ABSTRACT
Whether or not populations diverge with respect to the genetic contribution to risk of specific complex diseases is relevant to understanding the evolution of susceptibility and origins of health disparities. Here, we describe a large-scale whole-genome sequencing study of inflammatory bowel disease encompassing 1,774 affected individuals and 1,644 healthy control Americans with African ancestry (African Americans). Although no new loci for inflammatory bowel disease are discovered at genome-wide significance levels, we identify numerous instances of differential effect sizes in combination with divergent allele frequencies. For example, the major effect at PTGER4 fine maps to a single credible interval of 22 SNPs corresponding to one of four independent associations at the locus in European ancestry individuals but with an elevated odds ratio for Crohn disease in African Americans. A rare variant aggregate analysis implicates Ca2+-binding neuro-immunomodulator CALB2 in ulcerative colitis. Highly significant overall overlap of common variant risk for inflammatory bowel disease susceptibility between individuals with African and European ancestries was observed, with 41 of 241 previously known lead variants replicated and overall correlations in effect sizes of 0.68 for combined inflammatory bowel disease. Nevertheless, subtle differences influence the performance of polygenic risk scores, and we show that ancestry-appropriate weights significantly improve polygenic prediction in the highest percentiles of risk. The median amount of variance explained per locus remains the same in African and European cohorts, providing evidence for compensation of effect sizes as allele frequencies diverge, as expected under a highly polygenic model of disease.
Subject(s)
Calbindin 2/genetics , Genetic Predisposition to Disease , Inflammatory Bowel Diseases/genetics , Receptors, Prostaglandin E, EP4 Subtype/genetics , Black or African American/genetics , Aged , Aged, 80 and over , Colitis, Ulcerative/genetics , Colitis, Ulcerative/pathology , Crohn Disease/genetics , Crohn Disease/pathology , Female , Gene Frequency , Genome-Wide Association Study , Humans , Inflammatory Bowel Diseases/pathology , Male , Multifactorial Inheritance/genetics , Polymorphism, Single Nucleotide/genetics , White People/genetics , Whole Genome SequencingABSTRACT
BACKGROUND & AIMS: Crohn's disease is a relapsing and remitting inflammatory disorder with a variable clinical course. Although most patients present with an inflammatory phenotype (B1), approximately 20% of patients rapidly progress to complicated disease, which includes stricturing (B2), within 5 years. We analyzed DNA methylation patterns in blood samples of pediatric patients with Crohn's disease at diagnosis and later time points to identify changes that associate with and might contribute to disease development and progression. METHODS: We obtained blood samples from 164 pediatric patients (1-17 years old) with Crohn's disease (B1 or B2) who participated in a North American study and were followed for 5 years. Participants without intestinal inflammation or symptoms served as controls (n = 74). DNA methylation patterns were analyzed in samples collected at time of diagnosis and 1-3 years later at approximately 850,000 sites. We used genetic association and the concept of Mendelian randomization to identify changes in DNA methylation patterns that might contribute to the development of or result from Crohn's disease. RESULTS: We identified 1189 5'-cytosine-phosphate-guanosine-3' (CpG) sites that were differentially methylated between patients with Crohn's disease (at diagnosis) and controls. Methylation changes at these sites correlated with plasma levels of C-reactive protein. A comparison of methylation profiles of DNA collected at diagnosis of Crohn's disease vs during the follow-up period showed that, during treatment, alterations identified in methylation profiles at the time of diagnosis of Crohn's disease more closely resembled patterns observed in controls, irrespective of disease progression to B2. We identified methylation changes at 3 CpG sites that might contribute to the development of Crohn's disease. Most CpG methylation changes associated with Crohn's disease disappeared with treatment of inflammation and might be a result of Crohn's disease. CONCLUSIONS: Methylation patterns observed in blood samples from patients with Crohn's disease accompany acute inflammation; with treatment, these change to resemble methylation patterns observed in patients without intestinal inflammation. These findings indicate that Crohn's disease-associated patterns of DNA methylation observed in blood samples are a result of the inflammatory features of the disease and are less likely to contribute to disease development or progression.
