Prenatal hypoxia exposure induced methylomic and transcriptomic alterations in rat hearts [BiSulfite-seq]
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ABSTRACT: Antenatal hypoxia has critial impacts on fetal heart development. The molecular mechanism of the antenaltal hypoxia effect on the heart development is still unknown. We performed DNA methylome and transcriptome analyses of antenatal hypoxia induced rat fetal and adult offspring hearts to understand the hypoxia-mediated epigenomic programming in the heart development. Heart tissue from fetal (E21) and adult rat (5 months old) were collected. mRNA and genomic DNA methylation profiles of the heart tissue were generated by RNAseq and reduced representation bisulfite seuqencing (RRBS) techniques. We found 323 and 112 differential expressed genes between control and hypoxia groups in the fetal and adult hearts, respectively. Meanwhile, 2828 and 2193 differential methylated regions were identified in the fetal and adult hearts. Furthermore, opposite gobal DNA methylation pattern changes in transcription start site regions (TSS ± 1kb) were observed between fetal and adult hearts. Combining transcriptome, data indicates a significant difference in the responding genes and pathways between fetal and adult hearts in responding to the antenatal hypoxia. Our study provides an initial framework and new insights into fetal hypoxia-mediated epigenetic programming of pro-inflammatory phenotype in the heart development, linking antenatal stress, and developmental programming of heart vulnerability to disease later in life.
Project description:Antenatal hypoxia has critial impacts on fetal heart development. The molecular mechanism of the antenaltal hypoxia effect on the heart development is still unknown. We performed DNA methylome and transcriptome analyses of antenatal hypoxia induced rat fetal and adult offspring hearts to understand the hypoxia-mediated epigenomic programming in the heart development. Heart tissue from fetal (E21) and adult rat (5 months old) were collected. mRNA and genomic DNA methylation profiles of the heart tissue were generated by RNAseq and reduced representation bisulfite seuqencing (RRBS) techniques. We found 323 and 112 differential expressed genes between control and hypoxia groups in the fetal and adult hearts, respectively. Meanwhile, 2828 and 2193 differential methylated regions were identified in the fetal and adult hearts. Furthermore, opposite gobal DNA methylation pattern changes in transcription start site regions (TSS ± 1kb) were observed between fetal and adult hearts. Combining transcriptome, data indicates a significant difference in the responding genes and pathways between fetal and adult hearts in responding to the antenatal hypoxia. Our study provides an initial framework and new insights into fetal hypoxia-mediated epigenetic programming of pro-inflammatory phenotype in the heart development, linking antenatal stress, and developmental programming of heart vulnerability to disease later in life.
Project description:Cardiac hypertrophy has been well-characterized at the level of transcription. During cardiac hypertrophy, genes normally expressed primarily during fetal heart development are re-expressed, and this fetal gene program is believed to be a critical component of the hypertrophic process. Recently, alternative splicing of mRNA transcripts has been shown to be temporally regulated during heart development, leading us to consider whether fetal patterns of splicing also reappear during hypertrophy.We hypothesized that patterns of alternative splicing occurring during heart development are recapitulated during cardiac hypertrophy. Here we present a whole-transcriptome study of isoform expression during pressure-overload cardiac hypertrophy induced by 10 days of transverse aortic constriction (TAC) in rats and in developing fetal rat hearts compared to sham-operated adult rat hearts, using high-throughput sequencing of poly(A) tail mRNA. Quantification of isoform expression in fetal rat hearts, pressure-overloaded rat hearts, and sham-operated rat hearts by Illumina GAIIx in triplicate
Project description:Cardiac hypertrophy has been well-characterized at the level of transcription. During cardiac hypertrophy, genes normally expressed primarily during fetal heart development are re-expressed, and this fetal gene program is believed to be a critical component of the hypertrophic process. Recently, alternative splicing of mRNA transcripts has been shown to be temporally regulated during heart development, leading us to consider whether fetal patterns of splicing also reappear during hypertrophy.We hypothesized that patterns of alternative splicing occurring during heart development are recapitulated during cardiac hypertrophy. Here we present a whole-transcriptome study of isoform expression during pressure-overload cardiac hypertrophy induced by 10 days of transverse aortic constriction (TAC) in rats and in developing fetal rat hearts compared to sham-operated adult rat hearts, using high-throughput sequencing of poly(A) tail mRNA.
