MP254VDR GENE EXPRESSION AND GLOBAL DNA METHYLATION ARE ASSOCIATED WITH INSULIN RESISTANCE IN ESRD PATIENTS ON CHRONIC HAEMODIALYSIS

Abstract Acute myeloid leukemia (AML) represents a highly heterogeneous myeloid stem cell disorder classified based on various genetic defects. Besides genetic alterations, epigenetic changes are recognized as an additional mechanism contributing to leukemogenesis, but insight into the latter process remains minor. Using a combination of Methyl-CpG-Immunoprecipitation (MCIp-chip) and MALDI-TOF analysis of bisulfite-treated DNA in a cohort of 196 AML patients we previously demonstrated that (cyto)genetically defined AML subtypes, including CBFB-MYH11, AML-ETO, NPM1-mut, CEBPA-mut or IDH1/2-mut subtypes, express specific DNA-methylation profiles (Gebhard et al, Leukemia, 2018). A fraction of AML patients (5/196) displayed a unique abnormal hypermethylation profile that was completely distinct from any other AML subtype. These patients present immature leukemia (FAB M0, M1) with various chromosomal aberrations but very few mutations (e.g. no IDH1/2, KRAS, DNMT3A) that might explain the CpG island methylator phenotype (CIMP) phenotype. The CIMP patients showed high resemblance with a recently reported CEBPA methylated subgroup (Wouters et al, 2007 and Figueroa et al, 2009), which we confirmed by MCIp-chip and MALDI-TOF analysis. To explore the whole range of epigenetic alterations in the CIMP-AML patients we performed in-depth global DNA methylation and gene expression analyses (MCIp-seq and RNA-seq) in 45 AML and 12 CIMP patients from both studies. Principle component analysis and t-distributed stochastic neighbor embedding (t-SNE) revealed that CIMP patients express a unique DNA-methylation and gene-expression signature that separated them from all other AMLs. We could discriminate promoter methylation from non-promoter methylation by selecting MCIp-seq peaks within 3kb around TSS. Promoter hypermethylation was highly associated with repression of genes (PCC = -0.053, p-value = 0.00075). Hypermethylation of non-promoter regions was more strongly associated with upregulation of genes (PCC = 0.046, p-value = 4.613e-06). Interestingly, differentially methylated regions also showed a positive association with myeloid lineage CTCF binding sites (27% vs 18% expected, p-value < 2.2e-16 in a chi-square test of independence). Methylation of CTCF sites causes loss of CTCF binding, which has been reported to disrupt boundaries between so-called topologically associated domains (TADs), allowing enhancers located in a particular TAD to become accessible to genes in adjacent TADs and affect their transcription. Whether this is the case is under investigation. In this study we particularly focused on the role of hypermethylation of promoters in CIMP-AMLs. Promoters of many transcriptional regulators that are involved in the differentiation of myeloid lineages of which several are frequently mutated in AML were hypermethylated and repressed, including CEBPA, CEBPD, IRF8, GATA2, KLF4, MITF or MAFB. Notably, HMGA2, a critical regulator of myeloid progenitor expansion, exhibited the largest degree of CIMP promoter hypermethylation compared to the other AMLs, accompanied by a reduction in gene expression. Moreover, multiple members of the HOXB family and KLF1 (erythroid differentiation) were methylated and repressed as well. In addition, these patients frequently showed hypermethylation of many chromatin factors (e.g. LMNA, CHD7 or TET2). Hypermethylation of the TET2 promoter could result in a loss of maintenance DNA demethylation and therefore successive hypermethylation at CpG islands. We carried out regulome-capture-bisulfite sequencing on CIMP-AMLs compared to other AML samples and normal blood cell controls and confirmed methylation of the same transcription and chromatin factor promoters. We conclude that these leukemias represent very primitive HSCPs which are blocked in differentiation into multiple hematopoietic lineages, due to the absence of regulators of these lineages. Although the underlying cause for the extreme hypermethylation signature is still subject to ongoing studies, the consequence of promoter hypermethylation is silencing of key lineage regulators causing the differentiation arrest in these cells. We argue that these patients may particularly benefit from therapies that revert DNA methylation. Disclosures Ehninger: Cellex Gesellschaft fuer Zellgewinnung mbH: Employment, Equity Ownership; GEMoaB Monoclonals GmbH: Employment, Equity Ownership; Bayer: Research Funding. Thiede:AgenDix: Other: Ownership; Novartis: Honoraria, Research Funding.

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THU0003 ALTERED DNA METHYLATION AND DIFFERENTIAL EXPRESSION OF GENES INFLUENCING CARDIOVASCULAR RISK AND IMMUNITY IN CD4+ T CELLS FROM SUBJECTS WITH PSORIATIC ARTHRITIS.

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2020-eular.4492 ◽

2020 ◽

Vol 79 (Suppl 1) ◽

pp. 214.1-214

Author(s):

I. Arias de la Rosa ◽

M. D. López Montilla ◽

J. Rodríguez ◽

E. Ballester ◽

C. Torres-Granados ◽

...

