G protein-coupled receptors (GPCRs) play a central physiological role in the regulation of cardiac function in both health and disease and thus represents one of the largest class of surface receptors targeted by drugs. Several antagonists of GPCRs, such as β adrenergic receptors and angiotensin II receptors, are now considered standard of therapy for a wide range of cardiovascular disease such as hypertension, coronary artery disease, and heart failure. While the mechanism of action for GPCRs was thought to be largely worked-out in the 1980’s and 90’s, recent discoveries have brought to the fore new and previously unappreciated mechanisms for GPCR activation and subsequent downstream signaling. In this review we focus on GPCRs most relevant to the cardiovascular system and discuss traditional components of GPCR signaling and highlight evolving concepts of such as ligand bias, β-arrestin-mediated signaling, and conformational heterogeneity.

Percutaneous revascularization revolutionized the therapy of patients with coronary artery disease. Despite continuous technical advances that substantially improved patients' outcome after percutaneous revascularization, some issues are still open. In particular, restenosis still represents a challenge, even though it was dramatically reduced with the advent of drug-eluting stents. At the same time, drug-eluting stent thrombosis emerged as a major concern because of incomplete or delayed re-endothelialization after vascular injury. The discovery of microRNAs revealed a previously unknown layer of regulation for several biological processes, increasing our knowledge on the biological mechanisms underlying restenosis and stent thrombosis, revealing novel promising targets for more efficient and selective therapies. The present review summarizes recent experimental and clinical evidence on the role of microRNAs after arterial injury, focusing on practical aspects of their potential therapeutic application for selective inhibition of smooth muscle cell proliferation, enhancement of endothelial regeneration, and inhibition of platelet activation after coronary interventions. Application of circulating microRNAs as potential biomarkers is also discussed. (Circ Res.

BackgroundDiabetes mellitus (DM) has multifactorial detrimental effects on myocardial tissue. Recently, carbonic anhydrases (CAs) have been shown to play a major role in diabetic microangiopathy but their role in the diabetic cardiomyopathy is still unknown.Methods and ResultsWe obtained left ventricular samples from patients with DM type 2 (DM‐T2) and nondiabetic (NDM) patients with postinfarct heart failure who were undergoing surgical coronary revascularization. Myocardial levels of CA‐I and CA‐II were 6‐ and 11‐fold higher, respectively, in DM‐T2 versus NDM patients. Elevated CA‐I expression was mainly localized in the cardiac interstitium and endothelial cells. CA‐I induced by high glucose levels hampers endothelial cell permeability and determines endothelial cell apoptosis in vitro. Accordingly, capillary density was significantly lower in the DM‐T2 myocardial samples (mean±SE=2152±146 versus 4545±211/mm2). On the other hand, CA‐II was mainly upregulated in cardiomyocytes. The latter was associated with sodium‐hydrogen exchanger‐1 hyperphosphorylation, exaggerated myocyte hypertrophy (cross‐sectional area 565±34 versus 412±27 μm2), and apoptotic death (830±54 versus 470±34 per 106 myocytes) in DM‐T2 versus NDM patients. CA‐II is activated by high glucose levels and directly induces cardiomyocyte hypertrophy and death in vitro, which are prevented by sodium‐hydrogen exchanger‐1 inhibition. CA‐II was shown to be a direct target for repression by microRNA‐23b, which was downregulated in myocardial samples from DM‐T2 patients. MicroRNA‐23b is regulated by p38 mitogen‐activated protein kinase, and it modulates high‐glucose CA‐II–dependent effects on cardiomyocyte survival in vitro.ConclusionsMyocardial CA activation is significantly elevated in human diabetic ischemic cardiomyopathy. These data may open new avenues for targeted treatment of diabetic heart failure.

The present findings suggest that circulating levels of miRNAs are differentially expressed in patients with HF of different aetiologies. The presence of a transcoronary concentration gradient suggests a selective release of miRNAs by the failing heart into the coronary circulation. The presence of aetiology-specific transcoronary concentration gradients in HF patients might provide important information to better understand their role in HF, and suggests they could be useful biomarkers to distinguish HF of different aetiologies.

We identify miR-23b as a novel regulator of VSMC phenotypic switch in vitro and following vascular injury in vivo.

