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Differential DNA methylation correlates with differential expression of angiogenic factors in human heart failure.

ABSTRACT: Epigenetic mechanisms such as microRNA and histone modification are crucially responsible for dysregulated gene expression in heart failure. In contrast, the role of DNA methylation, another well-characterized epigenetic mark, is unknown. In order to examine whether human cardiomyopathy of different etiologies are connected by a unifying pattern of DNA methylation pattern, we undertook profiling with ischaemic and idiopathic end-stage cardiomyopathic left ventricular (LV) explants from patients who had undergone cardiac transplantation compared to normal control. We performed a preliminary analysis using methylated-DNA immunoprecipitation-chip (MeDIP-chip), validated differential methylation loci by bisulfite-(BS) PCR and high throughput sequencing, and identified 3 angiogenesis-related genetic loci that were differentially methylated. Using quantitative RT-PCR, we found that the expression of these genes differed significantly between CM hearts and normal control (p<0.01). Moreover, for each individual LV tissue, differential methylation showed a predicted correlation to differential expression of the corresponding gene. Thus, differential DNA methylation exists in human cardiomyopathy. In this series of heterogeneous cardiomyopathic LV explants, differential DNA methylation was found in at least 3 angiogenesis-related genes. While in other systems, changes in DNA methylation at specific genomic loci usually precede changes in the expression of corresponding genes, our current findings in cardiomyopathy merit further investigation to determine whether DNA methylation changes play a causative role in the progression of heart failure.

SUBMITTER: Movassagh M

PROVIDER: S-EPMC2797324 | biostudies-literature | 2010 Jan

REPOSITORIES: biostudies-literature

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Differential DNA methylation correlates with differential expression of angiogenic factors in human heart failure.

Movassagh Mehregan M Choy Mun-Kit MK Goddard Martin M Bennett Martin R MR Down Thomas A TA Foo Roger S-Y RS

PloS one 20100113 1

Epigenetic mechanisms such as microRNA and histone modification are crucially responsible for dysregulated gene expression in heart failure. In contrast, the role of DNA methylation, another well-characterized epigenetic mark, is unknown. In order to examine whether human cardiomyopathy of different etiologies are connected by a unifying pattern of DNA methylation pattern, we undertook profiling with ischaemic and idiopathic end-stage cardiomyopathic left ventricular (LV) explants from patients ...[more]

PMID: 20084101

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Project description:Heart failure (HF) is a leading cause of morbidity and mortality in the United States and worldwide. As a multifactorial syndrome with unpredictable clinical outcomes, identifying the common molecular underpinnings that drive HF pathogenesis remains a major focus of investigation. Disruption of cardiac gene expression has been shown to mediate a common final cascade of pathological hallmarks wherein the heart reactivates numerous developmental pathways. Although the central regulatory mechanisms that drive this cardiac transcriptional reprogramming remain unknown, epigenetic contributions are likely. In the current study, we examined whether the epigenome, specifically DNA methylation, is reprogrammed in HF to potentiate a pathological shift in cardiac gene expression. To accomplish this, we used paired-end whole genome bisulfite sequencing and next-generation RNA sequencing of left ventricle tissue obtained from seven patients with end-stage HF and three nonfailing donor hearts. We found that differential methylation was localized to promoter-associated cytosine-phosphate-guanine islands, which are established regulatory regions of downstream genes. Hypermethylated promoters were associated with genes involved in oxidative metabolism, whereas promoter hypomethylation enriched glycolytic pathways. Overexpression of plasmid-derived DNA methyltransferase 3A in vitro was sufficient to lower the expression of numerous oxidative metabolic genes in H9c2 rat cardiomyoblasts, further supporting the importance of epigenetic factors in the regulation of cardiac metabolism. Last, we identified binding-site competition via hypermethylation of the nuclear respiratory factor 1 (NRF1) motif, an established upstream regulator of mitochondrial biogenesis. These preliminary observations are the first to uncover an etiology-independent shift in cardiac DNA methylation that corresponds with altered metabolic gene expression in HF.NEW & NOTEWORTHY The failing heart undergoes profound metabolic changes because of alterations in cardiac gene expression, reactivating glycolytic genes and suppressing oxidative metabolic genes. In the current study, we discover that alterations to cardiac DNA methylation encode this fetal-like metabolic gene reprogramming. We also identify novel epigenetic interference of nuclear respiratory factor 1 via hypermethylation of its downstream promoter targets, further supporting a novel contribution of DNA methylation in the metabolic remodeling of heart failure.