Subject(s)
Crohn Disease/genetics , DNA Methylation/genetics , Gene Expression Regulation/genetics , Genome-Wide Association Study , Mendelian Randomization Analysis/methods , Adolescent , Age Factors , Case-Control Studies , Child , Child, Preschool , Crohn Disease/blood , Disease Progression , Female , Follow-Up Studies , Genotype , Humans , Infant , Inflammation/genetics , Male , North America , Risk Assessment , Severity of Illness Index , Sex FactorsABSTRACT
Microbial dysbiosis characterized by alterations in the structure and function of the gut microbiome has long been implicated in the pathogenesis of inflammatory bowel disease (IBD). To date, most human IBD microbiome studies are focused on microbial composition rather than function, however, with the latest technical advancements complemented by the rapidly dropping costs, studies focusing on the functional aspects of microbial dysbiosis are on the rise. Several compelling and complimentary pieces of evidence support the notion that the gut microbiome and their metabolites play an important role in the development of IBD. Data from preclinical studies overwhelmingly support the notion that changes in the gut microbiome causally underlie IBD pathogenesis. Hence, there is considerable interest in modulating the state and function of the gut microbiome to achieve therapeutic benefits. While the causal potential of the gut microbiome remains an active area of current research in the clinical setting, accumulating correlative evidence support the view that microbial dysbiosis parallels increased incidence of IBD. In this review, we intend to provide a brief overview of the current human IBD microbiome findings, describe the cause-effect relationships between the gut microbiome and IBD, and discuss the possibility of using microbiome-based approaches in the diagnosis, therapy, and management of disease. In addition, the potential role of microbiome-based interventions in the treatment of human IBD is also discussed.
Subject(s)
Gastrointestinal Microbiome , Host Microbial Interactions , Inflammatory Bowel Diseases/microbiology , Inflammatory Bowel Diseases/physiopathology , Microbiota , HumansABSTRACT
OBJECTIVES: Environmental factors play an important role in the pathogenesis of Crohn's Disease (CD). In particular, by virtue of the instability of the microbiome and development of immunologic tolerance, early life factors may exert the strongest influence on disease risk and phenotype. METHODS: We used data from 1119 CD subjects recruited from RISK inception cohort to examine the impact of early life environment on disease progression. Our primary exposures of interest were breastfeeding in infancy and exposure to maternal, active, or passive smoke. Our primary outcomes were development of complicated (stricturing or penetrating) disease, and need for CD-related hospitalization, and surgery. Multivariable logistic regression models were used to define independent associations, adjusting for relevant covariates. RESULTS: Our study cohort included 1119 patients with CD among whom 15% had stricturing (B2) or penetrating disease (B3) by 3 years. 331 patients (35%) and 95 patients (10.6%) required CD-related hospitalizations and surgery respectively. 74.5% were breastfed in infancy and 31% were exposed to smoking among whom 7% were exposed to maternal smoke. On multivariable analysis, a history of breastfeeding was inversely associated with complicated (B2/B3 disease) 0.65, CI 95% 0.44-96; P = 0.03) in pediatric CD. Maternal smoking during pregnancy was associated with increased risk of hospitalization during the 3-year follow-up period (OR 1.75, CI 95% 1.05-2.89; P = 0.03). CONCLUSIONS: Early life environmental factors influence the eventual phenotypes and disease course in CD.
