Silencing of circHIPK3 Inhibits Pressure Overload-Induced Cardiac Hypertrophy and Dysfunction by Sponging miR-185-3p

Background: Cardiac hypertrophy is induced by diverse patho-physiological stimuli and indicates an increase in cardiomyocyte size. Circular RNAs (circRNAs) and microRNAs (miRNAs), members of noncoding RNAs, are involved in several biological processes and cardiovascular diseases (CVD). Here, we investigated the potential role of circHIPK3, which is produced by the third exon of the HIPK3 gene in cardiac hypertrophy.

Methods: qRT-PCR and Sanger sequencing were conducted to identify the expression and characteristics (head-to-tail structure, stability, and location) of circHIPK3 in cardiac hypertrophy; Immunostaining of α-SMA was performed to evaluate the size of the cardiomyocytes; Transverse aortic constriction (TAC) induced hypertrophy models of mice were established to investigate the effect of circHIPK3 in vivo. Bioinformatics analysis and luciferase reporter assays, RNA immunoprecipitation, and fluorescence in situ hybridization (FISH) experiments were conducted to investigate the mechanism of circHIPK3-mediated cardiac hypertrophy.

Results: circHIPK3 is circular, more stable, and mainly located in the cytoplasm. Silencing of circHIPK3 inhibited the TAC induced cardiac hypertrophy, and reversed the effect of TAC on the echocardiograph parameters, such as left ventricular end-diastolic pressure (LVEDPS), left ventricular fraction shortening (LVFS), left ventricular ejection fraction (LVEF), and left ventricular systolic dysfunction (LVSD), and also the heart weight to tibial length (HW/TL). Angiotensin II (Ang II) Ang II-treated cardiomyocytes showed larger size of cardiomyocyte and upregulation of fetal genes, biomarkers of cardiac hypertrophy, peptide hormones, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), and myofilament protein, β-myosin heavy chain (β-MHC). These effects were reversed by circHIPK3 knockdown. Mechanically, circHIPK3 sponges miR-185-3p. In addition, miR-185-3p targets CASR. The rescue experiments confirmed the interaction between circHIPK3 and miR-185-3p as well as miR-185-3p and CASR.

Discussion: Our data suggested that circHIPK3 serve as a miR-185-3p sponge to regulate cardiac hypertrophy revealing a potential new target for the prevention of TAC- and Ang-II induced cardiac hypertrophy.