decreased anti-fibrotic miR-29b regulate TGF-β1 signaling, TGF-β1-dependent collagen-I expression and fibrosis in fibrostenotic (B2) Crohn’s disease. Inflamm Bowel Dis 2014; 20: 2–2.

47.

Mariño-Ramírez

Kann

Shoemaker

. Histone structure and nucleosome stability. Exp Rev Proteomics 2005; 2: 719–729.

48.

Carey

Workman

. The role of chromatin during transcription. Cell 2007; 128: 707–719.

49.

Karlic

Chung

Lasserre

. Histone modification levels are predictive for gene expression. Proc Natl Acad Sci USA 2010; 107: 2926–2931.

50.

Marmorstein

Trievel

. Histone modifying enzymes: Structures, mechanisms, and specificities. Biochim Biophys Acta 2009; 1789: 58–68.

51.

ENCODE Project Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature 2012; 489: 57–74.

52.

Tzouvelekis

Kaminski

. Epigenetics in idiopathic pulmonary fibrosis. Biochem Cell Biol 2015; 93: 159–170.

53.

Wang

Chen

Finger

. Suberoylanilide hydroxamic acid: A potential epigenetic therapeutic agent for lung fibrosis? Eur Respir J 2009; 34: 145–155.

54.

Coward

Feghali-Bostwick

Jenkins

. A central role for G9a and EZH2 in the epigenetic silencing of cyclooxygenase-2 in idiopathic pulmonary fibrosis. FASEB J 2014; 28: 3183–3196.

55.

Coward

Watts

Feghali-Bostwick

. Repression of IP-10 by interactions between histone deacetylation and hypermethylation in idiopathic pulmonary fibrosis. Mol Cell Biol 2010; 30: 2874–2886.

56.

Perugorria

Wilson

Zeybel

. Histone methyltransferase ASH1 orchestrates fibrogenic gene transcription during myofibroblast transdifferentiation. Hepatology 2012; 56: 1129–1139.

57.

Tsaprouni

Ito

Powell

. Differential patterns of histone acetylation in inflammatory bowel diseases. J Inflamm 2011; 8: 1–1.

58.

Ventham

Kennedy

Nimmo

. Beyond gene discovery in inflammatory bowel disease: The emerging role of epigenetics. Gastroenterology 2013; 145: 293–308.

59.

Mokry

Middendorp

Wiegerinck

. Many inflammatory bowel disease risk loci include regions that regulate gene expression in immune cells and the intestinal epithelium. Gastroenterology 2014; 146: 1040–1047.

60.

Sadler

Scarpa

Rieder

. Cytokine-induced chromatin modifications of the type I collagen alpha 2 gene during intestinal endothelial-to-mesenchymal transition. Inflamm Bowel Dis 2013; 19: 1354–1364.

61.

Sato

Tsuchiya

Meltzer

. MicroRNAs and epigenetics. FEBS J 2011; 278: 1598–1609.

62.

Iorio

Piovan

Croce

. Interplay between microRNAs and the epigenetic machinery: An intricate network. Biochim Biophys Acta 2010; 1799: 694–701.

63.

Saini

Griffiths-Jones

Enright

. Genomic analysis of human microRNA transcripts. Proc Natl Acad Sci USA 2007; 104: 17719–17724.

64.

Krichevsky

Gabriely

. miR-21: A small multi-faceted RNA. J Cell Mol Med 2009; 13: 39–53.

65.

Liu

Friggeri

Yang

. miR-21 mediates fibrogenic activation of pulmonary fibroblasts and lung fibrosis. J Exp Med 2010; 207: 1589–1597.

66.

Zhong

Chung

ACK

Chen

H-Y

. Smad3-mediated upregulation of miR-21 promotes renal fibrosis. J Am Soc Nephrol 2011; 22: 1668–1681.

67.

Ribas

Castanares

. A novel source for miR-21 expression through the alternative polyadenylation of VMP1 gene transcripts. Nucleic Acids Res 2012; 40: 6821–6833.

68.

Maurer

Stanczyk

Jüngel

. MicroRNA-29, a key regulator of collagen expression in systemic sclerosis. Arthritis Rheum 2010; 62: 1733–1743.

69.

Nijhuis

Biancheri

Lewis

. In Crohn's disease fibrosis-reduced expression of the miR-29 family enhances collagen expression in intestinal fibroblasts. Clin Sci 2014; 127: 341–350.

70.