Subject(s)
Breast Feeding/statistics & numerical data , Crohn Disease/diagnosis , Environmental Exposure/adverse effects , Prenatal Exposure Delayed Effects/epidemiology , Smoking/adverse effects , Tobacco Smoke Pollution/adverse effects , Adolescent , Child , Colon/pathology , Constriction, Pathologic/epidemiology , Constriction, Pathologic/etiology , Crohn Disease/complications , Crohn Disease/etiology , Crohn Disease/therapy , Disease Progression , Environmental Exposure/statistics & numerical data , Female , Follow-Up Studies , Hospitalization/statistics & numerical data , Humans , Infant, Newborn , Longitudinal Studies , Male , North America/epidemiology , Phenotype , Pregnancy , Prospective Studies , Risk Factors , Severity of Illness Index , Smoking/epidemiology , Time Factors , Tobacco Smoke Pollution/statistics & numerical dataABSTRACT
BACKGROUND: Asthma is a complex disorder influenced by genetics and the environment. Recent findings have linked abnormal DNA methylation in T cells with asthma; however, the potential dysregulation of methylation in airway epithelial cells is unknown. Studies of mouse models of asthma have observed greater levels of 5-hydroxymethylcytosine (5-hmC) and ten-eleven translocation 1 (TET1) expression in lungs. TET proteins are known to catalyze methylation through modification of 5-methylcytosine to 5-hmC. OBJECTIVE: We sought to examine the association of TET1 methylation with asthma and traffic-related air pollution (TRAP). METHODS: TET1 methylation levels from DNA derived from nasal airway epithelial cells collected from 12 African American children with physician-diagnosed asthma and their nonasthmatic siblings were measured by using Illumina 450K arrays. Regions of interest were verified by means of locus-specific pyrosequencing in 35 sibling pairs and replicated in an independent population (n = 186). Exposure to TRAP in participants' early life and at current home addresses was estimated by using a land-use regression model. Methylation studies in saliva, PBMCs, and human bronchial epithelial cells were done to support our findings. RESULTS: Loss of methylation at a single CpG site in the TET1 promoter (cg23602092) and increased global 5-hmC levels were significantly associated with asthma. In contrast, TRAP exposure at participants' current homes significantly increased methylation at the same site. Patterns were consistent across tissue sample types. 5-Aza-2'-deoxycytidine and diesel exhaust particle exposure in human bronchial epithelial cells was associated with altered TET1 methylation and expression and global 5-hmC levels. CONCLUSIONS: Our findings suggest a possible role of TET1 methylation in asthmatic patients and response to TRAP.
Subject(s)
Air Pollution/adverse effects , Asthma/etiology , DNA Methylation , DNA-Binding Proteins/genetics , Proto-Oncogene Proteins/genetics , Vehicle Emissions , 5-Methylcytosine/analogs & derivatives , Adolescent , Alleles , Asthma/metabolism , Case-Control Studies , Child , CpG Islands , Cytosine/analogs & derivatives , Cytosine/metabolism , Epigenesis, Genetic , Epithelial Cells , Female , Gene Expression , Humans , Male , Mixed Function Oxygenases , Nasal Mucosa/metabolism , Promoter Regions, Genetic , RNA, Messenger/genetics , RNA, Messenger/metabolism , Risk FactorsABSTRACT
BACKGROUND: Induced pluripotent stem cells (iPSCs) hold tremendous potential, both as a biological tool to uncover the pathophysiology of disease by creating relevant human cell models and as a source of cells for cell-based therapeutic applications. Studying the reprogramming process will also provide significant insight into tissue development. OBJECTIVE: We sought to characterize the derivation of iPSC lines from nasal epithelial cells (NECs) isolated from nasal mucosa samples of children, a highly relevant and easily accessible tissue for pediatric populations. METHODS: We performed detailed comparative analysis on the transcriptomes and methylomes of NECs, iPSCs derived from NECs (NEC-iPSCs), and embryonic stem cells (ESCs). RESULTS: NEC-iPSCs express pluripotent cell markers, can differentiate into all 3 germ layers in vivo and in vitro, and have a transcriptome and methylome remarkably similar to those of ESCs. However, residual DNA methylation marks exist, which are differentially methylated between NEC-iPSCs and ESCs. A subset of these methylation markers related to epithelium development and asthma and specific to NEC-iPSCs persisted after several passages in vitro, suggesting the retention of an epigenetic memory of their tissue of origin. Our analysis also identified novel candidate genes with dynamic gene expression and DNA methylation changes during reprogramming, which are indicative of possible roles in airway epithelium development. CONCLUSION: NECs are an excellent tissue source to generate iPSCs in pediatric asthmatic patients, and detailed characterization of the resulting iPSC lines would help us better understand the reprogramming process and retention of epigenetic memory.
