Integrated analysis of miRNA-mRNA interaction in pediatric dilated cardiomyopathy.

BACKGROUND: MicroRNAs (miRNAs) are short single-stranded nucleotides that can regulate gene expression. Although we previously evaluated the expression of miRNAs in pediatric dilated cardiomyopathy (DCM) by miRNA array, pathway prediction based on changes in mRNA expression has not been previously analyzed in this population. The current study aimed to determine the regulation of miRNA expression by miRNA-sequencing (miRNA-seq) and, through miRNA-sequencing (mRNA-seq), analyze their putative target genes and altered pathways in pediatric DCM hearts. METHODS: miRNA expression was determined by miRNA-seq [n = 10 non-failing (NF), n = 20 DCM]. Expression of a subset of miRNAs was evaluated in adult DCM patients (n = 11 NF, n = 13 DCM). miRNA-mRNA prediction analysis was performed using mRNA-seq data (n = 7 NF, n = 7 DCM) from matched samples. RESULTS: Expression of 393 miRNAs was significantly different (p < 0.05) in pediatric DCM patients compared to NF controls. TargetScan-based miRNA-mRNA analysis revealed 808 significantly inversely expressed genes. Functional analysis suggests upregulated pathways related to the regulation of stem cell differentiation and cardiac muscle contraction, and downregulated pathways related to the regulation of protein phosphorylation, signal transduction, and cell communication. CONCLUSIONS: Our results demonstrated a unique age-dependent regulation of miRNAs and their putative target genes, which may contribute to distinctive phenotypic characteristics of DCM in children. IMPACT: This is the first study to compare miRNA expression in the heart of pediatric DCM patients to age-matched healthy controls by RNA sequencing. Expression of a subset of miRNAs is uniquely dysregulated in children. Using mRNA-seq and miRNA-seq from matched samples, target prediction was performed. This study underscores the importance of pediatric-focused studies.

Serum response factor deletion 5 regulates phospholamban phosphorylation and calcium uptake.

AIMS: Pediatric dilated cardiomyopathy (pDCM) is characterized by unique age-dependent molecular mechanisms that include myocellular responses to therapy. We previously showed that pDCM, but not adult DCM patients respond to phosphodiesterase 3 inhibitors (PDE3i) by increasing levels of the second messenger cAMP and consequent phosphorylation of phospholamban (PLN). However, the molecular mechanisms involved in the differential pediatric and adult response to PDE3i are not clear. METHODS AND RESULTS: Quantification of serum response factor (SRF) isoforms from the left ventricle of explanted hearts showed that PDE3i treatment affects expression of SRF isoforms in pDCM hearts. An SRF isoform lacking exon 5 (SRFdel5) was highly expressed in the hearts of pediatric, but not adult DCM patients treated with PDE3i. To determine the functional consequence of expression of SRFdel5, we overexpressed full length SRF or SRFdel5 in cultured cardiomyocytes with and without adrenergic stimulation. Compared to a control adenovirus, expression of SRFdel5 increased phosphorylation of PLN, negatively affected expression of the phosphatase that promotes dephosphorylation of PLN (PP2Cε), and promoted faster calcium reuptake, whereas expression of full length SRF attenuated calcium reuptake through blunted phosphorylation of PLN. CONCLUSIONS: Taken together, these data indicate that expression of SRFdel5 in pDCM hearts in response to PDE3i contributes to improved function through regulating PLN phosphorylation and thereby calcium reuptake.

Dysregulated micro-RNAs and long noncoding RNAs in cardiac development and pediatric heart failure.

Noncoding RNAs (ncRNAs) are broadly described as RNA molecules that are not translated into protein. The investigation of dysregulated ncRNAs in human diseases such as cancer, neurological, and cardiovascular diseases has been under way for well over a decade. Micro-RNAs and long noncoding RNAs (lncRNAs) are the best characterized ncRNAs. These ncRNAs can have profound effects on the regulation of gene expression during cardiac development and disease. Importantly, ncRNAs are significant regulators of gene expression in several congenital heart diseases and can positively or negatively impact cardiovascular development. In this review, we focus on literature involving micro-RNAs and lncRNAs in the context of pediatric cardiovascular diseases, preclinical models of heart failure, and cardiac development.

Circulating and Cardiac Tissue miRNAs in Children with Dilated Cardiomyopathy.

microRNAs (miRs) are small non-coding single-stranded RNAs that regulate gene expression. We previously evaluated expression of miRs in the cardiac tissue of children with dilated cardiomyopathy (DCM) using miRNA-seq. However, a comparative analysis of serum and cardiac miRs has not been performed in this population. The current study aimed to evaluate miR levels in the serum of pediatric DCM patients compared to healthy non-failing (NF) donor controls and investigate the association between miR levels in tissue and sera from the same pediatric DCM patients. Defining the relationship between serum and tissue miRs may allow the use of circulating miRs as surrogate markers of cardiac miRs. miR levels were investigated through miR-array in sera [n = 10 NF, n = 12 DCM] and miR-seq in tissue (n = 10 NF, n = 12 DCM). Pathway analysis was investigated using the miR enrichment analysis and annotation tool (miEAA) for the five miRs commonly dysregulated in the sera and tissue of pediatric DCM patients. Functional analysis of miRs commonly dysregulated in the sera and tissue of pediatric DCM patients suggests altered pathways related to cell growth, differentiation and proliferation, inflammation, mitochondrial function, and metabolism. These findings suggest that circulating miRs could reflect altered levels of cardiac tissue miRs.