Noetel

Kwiecinski

Elfimova

. microRNA are central players in anti- and profibrotic gene regulation during liver fibrosis. Front Physiol 2012; 3: 49–49.

71.

Qin

Chung

ACK

Huang

. TGF-β/Smad3 signaling promotes renal fibrosis by inhibiting miR-29. J Am Soc Nephrol 2011; 22: 1462–1474.

72.

Zhang

Huang

X-R

Wei

L-H

. miR-29b as a therapeutic agent for Angiotensin II-induced cardiac fibrosis by targeting TGF-[beta]/Smad3 signaling. Mol Ther 2014; 22: 974–985.

73.

Mogilyansky

Rigoutsos

. The miR-17/92 cluster: A comprehensive update on its genomics, genetics, functions and increasingly important and numerous roles in health and disease. Cell Death Differ 2013; 20: 1603–1614.

74.

Agarwal

Bell

Nam

J-W

. Predicting effective microRNA target sites in mammalian mRNAs. eLife 2015; 4.

75.

Milosevic

Pandit

Magister

. Profibrotic role of miR-154 in pulmonary fibrosis. Am J Respir Cell Mol Biol 2012; 47: 879–887.

76.

Lakos

Takagawa

Chen

. Targeted disruption of TGF-beta/Smad3 signaling modulates skin fibrosis in a mouse model of scleroderma. Am J Pathol 2004; 165: 203–217.

77.

Tang

Ding

Guo

. Epigenetic regulation of Smad2 and Smad3 by profilin-2 promotes lung cancer growth and metastasis. Nat Commun 2015; 6: 8230–8230.

78.

Bian

Huang

Wang

. Repression of Smad7 mediated by DNMT1 determines hepatic stellate cell activation and liver fibrosis in rats. Toxicol Lett 2014; 224: 175–185.

79.

Zhou

Zang

Ponnusamy

. Enhancer of Zeste Homolog 2 inhibition attenuates renal fibrosis by maintaining Smad7 and Phosphatase and Tensin homolog expression. J Am Soc Nephrol 2015. Dec 23 [Epub ahead of print].

80.

Liu

Mih

Shea

. Feedback amplification of fibrosis through matrix stiffening and COX-2 suppression. J Cell Biol 2010; 190: 693–706.

81.

Coward

Feghali-Bostwick

Jenkins

. A central role for G9a and EZH2 in the epigenetic silencing of cyclooxygenase-2 in idiopathic pulmonary fibrosis. FASEB J 2014; 28: 3183–3196.

82.

Davids

Carothers

Damas

. Chronic cyclooxygenase-2 inhibition promotes myofibroblast-associated intestinal fibrosis. Cancer Prev Res (Phila) 2010; 3: 348–358.

83.

Chu

. Elucidation of the role of COX-2 in liver fibrogenesis using transgenic mice. Biochem Biophys Res Commun 2008; 372: 571–577.

84.

Feitoza

Gonçalves

Semedo

. Inhibition of COX 1 and 2 prior to renal ischemia/reperfusion injury decreases the development of fibrosis. Mol Med 2008; 14: 724–730.

85.

Abdou

Maraee

Saif

. Immunohistochemical evaluation of COX-1 and COX-2 expression in keloid and hypertrophic scar. Am J Dermatopathol 2014; 36: 311–317.

86.

Cox

Crusius

Peeters

. Haplotype of prostaglandin synthase 2/cyclooxygenase 2 is involved in the susceptibility to inflammatory bowel disease. World J Gastroenterol 2005; 11: 6003–6008.

87.

Honma

Shinohara

Takahashi

. Effect of cyclooxygenase (COX)-2 inhibition on mouse renal interstitial fibrosis. Eur J Pharmacol 2014; 740: 578–583.

88.

Meike

Ochs

Suess

. MicroRNA involved in inflammation: Control of eicosanoid pathway. Front Pharmacol 2011; 2: 39–39.

89.

Wang

Fan

Kahaleh

. Association between enhanced type I collagen expression and epigenetic repression of the FLI1 gene in scleroderma fibroblasts. Arthritis Rheum 2006; 54: 2271–2279.

90.

Kubo

Czuwara-Ladykowska

Moussa

. Persistent down-regulation of Fli1, a suppressor of collagen transcription, in fibrotic scleroderma skin. Am J Pathol 2003; 163: 571–581.

91.