Subject(s)
Asthma/genetics , Embryonic Stem Cells/metabolism , Epithelial Cells , Induced Pluripotent Stem Cells , Nasal Mucosa/cytology , Adolescent , Animals , Cell Line , Cells, Cultured , DNA Methylation , Epigenomics , Epithelial Cells/cytology , Epithelial Cells/metabolism , Fibroblasts , Foreskin/cytology , Gene Expression Profiling , Humans , Induced Pluripotent Stem Cells/cytology , Induced Pluripotent Stem Cells/metabolism , Male , MiceABSTRACT
BACKGROUND: There is considerable heterogeneity in asthma treatment response. OBJECTIVE: We sought to identify biomarkers of corticosteroid treatment response in children with asthma and evaluate the utility and mechanistic basis of these biomarkers. METHODS: Children (5-18 years) presenting to the emergency department with an acute asthma exacerbation were recruited and followed during hospitalization. Nasal epithelial cells were collected on presentation to the emergency department (T0) and 18 to 24 hours later (T1), and T1/T0 gene expression ratios were analyzed to identify genes associated with good and poor corticosteroid treatment response phenotypes. The utility of these genes in discriminating between systemic corticosteroid treatment response groups was then tested prospectively in a new cohort of patients. A gene candidate (vanin-1 [VNN1]) that consistently distinguished good versus poor response phenotypes was further studied in an experimental asthma model, and VNN1 promoter methylation was measured by means of bisulfite pyrosequencing in patients. RESULTS: VNN1 mRNA expression changes were associated with systemic corticosteroid treatment response in children with acute asthma, and VNN1 was required for optimal response to corticosteroid treatment in an experimental asthma model. A CpG site within the VNN1 promoter was differentially methylated between good versus poor treatment response groups, and methylation at this site correlated with VNN1 mRNA expression. CONCLUSIONS: We have identified a biological basis for poor corticosteroid treatment response that can be used to distinguish a subgroup of asthmatic children who respond poorly to systemic corticosteroid treatment. VNN1 contributes to corticosteroid responsiveness, and changes in VNN1 nasal epithelial mRNA expression and VNN1 promoter methylation might be clinically useful biomarkers of treatment response in asthmatic children.
