Role of Circular RNAs in Cardiovascular Disease.

Circular RNAs (circRNAs) have recently been identified as a new class of long noncoding RNAs with gene regulatory roles. These covalently closed transcripts are generated when the pre-mRNA splicing machinery back splices to join a downstream 5' splice site to an upstream 3' splice site. CircRNAs are naturally resistant to degradation by exonucleases and have long half-lives compared with their linear counterpart that potentially could serve as biomarkers for disease. Recent evidence highlights that circRNAs may play an essential role in cardiovascular injury and repair. However, our knowledge of circRNA is still in its infancy with limited direct evidence to suggest that circRNA may play critical roles in the mechanism and treatment of cardiac dysfunction. In this review, we focus on our current understanding of circRNA in the cardiovascular system.

Interleukin-10 Inhibits Bone Marrow Fibroblast Progenitor Cell-Mediated Cardiac Fibrosis in Pressure-Overloaded Myocardium.

BACKGROUND: Activated fibroblasts (myofibroblasts) play a critical role in cardiac fibrosis; however, their origin in the diseased heart remains unclear, warranting further investigation. Recent studies suggest the contribution of bone marrow fibroblast progenitor cells (BM-FPCs) in pressure overload-induced cardiac fibrosis. We have previously shown that interleukin-10 (IL10) suppresses pressure overload-induced cardiac fibrosis; however, the role of IL10 in inhibition of BM-FPC-mediated cardiac fibrosis is not known. We hypothesized that IL10 inhibits pressure overload-induced homing of BM-FPCs to the heart and their transdifferentiation to myofibroblasts and thus attenuates cardiac fibrosis. METHODS: Pressure overload was induced in wild-type (WT) and IL10 knockout (IL10KO) mice by transverse aortic constriction. To determine the bone marrow origin, chimeric mice were created with enhanced green fluorescent protein WT mice marrow to the IL10KO mice. For mechanistic studies, FPCs were isolated from mouse bone marrow. RESULTS: Pressure overload enhanced BM-FPC mobilization and homing in IL10KO mice compared with WT mice. Furthermore, WT bone marrow (from enhanced green fluorescent protein mice) transplantation in bone marrow-depleted IL10KO mice (IL10KO chimeric mice) reduced transverse aortic constriction-induced BM-FPC mobilization compared with IL10KO mice. Green fluorescent protein costaining with α-smooth muscle actin or collagen 1α in left ventricular tissue sections of IL10KO chimeric mice suggests that myofibroblasts were derived from bone marrow after transverse aortic constriction. Finally, WT bone marrow transplantation in IL10KO mice inhibited transverse aortic constriction-induced cardiac fibrosis and improved heart function. At the molecular level, IL10 treatment significantly inhibited transforming growth factor-ß-induced transdifferentiation and fibrotic signaling in WT BM-FPCs in vitro. Furthermore, fibrosis-associated microRNA (miRNA) expression was highly upregulated in IL10KO-FPCs compared with WT-FPCs. Polymerase chain reaction-based selective miRNA analysis revealed that transforming growth factor-ß-induced enhanced expression of fibrosis-associated miRNAs (miRNA-21, -145, and -208) was significantly inhibited by IL10. Restoration of miRNA-21 levels suppressed the IL10 effects on transforming growth factor-ß-induced fibrotic signaling in BM-FPCs. CONCLUSIONS: Our findings suggest that IL10 inhibits BM-FPC homing and transdifferentiation to myofibroblasts in pressure-overloaded myocardium. Mechanistically, we show for the first time that IL10 suppresses Smad-miRNA-21-mediated activation of BM-FPCs and thus modulates cardiac fibrosis.

Therapeutic inhibition of miR-375 attenuates post-myocardial infarction inflammatory response and left ventricular dysfunction via PDK-1-AKT signalling axis.

AIMS: Increased miR-375 levels has been implicated in rodent models of myocardial infarction (MI) and with patients with heart failure. However, no prior study had established a therapeutic role of miR-375 in ischemic myocardium. Therefore, we assessed whether inhibition of MI-induced miR-375 by LNA anti-miR-375 can improve recovery after acute MI. METHODS AND RESULTS: Ten weeks old mice were treated with either control or LNA anti miR-375 after induction of MI by LAD ligation. The inflammatory response, cardiomyocyte apoptosis, capillary density and left ventricular (LV) functional, and structural remodelling changes were evaluated. Anti-miR-375 therapy significantly decreased inflammatory response and reduced cardiomyocyte apoptosis in the ischemic myocardium and significantly improved LV function and neovascularization and reduced infarct size. Repression of miR-375 led to the activation of 3-phosphoinositide-dependent protein kinase 1 (PDK-1) and increased AKT phosphorylation on Thr-308 in experimental hearts. In corroboration with our in vivo findings, our in vitro studies demonstrated that knockdown of miR-375 in macrophages modulated their phenotype, enhanced PDK-1 levels, and reduced pro-inflammatory cytokines expression following LPS challenge. Further, miR-375 levels were elevated in failing human heart tissue. CONCLUSION: Taken together, our studies demonstrate that anti-miR-375 therapy reduced inflammatory response, decreased cardiomyocyte death, improved LV function, and enhanced angiogenesis by targeting multiple cell types mediated at least in part through PDK-1/AKT signalling mechanisms.

Endothelial Progenitor Cells: Procedure for Cell Isolation and Applications.

Bone marrow endothelial progenitor cells (EPCs) have shown a great promise to promote ischemic tissue neovascularization and to attenuate ischemic injury in a variety of animal models, which led to EPC-based clinical trials that yielded modest but promising results. Some of the variables in the use of EPCs relate to their differential isolation and characterization protocols since the EPC literature does not identify a unique marker for these vascular progenitors. In this chapter, we present step-by-step protocols for the isolation of EPCs, their characterization and culture conditions, and their potential use in basic and clinical research.