Hintermann

Bayer

Pfeilschifter

. CXCL10 promotes liver fibrosis by prevention of NK cell mediated hepatic stellate cell inactivation. J Autoimmun 2010; 35: 424–435.

92.

Jiang

Liang

Campanella

. Inhibition of pulmonary fibrosis in mice by CXCL10 requires glycosaminoglycan binding and syndecan-4. J Clin Invest 2010; 120: 2049–2057.

93.

Pociask

Chen

Choi

. γδ T cells attenuate bleomycin-induced fibrosis through the production of CXCL10. Am J Pathol 2011; 178: 1167–1176.

94.

Sahin

Wasmuth

. Chemokines in tissue fibrosis. Biochim Biophys Acta 2013; 1832: 1041–1048.

95.

Zhang

Chen

Hong

. TIMP-1 promotes age-related renal fibrosis through upregulating ICAM-1 in human TIMP-1 transgenic mice. J Gerontol A Biol Sci Med Sci 2006; 61: 1130–1143.

96.

Giannandrea

Parks

. MMPs & TIMPs: Diverse functions of matrix metalloproteinases during fibrosis. Dis Model Mech 2014; 7: 193–203.

97.

Xie

Zheng

. Effects of the tumor suppressor PTEN on the pathogenesis of idiopathic pulmonary fibrosis in Chinese patients. Mol Med Rep 2016; 13: 2715–2723.

98.

Zheng

Xie

. Regulatory effects and mechanism of adenovirus-mediated PTEN gene on hepatic stellate cells. Dig Dis Sci 2016; 61: 1107–1120.

99.

Chen

Zhu

Yang

. PTEN Interacts with Histone H1 and controls chromatin condensation. Cell Rep 2014; 8: 2003–2014.

100.

Song

Van Buren

Yosypiv

. Histone deacetylases are critical regulators of the renin-angiotensin system during ureteric bud branching morphogenesis. Pediatr Res 2010; 67: 573–578.

101.

Elbjeirami

. PPAR-α targeting in kidney fibrosis: Is BAY PP1 just another renoprotector? Kidney Int 2011; 80: 1115–1117.

102.

Lakatos

Thatcher

Kottmann

. The role of PPARs in lung fibrosis. PPAR Res 2007; 2007: 71323–71323.

103.

Nan

Kong

Ren

. Activation of peroxisome proliferator activated receptor alpha ameliorates ethanol mediated liver fibrosis in mice. Lipids Health Dis 2013; 12: 11–11.

104.

Zeybel

Hardy

Robinson

. Differential DNA methylation of genes involved in fibrosis progression in non-alcoholic fatty liver disease and alcoholic liver disease. Clin Epigen 2015; 7: 25–25.

105.

Zheng

Sheng

. Effect of miRNA-10b in regulating cellular steatosis level by targeting PPAR-α expression, a novel mechanism for the pathogenesis of NAFLD. J Gastroenterol Hepatol 2010; 25: 156–163.

106.

Yang

Huang

Dasgupta

. Reversible methylation of promoter-bound STAT3 by histone-modifying enzymes. Proc Natl Acad Sci USA 2010; 107: 21499–21504.

107.

Kim

Woo

. Phosphorylation of EZH2 activates STAT3 signaling via STAT3 methylation and promotes tumorigenicity of glioblastoma stem-like cells. Cancer Cell 2013; 23: 839–852.

108.

Gao

Zhou

. Arsenic trioxide prevents rat pulmonary fibrosis via miR-98 overexpression. Life Sci 2014; 114: 20–28.

109.

Lim

Phan

Lim

. Stat3 contributes to keloid pathogenesis via promoting collagen production, cell proliferation and migration. Oncogene 2006; 25: 5416–5425.

110.

Dudas

Mansuroglu

Batusic

. Thy-1 is expressed in myofibroblasts but not found in hepatic stellate cells following liver injury. Histochem Cell Biol 2009; 131: 115–127.

111.

McIntosh

JC1

Hagood

Richardson

. Thy1 (+) and (–) lung fibrosis subpopulations in LEW and F344 rats. Eur Respir J 1994; 7: 2131–2138.

112.

Yuasa

Juniantito

Ichikawa

. Thy-1 expression, a possible marker of early myofibroblast development, in renal tubulointerstitial fibrosis induced in rats by cisplatin. Exp Toxicol Pathol 2013; 65: 651–659.

113.