Keyword(s):

Gene Expression ◽

Risk Factors ◽

Insulin Resistance ◽

Dna Methylation ◽

T Cells ◽

Cardiovascular Risk ◽

Research Support ◽

Insulin Resistant ◽

Genome Wide ◽

Differentially Methylated Genes

Background:Cardiovascular risk factors are increased in Psoriatic Arthritis (PsA). In fact, around 60% out of PsA patients display insulin resistance (IR), a hallmark of metabolic syndrome, which might significantly contribute to the cardiovascular disease. Latest studies suggested that inflammatory and metabolic disorders may be under epigenetic control, including DNA methylation. DNA methylation is an unexplored area in the field of PsA.Objectives:To study the alterations in the genome-wide DNA methylation profile of CD4+T cells from PsA patients and its relationship with its pathology and the risk of cardiovascular comorbidity.Methods:Twenty healthy controls (HC) and 20 PsA patients were included in the study. PsA patients were classified into insulin resistant and non-insulin resistant according to HOMA-IR index. CD4+T lymphocytes were isolated from peripheral blood by positive immunomagnetic selection. The Illumina Infinium MethylationEPIC Beadchip was used to obtain DNA methylation profiles across approximately 850,000 CpGs (TSS1500, TSS200, 5UTR, 3UTR, first exon, gene body). Beta values (β) estimating methylation levels were obtained at each CpG site, and differentially methylated genes (DMG) between PsA and HC were identified. Functional classification of these genes was carried out through gene ontology analysis (PANTHER database). Gene expression analysis of the selected genes was also evaluated by RT-PCR. Vascular parameters including carotid intima-media thickness (cIMT) and endothelial function was analyzed by ecodoppler and periflux respectively.Results:The genome-wide methylation analysis identified 112 DMGs including 41 hypomethylated and 71 hypermethylated. These differentially methylated genes were enriched with several signaling pathways and disease categories including immune response, metabolic processes, oxidative stress, vascular and inflammatory pathways. The altered gene expression of selected genes with altered methylation levels in PsA was also validated. Correlation and association analysis of these DMGs with clinical and analytical variables, cardiovascular risk factors and endothelial microvascular function revealed that the degree of methylation of these genes was significantly associated with cIMT (IGF1R, NDRG3, SMYD3, HLA-DRB1, WDR70), arterial pressure (METT5D1, NRDG3, ADAM17, SMYD3, WNK1, CBX1), insulin resistance (AKAP13, SEMA6D, PLCB1), altered lipid profile and atherogenic index (MYBL1, METT5D1, MAN2A1, SLC1A7, SEMA6D, PLCB1, TLK1, SDK1, CBX1), inflammation (MYBL1, NDUFA5, METT5D1, SEMA6D, PLCB1, TLK1), and endothelial dysfunction (ADAMST10, GPCPD1, CCDC88A). In addition, this analysis also identified 435 DMGs including 280 hypomethylated and 155 hypermethylated in CD4+T cells from IR-PsA vs non IR-PsA patients. Between these two groups of PsA patients, CHUK, SERINC1, RUNX1, TTYH2, TXNDC11, FAF1, BICD1, SCD5, PDE5A, FAS, NFIA and GRP75 displayed the most significantly altered methylation, suggesting the role of these genes in the metabolic complications associated with PsA.Conclusion:These findings help our understanding of the pathogenesis of PsA and advance epigenetic studies in regards to this disease and the cardiometabolic comorbidities associated. Funded by ISCIII (PI17/01316 and RIER RD16/0012/0015) co-funded with FEDER.Disclosure of Interests:Iván Arias de la Rosa: None declared, María Dolores López Montilla Speakers bureau: Celgene, Javier Rodríguez: None declared, Esteban Ballester: None declared, Carmen Torres-Granados: None declared, Carlos Perez-Sanchez: None declared, Maria del Carmen Abalos-Aguilera: None declared, Gómez García Ignacio: None declared, Desiree Ruiz: None declared, Alejandra M. Patiño-Trives: None declared, María Luque-Tévar: None declared, Eduardo Collantes-Estévez Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Chary Lopez-Pedrera Grant/research support from: ROCHE and Pfizer., Alejandro Escudero Contreras Grant/research support from: ROCHE and Pfizer, Speakers bureau: ROCHE, Lilly, Bristol and Celgene., Nuria Barbarroja Puerto Grant/research support from: ROCHE and Pfizer., Speakers bureau: ROCHE and Celgene.

