Identification of cardiomyocyte-enriched long non-coding RNAs as potential targets for induction of cardiac regeneration

During myocardial infarction nearly one billion cardiomyocytes (CMs) die. In the neonatal mammalian hearts the lost CMs are renewed after injury, but this regenerative capacity is lost within a week after birth, and the adult mammalian heart is unable to regenerate significantly. Understanding of the factors regulating CM proliferation is limited. The aim of this work was to identify long non-coding RNAs (lncRNAs) that regulate CM proliferation and could therefore be targeted for induction of CM regeneration. Methods: RNA sequencing (RNAseq) was performed in cardiac tissue samples obtained from 1-, 4-, 9-and 23-day old mice (P1, P4, P9 and P23, respectively) and differential expression of lncRNAs was calculated with DESeq2 software. Cardiac enrichment was evaluated using Gene database by NCBI and Expression Atlas by EMBL-EBI. Ten cardiac-enriched lncRNAs with significant expression changes within the early postnatal period were chosen for validation with quantitative reverse transcription PCR (qRT-PCR). The relative lncRNA expression was calculated with the delta-delta-Ct method using Rn18s and Actb as the reference genes. To identify CM-specific lncRNAs, the expression of selected lncRNAs in neonatal mouse CMs and non-CMs was analyzed. Results: In total, 4 688 lncRNAs were detected in the postnatal hearts by RNAseq. Nearly 600 lncRNAs showed differential expression with more than 1.5-fold change between the time points P1, P4 and P9. Expression of 221 lncRNAs was increased and expression of 130 lncRNAs was decreased at P9 compared to P1. Nine of the ten selected lncRNAs were detected with qRT-PCR and eight of these exhibited similar expression patterns compared to RNAseq. Figure In addition, lncRNA-F was undetectable at P1, but showed significant increased expression at P9 and P23. Three of the selected lncRNAs were strongly enriched in CMs.