Subject(s)
Adrenal Cortex Hormones/therapeutic use , Amidohydrolases/metabolism , Asthma/drug therapy , Biomarkers, Pharmacological/metabolism , Nasal Mucosa/physiology , Adolescent , Amidohydrolases/genetics , Animals , Asthma/diagnosis , Cells, Cultured , Child , Child, Preschool , Cohort Studies , DNA Methylation , Disease Progression , Emergency Medical Services , Female , GPI-Linked Proteins/genetics , GPI-Linked Proteins/metabolism , Gene Expression Regulation , Hospitalization , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Knockout , Models, Animal , Prospective Studies , Treatment OutcomeABSTRACT
The renin angiotensin system (RAS) plays a vital role in the regulation of the cardiovascular and renal functions. COS7 is a robust and easily transfectable cell line derived from the kidney of the African green monkey, Cercopithecus aethiops. The aims of this study were to 1) demonstrate the presence of an endogenous and functional RAS in COS7, and 2) investigate the role of a disintegrin and metalloproteinase-17 (ADAM17) in the ectodomain shedding of angiotensin converting enzyme-2 (ACE2). Reverse transcription coupled to gene-specific polymerase chain reaction demonstrated expression of ACE, ACE2, angiotensin II type 1 receptor (AT1R), and renin at the transcript levels in total RNA cell extracts. Western blot and immunohistochemistry identified ACE (60 kDa), ACE2 (75 kDa), AT1R (43 kDa), renin (41 kDa), and ADAM17 (130 kDa) in COS7. At the functional level, a sensitive and selective mass spectrometric approach detected endogenous renin, ACE, and ACE2 activities. ANG-(1-7) formation (m/z 899) from the natural substrate ANG II (m/z 1,046) was detected in lysates and media. COS7 cells stably expressing shRNA constructs directed against endogenous ADAM17 showed reduced ACE2 shedding into the media. This is the first study demonstrating endogenous expression of the RAS and ADAM17 in the widely used COS7 cell line and its utility to study ectodomain shedding of ACE2 mediated by ADAM17 in vitro. The transfectable nature of this cell line makes it an attractive cell model for studying the molecular, functional, and pharmacological properties of the renal RAS.
Subject(s)
ADAM Proteins/metabolism , Peptidyl-Dipeptidase A/metabolism , Renin-Angiotensin System , ADAM Proteins/genetics , ADAM17 Protein , Angiotensin I/metabolism , Angiotensin II/genetics , Angiotensin II/metabolism , Angiotensin-Converting Enzyme 2 , Animals , Base Sequence , COS Cells , Chlorocebus aethiops , Molecular Sequence Data , Peptide Fragments/metabolism , Peptidyl-Dipeptidase A/genetics , RNA Interference , RNA, Messenger/metabolism , Receptor, Angiotensin, Type 1/genetics , Receptor, Angiotensin, Type 1/metabolism , Renin/genetics , Renin/metabolism , Renin-Angiotensin System/genetics , Transcription, Genetic , TransfectionABSTRACT
BACKGROUND: In peripheral blood, DNA methylation (DNAm) patterns in inflammatory bowel disease patients reflect inflammatory status rather than disease status. Here, we examined DNAm in diseased rectal mucosa from ulcerative colitis (UC) patients, focusing on constituent cell types with the goal of identifying therapeutic targets for UC other than the immune system. We profiled DNAm of rectal mucosal biopsies of pediatric UC at diagnosis (n = 211) and non-IBD control (n = 85) patients and performed epigenome-wide association studies (EWAS) of specific cell types to understand DNAm changes in epithelial, immune and fibroblast cells across disease states, course, and clinical outcomes. We also examined longitudinal analysis on follow-up samples (n = 73), and comparisons were made among patients with clinical outcomes including those undergoing colectomy versus those who did not. Additionally, we included RNA-seq from the same subjects to assess the impact of CpG sites on the transcription of nearby genes during the disease course. RESULTS: At diagnosis, UC rectal mucosa exhibited a lower proportion of epithelial cells and fibroblasts, and higher proportion of immune cells, in conjunction with variation in the DNAm pattern. While treatment had significant effects on the methylation signature of immune cells, its effects on fibroblasts and epithelial cells were attenuated. Individuals who required colectomy exhibited cell composition and DNAm patterns at follow-up more similar to disease onset than patients who did not require colectomy. Combining these results with gene expression profiles, we identify CpG sites whose methylation patterns are most consistent with a contribution to poor disease outcomes and could thus be potential therapeutic targets. CONCLUSIONS: Cell-specific epigenetic changes in the rectal mucosa in UC are associated with disease severity and outcome. Current therapeutics may more effectively target the immune than the epithelial and fibroblast compartments.
