MiR487b is edited in the seed sequence in mice and humans, resulting in a novel, proangiogenic microRNA with a unique targetome. The rate of miR487b editing, as well as 2'-O-ribose-methylation, is increased in murine muscle tissue during postischemic neovascularization. Our findings suggest miR487b editing plays an intricate role in postischemic neovascularization.

Aims We have shown that 14q32 microRNAs are highly involved in vascular remodelling and cardiovascular disease. However, the 14q32 locus also encodes 41 ‘orphan’ small nucleolar RNAs (snoRNAs). We aimed to gather evidence for an independent role for 14q32 snoRNAs in human cardiovascular disease. Methods and results We performed a lookup of the 14q32 region within the dataset of a genome wide association scan in 5244 participants of the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER). Single nucleotide polymorphisms (SNPs) in the snoRNA-cluster were significantly associated with heart failure. These snoRNA-cluster SNPs were not linked to SNPs in the microRNA-cluster or in MEG3, indicating that snoRNAs modify the risk of cardiovascular disease independently. We looked at expression of 14q32 snoRNAs throughout the human cardio-vasculature. Expression profiles of the 14q32 snoRNAs appeared highly vessel specific. When we compared expression levels of 14q32 snoRNAs in human vena saphena magna (VSM) with those in failed VSM-coronary bypasses, we found that 14q32 snoRNAs were up-regulated. SNORD113.2, which showed a 17-fold up-regulation in failed bypasses, was also up-regulated two-fold in plasma samples drawn from patients with ST-elevation myocardial infarction directly after hospitalization compared with 30 days after start of treatment. However, fitting with the genomic associations, 14q32 snoRNA expression was highest in failing human hearts. In vitro studies show that the 14q32 snoRNAs bind predominantly to methyl-transferase Fibrillarin, indicating that they act through canonical mechanisms, but on non-canonical RNA targets. The canonical C/D-box snoRNA seed sequences were highly conserved between humans and mice. Conclusion 14q32 snoRNAs appear to play an independent role in cardiovascular pathology. 14q32 snoRNAs are specifically regulated throughout the human vasculature and their expression is up-regulated during cardiovascular disease. Our data demonstrate that snoRNAs merit increased effort and attention in future basic and clinical cardiovascular research.

Adenosine-to-inosine (A-to-I) editing in the seed sequence of microRNAs can shift the microRNAs’ targetomes and thus their function. Using public RNA-sequencing data, we identified 35 vasoactive microRNAs that are A-to-I edited. We quantified A-to-I editing of the primary (pri-)microRNAs in vascular fibroblasts and endothelial cells. Nine pri-microRNAs were indeed edited, and editing consistently increased under ischemia. We determined mature microRNA editing for the highest expressed microRNAs, i.e., miR-376a-3p, miR-376c-3p, miR-381-3p, and miR-411-5p. All four mature microRNAs were edited in their seed sequence. We show that both ADAR1 and ADAR2 (adenosine deaminase acting on RNA 1 and RNA 2) can edit pri-microRNAs in a microRNA-specific manner. MicroRNA editing also increased under ischemia in vivo in a murine hindlimb ischemia model and ex vivo in human veins. For each edited microRNA, we confirmed a shift in targetome. Expression of the edited microRNA targetomes, not the wild-type targetomes, was downregulated under ischemia in vivo . Furthermore, microRNA editing enhanced angiogenesis in vitro and ex vivo . In conclusion, we show that microRNA A-to-I editing is a widespread phenomenon, induced by ischemia. Each editing event results in a novel microRNA with a unique targetome, leading to increased angiogenesis.

In the last decade, the use of microRNA (miRNA) and extracellular vesicle (EV) therapies has emerged as an alternative approach to mitigate the negative effects of several disease pathologies ranging from cancer to tissue and organ regeneration; however, delivery approaches towards target tissues have not been optimized. To alleviate these challenges, including rapid diffusion upon injection and susceptibility to degradation, porcine-derived decellularized extracellular matrix (ECM) hydrogels are examined as a potential delivery platform for miRNA and EV therapeutics. The incorporation of EVs and miRNA antagonists, including anti-miR and antago-miR, in ECM hydrogels results in a prolonged release as compared to the biologic agents alone. In addition, individual in vitro assessments confirm the bioactivity of the therapeutics upon release from the ECM hydrogels. This work demonstrates the feasibility of encapsulating miRNA and EV therapeutics in ECM hydrogels to enhance delivery and potentially efficacy in later in vivo applications.