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Global DNA Methylation Is Associated With Insulin Resistance: A Monozygotic Twin Study

Diabetes ◽

10.2337/db11-1048 ◽

2011 ◽

Vol 61 (2) ◽

pp. 542-546 ◽

Cited By ~ 114

Author(s):

J. Zhao ◽

J. Goldberg ◽

J. D. Bremner ◽

V. Vaccarino

Keyword(s):

Insulin Resistance ◽

Dna Methylation ◽

Twin Study ◽

Monozygotic Twin ◽

Global Dna Methylation

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Intergenerational Effect of Early Life Exposure to Permethrin: Changes in Global DNA Methylation and in Nurr1 Gene Expression

Toxics ◽

10.3390/toxics3040451 ◽

2015 ◽

Vol 3 (4) ◽

pp. 451-461 ◽

Cited By ~ 20

Author(s):

Laura Bordoni ◽

Cinzia Nasuti ◽

Maria Mirto ◽

Fabio Caradonna ◽

Rosita Gabbianelli

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Early Life ◽

Global Dna Methylation ◽

Early Life Exposure ◽

Intergenerational Effect

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Integrative analysis of glucometabolic traits, adipose tissue DNA methylation and gene expression identifies epigenetic regulatory mechanisms of insulin resistance and obesity in African Americans

10.2337/figshare.12896636.v1 ◽

2020 ◽

Author(s):

Ada Admin ◽

Neeraj K. Sharma ◽

Mary E. Comeau ◽

Dennis Montoya ◽

Matteo Pellegrini ◽

...

Keyword(s):

Gene Expression ◽

Insulin Resistance ◽

Dna Methylation ◽

Adipose Tissue ◽

Insulin Sensitivity ◽

African Americans ◽

Association Studies ◽

Regulation Of Gene Expression ◽

Transcript Level ◽

Matsuda Index

Decline in insulin sensitivity due to dysfunction of adipose tissue (AT) is one of the earliest pathogenic events in Type 2 Diabetes. We hypothesize that differential DNA methylation (DNAm) controls insulin sensitivity and obesity by modulating transcript expression in AT. Integrating AT DNAm profiles with transcript profile data measured in a cohort of 230 African Americans from AAGMEx cohort, we performed cis-expression quantitative trait methylation (cis-eQTM) analysis to identify epigenetic regulatory loci for glucometabolic trait-associated transcripts. We identified significantly associated CpG-regions for 82 transcripts (FDR-P<0.05). The strongest eQTM locus was observed for the proopiomelanocortin (POMC; r= -0.632, P= 4.70X10-27) gene. Epigenome-wide association studies (EWAS) further identified 155, 46, and 168 CpG regions associated (FDR-P <0.05) with Matsuda index, SI and BMI, respectively. Intersection of EWAS, transcript level to trait association, and eQTM results, followed by causal inference test identified significant eQTM loci for 23 genes that were also associated with Matsuda index, SI and/or BMI in EWAS. These associated genes include FERMT3, ITGAM, ITGAX, and POMC. In summary, applying an integrative multi-omics approach, our study provides evidence for DNAm-mediated regulation of gene expression at both previously identified and novel loci for many key AT transcripts influencing insulin resistance and obesity.

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Genetic Profiling of Orbital Fibroblasts from Patients with Graves’ Orbitopathy

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/clinem/dgab035 ◽

2021 ◽

Author(s):

Giovanna Rotondo Dottore ◽

Ilaria Bucci ◽

Giulia Lanzolla ◽

Iacopo Dallan ◽

Angela Sframeli ◽

...

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Cell Proliferation ◽

Differential Gene Expression ◽

Global Dna Methylation ◽

Whole Genome ◽

Graves Orbitopathy ◽

Differential Gene ◽

And Control ◽

Orbital Fibroblasts

Abstract Context Graves’ orbitopathy (GO) is an autoimmune disease that persists when immunosuppression is achieved. Orbital fibroblasts from GO patients display peculiar phenotypes even if not exposed to autoimmunity, possibly reflecting genetic or epigenetic mechanisms, which we investigated here. Objective We aimed to explore potential genetic or epigenetic differences using primary cultures of orbital fibroblasts from GO and control patients. Methods Cell proliferation, hyaluronic acid (HA) secretion, and HA synthases (HAS) were measured. Next-generation sequencing and gene expression analysis of the whole genome were performed, as well as real-time-PCR of selected genes and global DNA methylation assay on orbital fibroblasts from 6 patients with GO and 6 control patients from a referral center. Results Cell proliferation was higher in GO than in control fibroblasts. Likewise, HA in the cell medium was higher in GO fibroblasts. HAS-1 and HAS-2 did not differ between GO and control fibroblasts, whereas HAS-3 was more expressed in GO fibroblasts. No relevant gene variants were detected by whole-genome sequencing. However, 58 genes were found to be differentially expressed in GO compared with control fibroblasts, and RT-PCR confirmed the findings in 10 selected genes. We postulated that the differential gene expression was related to an epigenetic mechanism, reflecting diverse DNA methylation, which we therefore measured. In support of our hypothesis, global DNA methylation was significantly higher in GO fibroblasts. Conclusions We propose that, following an autoimmune insult, DNA methylation elicits differential gene expression and sustains the maintenance of GO.

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