Subject(s)
Colitis, Ulcerative , Crohn Disease , Child , Humans , Colitis, Ulcerative/genetics , DNA Methylation , Crohn Disease/genetics , Rectum/surgery , Mucous Membrane/metabolismABSTRACT
Recently, we identified 1,189 CpG sites whose DNA methylation level in blood associated with Crohn's disease. Here, we examined associations between DNA methylation and genetic variants to identify methylation quantitative trait loci across disease states in (1) 402 blood samples from 164 newly diagnosed pediatric Crohn's disease patients taken at 2 time points (diagnosis and follow-up), and 74 non-inflammatory bowel disease controls, (2) 780 blood samples from a non-Crohn's disease adult population, and (3) 40 ileal biopsies (17 Crohn's disease cases and 23 non-inflammatory bowel disease controls) from group (1). Genome-wide DNAm profiling and genotyping were performed using the Illumina MethylationEPIC and Illumina Multi-Ethnic arrays. SNP-CpG associations were identified via linear models adjusted for age, sex, disease status, disease subtype, estimated cell proportions, and genotype-based principal components. In total, we observed 535,448 SNP-CpG associations between 287,881 SNPs and 12,843 CpG sites (P < 8.21 × 10-14). Associations were highly consistent across different ages, races, disease states, and tissue types, suggesting that the majority of these methylation quantitative trait loci participate in common gene regulation. However, genes near CpGs associated with inflammatory bowel disease SNPs were enriched for 18 KEGG pathways relevant to inflammatory bowel disease-linked immune function and inflammatory responses. We observed suggestive evidence for a small number of tissue-specific associations and disease-specific associations in ileum, though larger studies will be needed to confirm these results. Our study concludes that the vast majority of blood-derived methylation quantitative trait loci are common across individuals, though a subset may be involved in processes related to Crohn's disease. Independent cohort studies will be required to validate these findings.
Subject(s)
Crohn Disease , Adult , Child , Crohn Disease/genetics , DNA Methylation/genetics , Genome-Wide Association Study/methods , Genotype , Humans , Polymorphism, Single Nucleotide , Quantitative Trait LociABSTRACT
BACKGROUND: The gut and oral microbiome have independently been shown to be associated with inflammatory bowel disease (IBD). However, it is not known to what extent gut and oral microbial disease markers converge in terms of their composition in IBD. Further, the spatial and temporal variation within the oral microenvironments of IBD remain to be elucidated. PATIENTS AND METHODS: We used a prospectively recruited cohort of patients with IBD (nâ =â 47) and unrelated healthy control patients (nâ =â 18) to examine the spatial and temporal distribution of microbiota within the various oral microenvironments, represented by saliva, tongue, buccal mucosa, and plaque, and compared them with stool. Microbiome characterization was performed using 16S rRNA gene sequencing. RESULTS: The oral microbiome displayed IBD-associated dysbiosis, in a site- and taxa-specific manner. Plaque samples depicted a relatively severe degree of dysbiosis, and the disease-associated dysbiotic bacterial groups were predominantly the members of the phylum Firmicutes. Our 16S rRNA gene analyses show that oral microbiota can distinguish patients with IBD from healthy control patients, with salivary microbiota performing the best, closely matched by stool and other oral sites. Longitudinal profiles of microbial composition suggest that some taxa are more consistently perturbed than others, preferentially in a site-dependent fashion. CONCLUSIONS: Collectively, these data indicate the potential of using oral microbial profiles in screening and monitoring patients with IBD. Furthermore, these results support the importance of spatial and longitudinal microbiome sampling to interpret disease-associated dysbiotic states and eventually to gain insights into disease pathogenesis.