Project description:BackgroundAnthracycline-induced cardiotoxicity is a well-known serious clinical entity. However, detailed mechanistic insights on how short-term administration leads to late and long-lasting cardiotoxicity, are still largely undiscovered. We hypothesize that chemotherapy provokes a memory effect at the level of epigenomic DNA modifications which subsequently lead to cardiotoxicity even years after cessation of chemotherapy.MethodsWe explored the temporal evolution of epigenetic modifiers in early and late cardiotoxicity due to anthracyclines by means of RNA-sequencing of human endomyocardial left ventricular biopsies and mass spectrometry of genomic DNA. Based on these findings, validation of differentially regulated genes was obtained by performing RT-qPCR. Finally, a proof-of-concept in vitro mechanistic study was performed to dissect some of the mechanistic aspects of epigenetic memory in anthracycline-induced cardiotoxicity.ResultsCorrelation of gene expression between late and early onset cardiotoxicity revealed an R 2 value of 0.98, demonstrating a total of 369 differentially expressed genes (DEGs, FDR < 0.05). of which 72% (n = 266) were upregulated, and 28% of genes, (n = 103) downregulated in later as compared to earlier onset cardiotoxicity. Gene ontology analysis showed significant enrichment of genes involved in methyl-CpG DNA binding, chromatin remodeling and regulation of transcription and positive regulation of apoptosis. Differential mRNA expression of genes involved in DNA methylation metabolism were confirmed by RT-qPCR in endomyocardial biopsies. In a larger biopsy cohort, it was shown that Tet2 was more abundantly expressed in cardiotoxicity biopsies vs. control biopsies and vs. non-ischemic cardiomyopathy patients. Moreover, an in vitro study was performed: following short-term doxorubicin treatment, H9c2 cells were cultured and passaged once they reached a confluency of 70%-80%. When compared to vehicle-only treated cells, in doxorubicin-treated cells, three weeks after short term treatment, Nppa, Nppb, Tet1/2 and other genes involved in active DNA demethylation were markedly upregulated. These alterations coincided with a loss of DNA methylation and a gain in hydroxymethylation, reflecting the epigenetic changes seen in the endomyocardial biopsies.ConclusionsShort-term administration of anthracyclines provokes long-lasting epigenetic modifications in cardiomyocytes both in vivo and in vitro, which explain in part the time lapse between the use of chemotherapy and the development of cardiotoxicity and, eventually, heart failure.

Project description:Background – Epigenetic alterations are stable modifications to chromatin structure that occur in response to environmental cues such as hypoxia or altered nutrient delivery. DNA methylation is a well-established and dynamic DNA modification that contributes to the regulation of gene expression. In the current study, we test the hypothesize that ischemic heart failure is defined by a distinct signature of DNA methylation that corresponds with altered expression of genes involved in cardiac ventricular dysfunction. Methods and Results – Using a methylation array, we quantified genome-wide DNA methylation of endomyocardial samples acquired from patients with ischemic (n = 6) or non-ischemic (n = 5) heart failure. RNA-sequencing analysis was performed in the same samples to identify transcriptomic changes (Fold Change > 1.5, Q < 0.05, FPKM > 2) associated with differential methylation (|Percent Change| > 5%, p < 0.05). Of the promoter-associated CpG Islands, which are well-established regions of negative transcriptional regulation, we identified a signature of robust hypermethylation. The methylation changes linked to significantly decreased transcripts included key fatty acid metabolic regulators (e.g. KLF15, AGPAT9, APOA1, and MXD4). Among the few hypomethylated and induced genes was PFKFB3, which encodes for the rate-limiting enzyme of glycolysis. Gene set enrichment analysis identified TGFβ as a nodal upstream regulator of the methylation changes, potentially supporting a role of DNA methylation in the increased fibrosis and apoptosis that accompanies ischemic heart failure. Conclusions – Our data identify that the DNA methylation signature recapitulates the pathologic hallmarks of ischemic heart failure. Furthermore, we show that differential DNA methylation of CpG islands within the promoter depict alterations in metabolic substrate utilization known to occur in ischemic heart failure, and may govern a return to the fetal-like metabolic program.

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Differential DNA methylation correlates with differential expression of angiogenic factors in human heart failure.

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Differential DNA methylation correlates with differential expression of angiogenic factors in human heart failure.

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