Urotensin-II (UII) is a vasoactive peptide that promotes vascular smooth muscle cells (VSMCs) proliferation and is involved in the pathogenesis of atherosclerosis, restenosis, and vascular remodelling. This study aimed to determine the role of calcium (Ca²⁺)-dependent signalling and alternative signalling pathways in UII-evoked VSMCs proliferation focusing on store-operated Ca²⁺ entry (SOCE) and epithelium growth factor receptor (EGFR) transactivation. We used primary cultures of VSMCs isolated from Wistar rat aorta to investigate the effects of UII on intracellular Ca²⁺ mobilization, and proliferation determined by the 5-bromo-2-deoxyuridine (BrdU) assay. We found that UII enhanced intracellular Ca²⁺ concentration ([Ca²⁺]_i) which was significantly reduced by classical SOCE inhibitors and by knockdown of essential components of the SOCE such as stromal interaction molecule 1 (STIM1), Orai1, or TRPC1. Moreover, UII activated a Gd³⁺-sensitive current with similar features of the Ca²⁺ release-activated Ca²⁺ current (I_CRAC). Additionally, UII stimulated VSMCs proliferation and Ca²⁺/cAMP response element-binding protein (CREB) activation through the SOCE pathway that involved STIM1, Orai1, and TRPC1. Co-immunoprecipitation experiments showed that UII promoted the association between Orai1 and STIM1, and between Orai1 and TRPC1. Moreover, we determined that EGFR transactivation, extracellular signal-regulated kinase (ERK) and Ca²⁺/calmodulin-dependent kinase (CaMK) signalling pathways were involved in both UII-mediated Ca²⁺ influx, CREB activation and VSMCs proliferation. Our data show for the first time that UII-induced VSMCs proliferation and CREB activation requires a complex signalling pathway that involves on the one hand SOCE mediated by STIM1, Orai1, and TRPC1, and on the other hand EGFR, ERK, and CaMK activation.

Urocortin 1 and 2 (Ucn-1 and Ucn-2) have established protective actions against myocardial ischemia-reperfusion (I/R) injuries. However, little is known about their role in posttranscriptional regulation in the process of cardioprotection. Herein, we investigated whether microRNAs play a role in urocortin-induced cardioprotection. Administration of Ucn-1 and Ucn-2 at the beginning of reperfusion significantly restored cardiac function, as evidenced ex vivo in Langendorff-perfused rat hearts and in vivo in rat subjected to I/R. Experiments using microarray and qRT-PCR determined that the addition of Ucn-1 at reperfusion modulated the expression of several miRNAs with unknown role in cardiac protection. Ucn-1 enhanced the expression of miR-125a-3p, miR-324-3p; meanwhile it decreased miR-139-3p. Similarly, intravenous infusion of Ucn-2 in rat model of I/R mimicked the effect of Ucn-1 on miR-324-3p and miR-139-3p. The effect of Ucn-1 involves the activation of corticotropin-releasing factor receptor-2, Epac2 and ERK1/2. Moreover, the overexpression of miR-125a-3p, miR-324-3p and miR-139-3p promoted dysregulation of genes expression involved in cell death and apoptosis (BRCA1, BIM, STAT2), in cAMP and Ca²⁺ signaling (PDE4a, CASQ1), in cell stress (NFAT5, XBP1, MAP3K12) and in metabolism (CPT2, FoxO1, MTRF1, TAZ). Altogether, these data unveil a novel role of urocortin in myocardial protection, involving posttranscriptional regulation with miRNAs.

Aims The aim of this study is to evaluate the positive inotropic effect of urocortin (Ucn) and to characterize its signalling pathways.

Background— Hypertension, a major cause of cardiovascular morbidity and mortality, can result from chronic hypoxia; however, the pathogenesis of this disorder is unknown. We hypothesized that downregulation of the maxi-K ⁺ channel β ₁ -subunit by hypoxia decreases the ability of these channels to hyperpolarize arterial smooth muscle cells, thus favoring vasoconstriction and hypertension. Methods and Results— Lowering O ₂ tension produced a decrease of maxi-K ⁺ β ₁ -subunit mRNA levels in rat (aortic and basilar) and human (mammary) arterial myocytes. This was paralleled by a reduction of the β ₁ -subunit protein level as determined by immunocytochemistry and flow cytometry. Exposure to hypoxia also produced a decrease of open probability, mean open time, and sensitivity to the xenoestrogen tamoxifen of single maxi-K ⁺ channels recorded from patch-clamped dispersed myocytes. The number of channels per patch and the single-channel conductance were not altered. The vasorelaxing force of maxi-K ⁺ channels was diminished in rat and human arterial rings exposed to low oxygen tension. Conclusions— These results indicate that a decrease of the maxi-K ⁺ channel β ₁ -subunit expression in arterial myocytes is a key factor in the vasomotor alterations induced by hypoxia.

Objective—: Human urotensin-II (UII) is considered the most potentendogenous vasoconstrictor discovered to date, although the precise mechanism activated downstream of its receptor UTS2R in blood vessels remains elusive. The aim of this study was to determine the role of the store operated Ca²⁺entry (SOCE) signaling pathway in UII-induced coronary artery vasoconstriction.Methods and Results—: We used a combination of isometric tension measurement, Ca²⁺imaging, pharmacology, and molecular approaches to study UII-mediated rat coronary artery vasoconstriction and intracellular Ca²⁺mobilization in coronary smooth muscle cells. We found that UII promoted dose-dependent vasoconstriction and elicited Ca²⁺and Mn²⁺influx, which were sensitive to classical SOCE inhibitors. In addition, knockdown of either STIM1 or Orai1 essentially inhibited UII-mediated SOCE and prevented UII but not high-KCL evoked contraction in transfected coronary artery. Moreover, we found that Ca²⁺-independent phospholipase A₂β was involved in UII effects and that is colocalized with STIM1 in different submembrane compartments. Importantly, STIM1 but not Orai1 downregulation inhibits significantly independent phospholipase A₂activation. Furthermore, lysophosphatidylcholine, an independent phospholipase A₂product, activated Orai1 but not STIM1-dependent contraction and SOCE.Conclusion—: Here, we demonstrated that different critical players of SOCE signaling pathway are required for UII-induced vasoconstriction of rat coronary artery.