Subject(s)
Inflammatory Bowel Diseases , Microbiota , Mouth/microbiology , Dysbiosis/diagnosis , Feces/microbiology , Humans , Inflammatory Bowel Diseases/classification , Inflammatory Bowel Diseases/microbiology , RNA, Ribosomal, 16S/geneticsABSTRACT
Background: Inflammatory bowel disease (IBD) mainly consists of Crohn's disease (CD) and ulcerative colitis (UC). About 10%-15% of patients with IBD cannot be firmly diagnosed with CD or UC; hence, they are initially diagnosed as inflammatory bowel disease unclassified (IBD-U). Having a firm diagnosis is clearly preferred to guide treatment choices, and better understanding of the nature of IBD-U is required. Methods: We performed an analysis of a subset of pediatric subjects from an inception IBD cohort of patients initially enrolled in a prospective multicenter study (the RISK study). Initial diagnosis and 2-year follow-up data from the subjects diagnosed with IBD-U were analyzed. An expert panel verified final diagnosis using predefined criteria as a guide. Serological and disease-relevant ileal and rectal tissue gene expression profiles were investigated. The use and the time to initiate anti-TNFα treatment was analyzed among the outcome groups. Results: A total of 1411 subjects were enrolled with initial diagnosis of IBD, and among them, 136 subjects were initially diagnosed as IBD-U at enrollment. And 26% were reclassified as UC and 14% as CD within 2 years of diagnosis, while 60% remained as IBD-U. Of those who were reclassified, there was a 2:1 ratio, UC (n = 35) to CD (n = 19). The molecular and serological features of IBD-U at the end of follow-up were very similar to UC and very different from CD. There was less likelihood of receiving anti-TNFα agents if the diagnosis was IBD-U compared with CD (P < 0.0001). Conclusions: In our cohort, 60% of the IBD-U subjects remained as unclassified at 2 years; of those subsequently classified, a higher percentage followed a course more similar to UC. Most of the IBD-U subjects at diagnosis had serological and molecular signatures that are very similar to UC. Although the atypical presentations made the clinician to make an interim diagnosis of IBD-U, results of the molecular and serological factors performed at the time of diagnosis suggests that they were very similar to UC. However, long-term studies are needed to better understand the natural history and molecular characterization of pediatric onset IBD-U. 10.1093/ibd/izy136_video1Video 1.Video 1. Watch now at https://academic.oup.com/ibd/article-lookup/doi/10.1093/ibd/izy136izy136.video15791389938001.
Subject(s)
Biomarkers/analysis , Inflammatory Bowel Diseases/classification , Inflammatory Bowel Diseases/diagnosis , Transcriptome , Adolescent , Child , Child, Preschool , Female , Follow-Up Studies , Humans , Immunosuppressive Agents/therapeutic use , Infant , Infant, Newborn , Inflammatory Bowel Diseases/blood , Inflammatory Bowel Diseases/genetics , Male , Prognosis , Prospective StudiesABSTRACT
Angiotensin II (Ang II) is involved in induction and progression of renal damage in diabetes. Angiotensin converting enzyme 2 (ACE2) is highly expressed in the kidney and has been shown to be renoprotective by degrading Ang II to Ang-(1-7). A disintegrin and metalloproteinase 17 (ADAM17)-mediated shedding of renal ACE2 contribute to diabetic nephropathy pathogenesis. Lifestyle modification and metformin are recommended as initial therapies for most patients with type 2 diabetes. The aim of this study was to investigate whether exercise training and/or metformin improve glucose homeostasis and albuminuria and downregulate renal ADAM17 and ACE2 shedding in db/db mice. Seven-week-old normal and db/db mice were subjected either to a sedentary existence or exercise training with and without metformin (150âmg/kg per day) for 10 weeks. Exercise training significantly lowered blood glucose, urinary albumin and ACE2 excretion in db/db mice. ADAM17 and ACE2 proteins were co-localized in cortical tubules of the kidney, indicating a possible interaction. Metformin treatment was effective in lowering hyperglycemia only during the first 2 weeks of treatment. Increased renal ADAM17 in 17-week-old db/db mice was corrected by physical exercise but not metformin. In addition, exercise training reduced plasma triglycerides and enhanced insulin levels of db/db mice. In conclusion, exercise training alone and in combination with metformin prevented shedding of renal ACE2 by decreasing ADAM17 protein. Urinary ACE2 could serve as a prognostic tool for the progression of kidney damage and its attenuation by exercise may partially contribute to its renal protection.
