Characterization of Circulating MicroRNA Expression in Patients with a Ventricular Septal Defect

Abstract

ObjectivesVentricular septal defect (VSD), one of the most common types of congenital heart disease (CHD), results from a combination of environmental and genetic factors. Recent studies demonstrated that microRNAs (miRNAs) are involved in development of CHD. This study was to characterize the expression of miRNAs that might be involved in the development or reflect the consequences of VSD.MethodsMiRNA microarray analysis and reverse transcription-polymerase chain reaction (RT-PCR) were employed to determine the miRNA expression profile from 3 patients with VSD and 3 VSD-free controls. 3 target gene databases were employed to predict the target genes of differentially expressed miRNAs. miRNAs that were generally consensus across the three databases were selected and then independently validated using real time PCR in plasma samples from 20 VSD patients and 15 VSD-free controls. Target genes of validated 8 miRNAs were predicted using bioinformatic methods.Results36 differentially expressed miRNAs were found in the patients with VSD and the VSD-free controls. Compared with VSD-free controls, expression of 15 miRNAs were up-regulated and 21 miRNAs were downregulated in the VSD group. 15 miRNAs were selected based on database analysis results and expression levels of 8 miRNAs were validated. The results of the real time PCR were consistent with those of the microarray analysis. Gene ontology analysis indicated that the top target genes were mainly related to cardiac right ventricle morphogenesis. NOTCH1, HAND1, ZFPM2, and GATA3 were predicted as targets of hsa-let-7e-5p, hsa-miR-222-3p and hsa-miR-433.ConclusionWe report for the first time the circulating miRNA profile for patients with VSD and showed that 7 miRNAs were downregulated and 1 upregulated when matched to VSD-free controls. Analysis revealed target genes involved in cardiac development were probably regulated by these miRNAs.

MicroRNAs: pleiotropic players in congenital heart disease and regeneration

“…Gene Ontology (GO) analysis revealed that target genes of these miRs include NOTCH1, HAND1, ZFPM2 (also known as transcription factor GATA4/FOG-2) and GATA3, genes known to be involved in cardiac right ventricle development and right ventricle morphogenesis (52)(53)(54). Dysregulation of these genes by miRs may therefore contribute to an altered cardiac morphogenesis that result in the development of VSDs (51). An overview of selected target genes and/or their miRs that are potentially involved in CHDs is provided in Table 1 and Figure 2.…”

“…However, miR-155 was shown to be down-regulated in patients with VSDs, indicating that VEGF levels were not increased in the cohort and thus did not activate their downstream miRs. This could be explained by the fact that the VSD cohort included in the study appeared to have relatively small shunt diameters (5-8 mm) (51,117), which in turn do not necessarily induce hypoxia. Therefore, miR-155 likely has a predominant impact on cardiac right ventricle development and right ventricle morphogenesis in patients with small VSDs rather than being involved in angiogenesis.…”

Circulating microRNAs as potential biomarkers for diagnosis of congenital heart defects

“…Lai et al [28] Serum samples of systemic right ventricular (n=5) versus serum samples of healthy controls (n=5) [29] Venous blood of 3 patients with VSD versus 3 VSDfree controls…”

“…Subsequent validation in 26 patients and 20 controls confirmed the up-regulation of 11 of the 23 microRNAs. [28] In the study by Li et al, [29] microRNA array initially was employed to determine the differential microRNAs between 3 patients with VSD and 3 VSD-free controls, which yielded a total of 36 candidate microRNAs, of which 15 were upregulated and 21 downregulated. The results of subsequent validation experiment confi rmed the up-regulation of miR-498 and down-regulation of miRlet-7e-5p, miR-155-5p, miR-222-3p, miR379-5p, miR-433, and miR-487b, and miR-409-3p in plasma samples of patients (n=20) compared with the controls (n=15).…”

“…The results of subsequent validation experiment confi rmed the up-regulation of miR-498 and down-regulation of miRlet-7e-5p, miR-155-5p, miR-222-3p, miR379-5p, miR-433, and miR-487b, and miR-409-3p in plasma samples of patients (n=20) compared with the controls (n=15). [29] By using microRNA array, de la Morena and colleagues [30] showed that 18 microRNAs were differentially expressed in peripheral blood of patients with CHD (associated with 22q11.2 deletion syndrome) relative to normal controls. The level of miR-185 in patient blood was found to be about 0.4 fold of normal level.…”