Project description:A few reports have implicated specific lncRNAs in cardiac development or failure, but precise details of lncRNAs expressed in hearts and how their expression may be altered during embryonic heart development or by adult heart disease is unknown. By comparing lncRNA profiles of normal embryonic (~E14), normal adult, and hypertrophied adult hearts we defined a distinct fetal lncRNA abundance signature that includes 157 lncRNAs differentially expressed compared to adults (fold-change ≥ 50%, FDR=0.02), and which was only poorly recapitulated in hypertrophied hearts (17 differentially expressed lncRNAs; 13 of these observed in embryonic hearts). Analysis of protein-coding mRNAs from the same samples identified 22 concordantly and 11 reciprocally regulated mRNAs within 10 kb of dynamically expressed lncRNAs, reciprocal relationships of lncRNA and mRNA levels was validated for the Mccc1 and Relb genes using in vitro lncRNA knockdown in C2C12 cells. Network analysis suggested a central role for lncRNAs in modulating NFkappaB- and CREB1-regulated genes during embryonic heart growth and identified multiple mRNAs within these pathways that are also regulated, but independently of lncRNAs. Cardiac polyadenylated RNA (mRNA and lncRNA) profiles were generated from C57BL/6J mouse hearts were generated on Illumina HiSeq 2000 instruments. 7 independent E13.5 hearts, 12 adult hearts (6 at 6 weeks of age, 6 at 16 weeks of age), 4 sham-operated hearts at 12 weeks of age, and 4 hearts after 4 weeks of pressure overload (TAC) at 12 weeks of age.
Project description:In response to heart failure (HF), the heart reacts by repressing adult genes and expressing fetal genes, thereby returning to a more fetal-like gene profile. To identify genes involved in this process, we carried out transcriptional analysis on murine hearts at different stages of development and adult mice with HF. Our screen identified 5-oxoprolinase (OPLAH), a member of the -Glutamyl cycle, that functions by scavenging 5-oxoproline. OPLAH depletion occurred as a result of cardiac injury, leading to elevated 5-oxoproline and oxidative stress, whereas OPLAH overexpression improved cardiac function after ischemic injury. In HF patients we observed elevated plasma 5-oxoproline levels, which were associated with a worse clinical outcome. Understanding and modulating fetal-like genes in the failing heart may lead to potential novel diagnostic, prognostic and therapeutic options in HF.
Project description:A few reports have implicated specific lncRNAs in cardiac development or failure, but precise details of lncRNAs expressed in hearts and how their expression may be altered during embryonic heart development or by adult heart disease is unknown. By comparing lncRNA profiles of normal embryonic (~E14), normal adult, and hypertrophied adult hearts we defined a distinct fetal lncRNA abundance signature that includes 157 lncRNAs differentially expressed compared to adults (fold-change ≥ 50%, FDR=0.02), and which was only poorly recapitulated in hypertrophied hearts (17 differentially expressed lncRNAs; 13 of these observed in embryonic hearts). Analysis of protein-coding mRNAs from the same samples identified 22 concordantly and 11 reciprocally regulated mRNAs within 10 kb of dynamically expressed lncRNAs, reciprocal relationships of lncRNA and mRNA levels was validated for the Mccc1 and Relb genes using in vitro lncRNA knockdown in C2C12 cells. Network analysis suggested a central role for lncRNAs in modulating NFkappaB- and CREB1-regulated genes during embryonic heart growth and identified multiple mRNAs within these pathways that are also regulated, but independently of lncRNAs.
