ABSTRACT
BACKGROUND: Myocardial infarction leads to cardiac remodeling and development of heart failure. Insufficient myocardial capillary density after myocardial infarction has been identified as a critical event in this process, although the underlying mechanisms of cardiac angiogenesis are mechanistically not well understood. METHODS AND RESULTS: Here, we show that the small noncoding RNA microRNA-24 (miR-24) is enriched in cardiac endothelial cells and considerably upregulated after cardiac ischemia. MiR-24 induces endothelial cell apoptosis, abolishes endothelial capillary network formation on Matrigel, and inhibits cell sprouting from endothelial spheroids. These effects are mediated through targeting of the endothelium-enriched transcription factor GATA2 and the p21-activated kinase PAK4, which were identified by bioinformatic predictions and validated by luciferase gene reporter assays. Respective downstream signaling cascades involving phosphorylated BAD (Bcl-XL/Bcl-2-associated death promoter) and Sirtuin1 were identified by transcriptome, protein arrays, and chromatin immunoprecipitation analyses. Overexpression of miR-24 or silencing of its targets significantly impaired angiogenesis in zebrafish embryos. Blocking of endothelial miR-24 limited myocardial infarct size of mice via prevention of endothelial apoptosis and enhancement of vascularity, which led to preserved cardiac function and survival. CONCLUSIONS: Our findings indicate that miR-24 acts as a critical regulator of endothelial cell apoptosis and angiogenesis and is suitable for therapeutic intervention in the setting of ischemic heart disease.
Subject(s)
Endothelial Cells/metabolism , MicroRNAs/physiology , Myocardial Infarction/physiopathology , Animals , Apoptosis/drug effects , Arterioles/pathology , Capillaries/pathology , Cell Hypoxia , Cells, Cultured/drug effects , Cells, Cultured/metabolism , Collagen , Drug Combinations , Drug Evaluation, Preclinical , Endothelial Cells/pathology , GATA2 Transcription Factor/biosynthesis , GATA2 Transcription Factor/genetics , Gene Expression Profiling , Heart Failure/etiology , Heme Oxygenase-1/biosynthesis , Heme Oxygenase-1/genetics , Laminin , Male , Mice , Mice, Inbred C57BL , MicroRNAs/antagonists & inhibitors , MicroRNAs/genetics , Myocardial Infarction/complications , Myocardial Infarction/genetics , Neovascularization, Physiologic/drug effects , Neovascularization, Physiologic/genetics , Oligoribonucleotides/pharmacology , Proteoglycans , RNA Interference , RNA, Small Interfering/pharmacology , RNA, Small Interfering/therapeutic use , Spheroids, Cellular , Ventricular Remodeling , Zebrafish/embryology , Zebrafish Proteins/biosynthesis , Zebrafish Proteins/genetics , p21-Activated Kinases/biosynthesis , p21-Activated Kinases/geneticsABSTRACT
T cell sensitivity to antigen is intrinsically regulated during maturation to ensure proper development of immunity and tolerance, but how this is accomplished remains elusive. Here we show that increasing miR-181a expression in mature T cells augments the sensitivity to peptide antigens, while inhibiting miR-181a expression in the immature T cells reduces sensitivity and impairs both positive and negative selection. Moreover, quantitative regulation of T cell sensitivity by miR-181a enables mature T cells to recognize antagonists-the inhibitory peptide antigens-as agonists. These effects are in part achieved by the downregulation of multiple phosphatases, which leads to elevated steady-state levels of phosphorylated intermediates and a reduction of the T cell receptor signaling threshold. Importantly, higher miR-181a expression correlates with greater T cell sensitivity in immature T cells, suggesting that miR-181a acts as an intrinsic antigen sensitivity "rheostat" during T cell development.