Rieder

Kessler

West

. Inflammation-induced endothelial-to-mesenchymal transition: A novel mechanism of intestinal fibrosis. Am J Pathol 2011; 179: 2660–2673.

114.

Kim

Akira

. TLR2-mediated production of IL-27 and chemokines by respiratory epithelial cells promotes bleomycin-induced pulmonary fibrosis in mice. J Immunol 2011; 187: 4007–4017.

115.

Dong

Zhao

Tai

. IL-27 attenuates the TGF-β1-induced proliferation, differentiation and collagen synthesis in lung fibroblasts. Life Sci 2016; 146: 24–33.

116.

Dibra

Xia

Mitra

. Mutant p53 in concert with an interleukin-27 receptor alpha deficiency causes spontaneous liver inflammation, fibrosis, and steatosis in mice. Hepatology 2016; 63: 1000–1012.

117.

Summers

SA1

Phoon

Ooi

. The IL-27 receptor has biphasic effects in crescentic glomerulonephritis mediated through Th1 responses. Am J Pathol 2011; 178: 580–590.

118.

Herro

Antunes Rda

Aguilera

. The tumor necrosis factor superfamily molecule LIGHT promotes keratinocyte activity and skin fibrosis. J Invest Dermatol 2015; 135: 2109–2118.

119.

Murdaca

Spanò

Contatore

. Potential use of TNF-α inhibitors in systemic sclerosis. Immunotherapy 2014; 6: 283–289.

120.

Sullivan

Reddy

Dietzmann

. Epigenetic regulation of tumor necrosis factor alpha. Mol Cell Biol 2007; 27: 5147–5160.

121.

Bala

Marcos

Kodys

. Up-regulation of MicroRNA-155 in macrophages contributes to increased Tumor Necrosis Factor α (TNFα) production via increased mrna half-life in alcoholic liver disease. J Biol Chem 2011; 286: 1436–1444.

122.

Dong

Cui

Jiang

. MicroRNA-23a modulates tumor necrosis factor-alpha-induced osteoblasts apoptosis by directly targeting Fas. J Cell Biochem 2013; 114: 2738–2745.

123.

Frenzel

Semaan

Alsaleh

. A new mode of TNF-[alpha] inhibition by microRNA. J Immunol 2009; 182. 99.26.

124.

Wang

Hartmann

Haimerl

. Nod2 deficiency protects mice from cholestatic liver disease by increasing renal excretion of bile acids. J Hepatol 2014; 60: 1259–1267.

125.

Brain

Owens

Pichulik

. The intracellular sensor NOD2 induces microRNA-29 expression in human dendritic cells to limit IL-23 release. Immunity 2013; 39: 521–536.

126.

Chuang

Zhai

. NOD2 expression is regulated by microRNAs in colonic epithelial HCT116 cells. Inflamm Bowel Dis 2014; 20: 126–135.

127.

Zheng

Abraham

. NF-κB1 inhibits NOD2-induced cytokine secretion through ATF3-dependent mechanisms. Mol Cell Biol 2013; 33: 4857–4871.

128.

Maisi

Prikk

Sepper

. Soluble membrane-type 1 matrix metalloproteinase (MT1-MMP) and gelatinase A (MMP-2) in induced sputum and bronchoalveolar lavage fluid of human bronchial asthma and bronchiectasis. APMIS 2002; 110: 771–782.

129.

Birukawa

Murase

Sato

. Activated hepatic stellate cells are dependent on self-collagen, cleaved by Membrane Type 1 Matrix Metalloproteinase for their growth. J Biol Chem 2014; 289: 20209–20221.

130.

Niarakis

Giannopoulou

Ravazoula

. Detection of a latent soluble form of membrane type 1 matrix metalloprotease bound with tissue inhibitor of matrix metalloproteinases-2 in periprosthetic tissues and fluids from loose arthroplasty endoprostheses. FEBS J 2013; 280: 6541–6555.

131.

Kazes

Delarue

Hagège

. Soluble latent membrane-type 1 matrix metalloprotease secreted by human mesangial cells is activated by urokinase. Kidney Int 1998; 54: 1976–1984.

Genetic and epigenetic regulation of intestinal fibrosis

Abstract

Keywords

Introduction

Genetics

Epigenetics

DNA methylation

DNA methylation and fibrosis

Histone modifications of DNA and post-translational modification of proteins

Histone modification and fibrosis

Small or non-coding RNA interference

MicroRNA and fibrosis

Summary

Footnotes

Conflict of interest

Funding

References