miR-96 and miR-183 differentially regulate neonatal and adult postinfarct neovascularization
Raphael F.P. Castellan, Milena Vitiello, Martina Vidmar, Steven Johnstone, Dominga Iacobazzi, David Mellis, Benjamin Cathcart, Adrian Thomson, Christiana Ruhrberg, Massimo Caputo, David E. Newby, Gillian A. Gray, Andrew H. Baker, Andrea Caporali, Marco Meloni
Raphael F.P. Castellan, Milena Vitiello, Martina Vidmar, Steven Johnstone, Dominga Iacobazzi, David Mellis, Benjamin Cathcart, Adrian Thomson, Christiana Ruhrberg, Massimo Caputo, David E. Newby, Gillian A. Gray, Andrew H. Baker, Andrea Caporali, Marco Meloni
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Research Article Angiogenesis Vascular biology

miR-96 and miR-183 differentially regulate neonatal and adult postinfarct neovascularization

Abstract

Following myocardial infarction (MI), the adult heart has minimal regenerative potential. Conversely, the neonatal heart can undergo extensive regeneration, and neovascularization capacity was hypothesized to contribute to this difference. Here, we demonstrate the higher angiogenic potential of neonatal compared with adult mouse cardiac endothelial cells (MCECs) in vitro and use this difference to identify candidate microRNAs (miRs) regulating cardiac angiogenesis after MI. miR expression profiling revealed miR-96 and miR-183 upregulation in adult compared with neonatal MCECs. Their overexpression decreased the angiogenic potential of neonatal MCECs in vitro and prevented scar resolution and neovascularization in neonatal mice after MI. Inversely, their inhibition improved the angiogenic potential of adult MCECs, and miR-96/miR-183–KO mice had increased peri-infarct neovascularization. In silico analyses identified anillin (ANLN) as a direct target of miR-96 and miR-183. In agreement, Anln expression declined following their overexpression and increased after their inhibition in vitro. Moreover, ANLN expression inversely correlated with miR-96 expression and age in cardiac ECs of cardiovascular patients. In vivo, ANLN+ vessels were enriched in the peri-infarct area of miR-96/miR-183–KO mice. These findings identify miR-96 and miR-183 as regulators of neovascularization following MI and miR-regulated genes, such as anillin, as potential therapeutic targets for cardiovascular disease.

Authors

Raphael F.P. Castellan, Milena Vitiello, Martina Vidmar, Steven Johnstone, Dominga Iacobazzi, David Mellis, Benjamin Cathcart, Adrian Thomson, Christiana Ruhrberg, Massimo Caputo, David E. Newby, Gillian A. Gray, Andrew H. Baker, Andrea Caporali, Marco Meloni

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Figure 3

Mimic-mediated overexpression of miR-96 and miR-183 prevents scar tissue resolution and neovascularization of the neonatal mouse heart after MI.

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Mimic-mediated overexpression of miR-96 and miR-183 prevents scar tissue...
(A and B) Survival curves show that, whereas all neonatal mice injected with a combination of miR-96 and miR-183 mimics at the dose of 200 pmol died within 24 hours after MI and mimic injection (A), the mortality rate of mice injected with miR-96 and miR-183 mimics at the dose of 50 pmol was similar to that of control mimic-injected mice (B). Representative microphotographs (C) and bar graph (D) (n = 6 control mimic, n = 8 miR-96/miR-183 mimic) showing differences in fibrosis deposition (in red, assessed by Picrosirius red staining) between neonatal infarcted hearts injected with either control mimics or miR-96 and miR-183 mimics. Scale bar: 1000 μm. Representative microphotographs (E) and bar graph (F) (n = 4 control mimic, n = 6 miR-96/miR-183 mimic) showing the reduced capillary density in the heart of neonatal mice subjected to MI and injected with miR-96 and miR-183 mimics compared with control injected mice. Capillaries are stained with isolectin-B4 (green fluorescence), and arterioles are stained with isolectin-B4 (green fluorescence) and α-smooth muscle actin (red fluorescence). Scale bar: 250 μm (left); 50 μm (right). *P < 0.05 vs. control mimic (Student’s t test). Analyses were performed at 21 days after MI. Data are shown as mean ± SEM.