MicroRNA-421 Dysregulation is Associated with Tetralogy of Fallot

Abstract

The importance of microRNAs for maintaining stability in the developing vertebrate heart has recently become apparent. In addition, there is a growing appreciation for the significance of microRNAs in developmental pathology, including the formation of congenital heart defects. We examined the expression of microRNAs in right ventricular (RV) myocardium from infants with idiopathic tetralogy of Fallot (TOF, without a 22q11.2 deletion), and found 61 microRNAs to be significantly changed in expression in myocardium from children with TOF compared to normally developing comparison subjects (O'Brien et al. 2012). Predicted targets of microRNAs with altered expression were enriched for gene networks that regulate cardiac development. We previously derived a list of 229 genes known to be critical to heart development, and found 44 had significantly changed expression in TOF myocardium relative to normally developing myocardium. These 44 genes had significant negative correlations with 33 microRNAs, each of which also had significantly changed expression. Here, we focus on miR-421, as it is significantly upregulated in RV tissue from infants with TOF; is predicted to interact with multiple members of cardiovascular regulatory pathways; and has been shown to regulate cell proliferation. We knocked down, and over expressed miR-421 in primary cells derived from the RV of infants with TOF, and infants with normally developing hearts, respectively. We found a significant inverse correlation between the expression of miR-421 and SOX4, a key regulator of the Notch pathway, which has been shown to be important for the cardiac outflow track. These findings suggest that the dysregulation of miR-421 warrants further investigation as a potential contributor to tetralogy of Fallot.

Figure 1

MiR-421 expression vector.

Figure 2

MiR-421 is more highly expressed in RV from infants with TOF compared to the RV from normally developing infants. Data is from qRT-PCR. * significant difference (p = 0.03).

Figure 3

Putative connections between miR-421 and genes associated with cardiac development or function with significant inverse correlations between expression in TOF RV relative to control RV. Genes with a blue outline are members of the 5 key cardiac regulatory pathways shown in Figure 1 in [15].

Figure 4

Transfections: (A) overexpression (OE) miR-421 in normal cardiomyocytes. (B) siRNA knockdown (KD) of miR-421 in primary cell cultures derived from the RV of infants with TOF. * significant change relative to starting level.