Human left ventricle circRNA-miRNA-mRNA network analyses reveals a novel proangiogenic role for circNPHP1 under ischemic conditions

ABSTRACT Background Ischemic heart disease (IHD) is characterized by insufficient myocardial blood flow in the left ventricle and aggravated by diabetes mellitus. Endothelial resilience and reparative angiogenesis are tightly controlled processes. Gene expression is regulated by multimodal interactions between RNA species. Circular RNAs (circRNAs) can sponge microRNAs (miRNAs) to reduce the repressive effects of miRNAs on its messenger RNA (mRNAs) targets. Methods Left ventricle whole RNA-sequencing (circRNAs, mRNAs) and small RNA- sequencing (miRNAs) datasets were obtained from 3 patient groups: IHD with/out T2DM and controls (N=11 to 12/group) as part of a prospective observational cardiac surgery study. The interactions between differentially expressed (DE) circRNAs, miRNA and mRNAs were identified with a customized bioinformatics pipeline. The emerging networks were screened using endothelial-specific RNA-sequencing datasets from GEO resulting in EC-rich networks. CircRNAs from these networks were subsequently screened (RT-PCR) in endothelial cells (ECs) exposed to disease-mimicking conditions vs control. Afterwards, circRNA pulldown allowed to interrogate the circRNA-miRNAs interactome in ECs. EC biology assays using loss-of-function and gain-of-function approaches corroborated the study. Results We identified novel circRNA-miRNA-mRNA interactions in the human diseased heart. CircNPHP1, which is upregulated in IHD with/out Type-2 diabetes mellitus (T2DM), sponges miR-221-3p to de-repress VEGF-A and BCL2, increasing the angiogenic capacity of ECs under disease and disease-mimicking conditions. Conclusions The interactions between individual members of different RNA species are affected by IHD. The therapeutic value of circNPHP1/miR-221-3p axis could be further investigated. Visual Abstract Highlights We found a novel circRNA-miRNA-mRNA network in IHD and Type 2 diabetes. CircNPHP1 regulates angiogenesis and proliferation in the cardiac ECs exposed to conditions mimicking IHD and Type 2 diabetes. We elucidated a novel pro-angiogenic subnetwork commanded by circNPHP1/miR-221-3p/BCL2/VEGFA. We identified circNPHP1 as a potential new target for therapeutic angiogenesis.