Large-scale analyses of mammalian transcriptomes have identified a significant number of different RNA molecules that are not translated into protein. In fact, the use of new sequencing technologies has identified that most of the genome is transcribed, producing a heterogeneous population of RNAs which do not encode for proteins (ncRNAs). Emerging data suggest that these transcripts influence the development of cardiovascular disease. The best characterized non-coding RNA family is represented by short highly conserved RNA molecules, termed microRNAs (miRNAs), which mediate a process of mRNA silencing through transcript degradation or translational repression. These microRNAs (miRNAs) are expressed in cardiovascular tissues and play key roles in many cardiovascular pathologies, such as coronary artery disease (CAD) and heart failure (HF). Potential links between other ncRNAs, like long non-coding RNA, and cardiovascular disease are intriguing but the functions of these transcripts are largely unknown. Thus, the functional characterization of ncRNAs is essential to improve the overall understanding of cellular processes involved in cardiovascular diseases in order to define new therapeutic strategies. This review outlines the current knowledge of the different ncRNA classes and summarizes their role in cardiovascular development and disease.

Ligand activation of the angiotensin II type 1 receptor (AT1R), a member of the G protein-coupled receptor (GPCR) family, stimulates intracellular signaling to mediate a variety of physiological responses. The AT1R is also known to be a mechanical sensor. When activated by mechanical stretch, the AT1R can signal via the multifunctional adaptor protein β-arrestin, rather than through classical heterotrimeric G protein pathways. To date, the AT1R conformation induced by membrane stretch in the absence of ligand was thought to be the same as that induced by β-arrestin-biased agonists, which selectively engage β-arrestin thereby preventing G protein coupling. Here, we show that in contrast to the β-arrestin-biased agonists TRV120023 and TRV120026, membrane stretch uniquely promotes the coupling of the inhibitory G protein (Gαi) to the AT1R to transduce signaling. Stretch-triggered AT1R-Gαi coupling is required for the recruitment of β-arrestin2 and activation of downstream signaling pathways, such as EGFR transactivation and ERK phosphorylation. Our findings demonstrate additional complexity in the mechanism of receptor bias in which the recruitment of Gαi is required for allosteric mechanoactivation of the AT1R-induced β-arrestin-biased signaling.

Peripheral ischemia is associated with higher degree of endothelial dysfunction and a worse prognosis after percutaneous coronary interventions (PCI). However, the role of peripheral ischemia on vascular remodeling in remote districts remains poorly understood. Here we show that the presence of hindlimb ischemia significantly enhances neointima formation and impairs endothelial recovery in balloon-injured carotid arteries. Endothelial-derived microRNAs are involved in the modulation of these processes. Indeed, endothelial miR-16 is remarkably upregulated after vascular injury in the presences of hindlimb ischemia and exerts a negative effect on endothelial repair through the inhibition of RhoGDIα and nitric oxide (NO) production. We showed that the repression of RhoGDIα by means of miR-16 induces RhoA, with consequent reduction of NO bioavailability. Thus, hindlimb ischemia affects negative carotid remodeling increasing neointima formation after injury, while systemic antagonizzation of miR-16 is able to prevent these negative effects.Peripheral Arterial Disease (PAD) affects approximately 20% of adults over the age of 50, exposing them to the negative impact of peripheral ischemia 1-3 . However, half of these subjects are asymptomatic. Hence, the actual impact of peripheral ischemia is largely under-estimated, leading to under-treatment and seriously undermining prevention strategies 4,5 . Interestingly, in patients with multidistrectual atherosclerosis the presence of peripheral ischemia is associated to a further increase in mortality independently of classical cardiovascular risk factors 6,7 . In line with these observations, it was shown that limb ischemia might remotely influence atherosclerotic vascular remodeling in coronary and cerebral districts 8,9 . Indeed, the presence of limb ischemia is often associated to endothelial dysfunction 10 , and consequently to a more severe atherosclerosis of remote districts 11 . Interestingly, patients with PAD and peripheral ischemia undergoing percutaneous coronary intervention (PCI) have a significantly poorer prognosis suggesting a remote negative impact of peripheral ischemia on coronary artery disease 12,13 . This picture has not improved with the recent significant progresses in medical therapy and introduction of the latest generation of drug eluting stents (DES) 14 . Indeed, it is known that limb ischemia can negatively influence vascular remodeling in other districts, such as coronary and cerebral arteries [12][13][14][15] . MicroRNA (miRNAs), small noncoding RNAs, have emerged in the last decade as epigenetic regulators of essential biological processes [15][16][17] . In particular, they play a key role in the pathophysiology of atherosclerosis, including restenosis after PCI [18][19][20] , and could be used as circulating biomarkers [21][22][23][24] . Recent studies showed that miRNAs could modulate both vascular smooth muscle cells (VSMC) and the endothelium in response to vascular injury 25 . Moreover, miRNAs can mediate intercellular crosstalk, involv...