Subject(s)
Albuminuria/prevention & control , Diabetes Mellitus, Experimental/therapy , Exercise Therapy/methods , Peptidyl-Dipeptidase A/urine , Physical Conditioning, Animal/physiology , Albuminuria/metabolism , Angiotensin-Converting Enzyme 2 , Animals , Combined Modality Therapy , Diabetes Complications/prevention & control , Diabetes Complications/urine , Diabetes Mellitus, Experimental/complications , Diabetes Mellitus, Experimental/genetics , Diabetes Mellitus, Experimental/urine , Diabetes Mellitus, Type 2/complications , Diabetes Mellitus, Type 2/therapy , Diabetes Mellitus, Type 2/urine , Hypoglycemic Agents/therapeutic use , Kidney/metabolism , Male , Metformin/therapeutic use , Mice , Mice, Transgenic , Protein Transport , Receptors, Leptin/geneticsABSTRACT
BACKGROUND: Dendritic cells (DCs) are important mediators of innate and adaptive immune responses, but the gene networks governing their lineage differentiation and maturation are poorly understood. To gain insight into the mechanisms that promote human DC differentiation and contribute to the acquisition of their functional phenotypes, we performed genome-wide base-resolution mapping of 5-methylcytosine in purified monocytes and in monocyte-derived immature and mature DCs. RESULTS: DC development and maturation were associated with a great loss of DNA methylation across many regions, most of which occurs at predicted enhancers and binding sites for known transcription factors affiliated with DC lineage specification and response to immune stimuli. In addition, we discovered novel genes that may contribute to DC differentiation and maturation. Interestingly, many genes close to demethylated CG sites were upregulated in expression. We observed dynamic changes in the expression of TET2, DNMT1, DNMT3A and DNMT3B coupled with temporal locus-specific demethylation, providing possible mechanisms accounting for the dramatic loss in DNA methylation. CONCLUSIONS: Our study is the first to map DNA methylation changes during human DC differentiation and maturation in purified cell populations and will greatly enhance the understanding of DC development and maturation and aid in the development of more efficacious DC-based therapeutic strategies.
ABSTRACT
Alterations within the renal renin angiotensin system play a pivotal role in the development and progression of cardiovascular and renal disease. Angiotensin converting enzyme 2 (ACE2) is highly expressed in renal tubules and has been shown to be renoprotective in diabetes. The protease, a disintegrin and metalloprotease (ADAM) 17, is involved in the ectodomain shedding of several transmembrane proteins including ACE2. Renal ACE2 and ADAM17 were significantly increased in db/db mice compared to controls. We investigated the effect of the insulin sensitizer, rosiglitazone, on albuminuria, renal ADAM17 protein expression and ACE2 shedding in db/db diabetic mice. Rosiglitazone treatment of db/db mice normalized hyperglycemia, attenuated renal injury and decreased urinary ACE2 and renal ADAM17 protein expression. Urinary excreted ACE2 is enzymatically active. Western blot analysis of urinary ACE2 demonstrated two prominent immunoreactive bands at approximately 70 & 90 kDa. The predominant immunoreactive band is approximately 20 kDa shorter than the one demonstrated for kidney lysate, indicating possible ectodomain shedding of active renal ACE2 in the urine. Therefore, it is tempting to speculate that renoprotection of rosiglitazone could be partially mediated via downregulation of renal ADAM17 and ACE2 shedding. In addition, there was a positive correlation between blood glucose, urinary albumin, plasma glucagon, and triglyceride levels with urinary ACE2 excretion. In conclusion, urinary ACE2 could be used as a sensitive biomarker of diabetic nephropathy and for monitoring the effectiveness of renoprotective medication.