Project description:Semilunar valve leaflets have a well-described trilaminar histoarchitecture with a sophisticated elastic fiber network. It was previously proposed that elastin-containing fibers play a subordinate role in early human cardiac valve development; however, this assumption was based on data obtained from mouse models and human second and third trimester tissues. Here, we systematically analyzed tissues from human fetal first (4-12 weeks) and second (13-18 weeks) trimester, adolescent (14-19 years) and adult (50-55 years) hearts to monitor the temporal and spatial distribution of elastic fibers, focusing on semilunar valves. Global gene expression analyses revealed that the transcription of genes essential for elastic fiber formation starts early within the first trimester. These data were confirmed by quantitative PCR and immunohistochemistry employing antibodies that recognize fibronectin, fibrilin-1, -2 and -3, EMILIN-1, fibulin-4 and fibulin-5, which were all expressed at the onset of cardiac cushion formation (~week 4 of development). Tropoelastin/ elastin protein expression was first detectable in leaflets of 7-week hearts. We revealed that immature elastic fibers are organized in early human cardiovascular development, and mature elastin-containing fibers first evolve in semilunar valves when blood pressure and heartbeat accelerate. Our findings provide a conceptual framework with the potential to lead to novel hypotheses in human cardiac valve development and disease. Total RNA obtained from fetal cardiac valve cushions, developed fetal heart valves, adolescent heart valves, and adult heart valves.
Project description:This project seeks to understand the metabolic consequences of gestational hypoxia on fetal, newborn, and adult plasma, arteries and other tissues using a sheep model of fetal growth restriction. Specifically we are interested testing the hypothesis that gestational hypoxia will result in discernable differences in glucose and lipid metabolism in tissues and plasma as well influence indicators of oxidative stress and inflammation. These studies aim to delineate pathways and biomarkers that help explain how hypoxia leads to the development of neonatal as well as adult-onset diseases associated with chronic hypoxia that are inter-related with fetal growth restriction. From a vascular perspective this includes cerebrovascular hemorrhage and pulmonary hypertension in the newborn, but more broadly it includes development of diseases later in life including diabetes, hypertension, and coronary artery disease.
Project description:To gain insight into the molecular regulation of human heart development, a detailed comparison of the mRNA and miRNA transcriptomes across differentiating human-induced pluripotent stem cell (hiPSC)–derived cardiomyocytes and biopsies from fetal, adult, and hypertensive human hearts was performed. Gene ontology analysis of the mRNA expression levels of the hiPSCs differentiating into cardiomyocytes revealed 3 distinct groups of genes: pluripotent specific, transitional cardiac specification, and mature cardiomyocyte specific. Hierarchical clustering of the mRNA data revealed that the transcriptome of hiPSC cardiomyocytes largely stabilizes 20 days after initiation of differentiation. Nevertheless, analysis of cells continuously cultured for 120 days indicated that the cardiomyocytes continued to mature toward a more adult-like gene expression pattern. Analysis of cardiomyocyte-specific miRNAs (miR-1, miR-133a/b, and miR-208a/b) revealed a miRNA pattern indicative of stem cell to cardiomyocyte specification. A biostatistitical approach integrated the miRNA and mRNA expression profiles revealing a cardiomyocyte differentiation miRNA network and identified putative mRNAs targeted by multiple miRNAs. Together, these data reveal the miRNA network in human heart development and support the notion that overlapping miRNA networks re-enforce transcriptional control during developmental specification. Comparison of mRNA expression profiling of differentiating human-induced pluripotent stem cell (hiPSC)–derived cardiomyocytes, biopsies from fetal, adult and hypertensive human hearts and primary cardiomyocytes
Project description:Background: Fetal hypoxia causes vital, systemic, developmental malformations in the fetus, particularly in the brain, and increases the risk of diseases in later life. We previously demonstrated that fetal hypoxia exposure increases the susceptibility of the neonatal brain to hypoxic-ischemic insult. Herein, we investigate the effect of fetal hypoxia on programming of cell-specific transcriptomes in the brain of neonatal rats.