Neuronatin Promotes Neural Lineage in ESCs via Ca2+ Signaling

Neural induction is the first step in the formation of the vertebrate central nervous system. The emerging consensus of the mechanisms underling neural induction is the combined influences from inhibiting bone morphogenetic protein (BMP) signaling and activating fibroblast growth factor (FGF)/Erk signaling, which act extrinsically via either autocrine or paracrine fashions. However, do intrinsic forces (cues) exist and do they play decisive roles in neural induction? These questions remain to be answered. Here, we have identified a novel neural initiator, neuronatin (Nnat), which acts as an intrinsic factor to promote neural fate in mammals and Xenopus. ESCs lacking this intrinsic factor fail to undergo neural induction despite the inhibition of the BMP pathway. We show that Nnat initiates neural induction in ESCs through increasing intracellular Ca2+ ([Ca2+]i) by antagonizing Ca2+-ATPase isoform 2 (sarco/endoplasmic reticulum Ca2+-ATPase isoform 2) in the endoplasmic reticulum, which in turn increases the phosphorylation of Erk1/2 and inhibits the BMP4 pathway and leads to neural induction in conjunction with FGF/Erk pathway. STEM CELLS 2010;28:1950–196

Role of SRC-Family Kinases in Hypoxic Vasoconstriction of Rat Pulmonary Artery

Aims: We investigated the role of src-family kinases (srcFKs) in hypoxic pulmonary vasoconstriction (HPV) and how this relates to Rho-kinase-mediated Ca(2+) sensitization and changes in intracellular Ca(2+) concentration ([Ca(2+)](i)). Methods and results: Intra-pulmonary arteries (IPAs) were obtained from male Wistar rats. HPV was induced in myograph-mounted IPAs. Auto-phosphorylation of srcFKs and phosphorylation of the regulatory subunit of myosin phosphatase (MYPT-1) and myosin light-chain (MLC(20)) in response to hypoxia were determined by western blotting. Translocation of Rho-kinase and effects of siRNA knockdown of src and fyn were examined in cultured pulmonary artery smooth muscle cells (PASMCs). [Ca(2+)](i) was estimated in Fura-PE3-loaded IPA. HPV was inhibited by two blockers of srcFKs, SU6656 and PP2. Hypoxia enhanced phosphorylation of three srcFK proteins at Tyr-416 (60, 59, and 54 kDa, corresponding to src, fyn, and yes, respectively) and enhanced srcFK-dependent tyrosine phosphorylation of multiple target proteins. Hypoxia caused a complex, time-dependent enhancement of MYPT-1 and MLC(20) phosphorylation, both in the absence and presence of pre-constriction. The sustained component of this enhancement was blocked by SU6656 and the Rho-kinase inhibitor Y27632. In PASMCs, hypoxia caused translocation of Rho-kinase from the nucleus to the cytoplasm, and this was prevented by anti-src siRNA and to a lesser extent by anti-fyn siRNA. The biphasic increases in [Ca(2+)](i) that accompany HPV were also inhibited by PP2. Conclusion: Hypoxia activates srcFKs and triggers protein tyrosine phosphorylation in IPA. Hypoxia-mediated Rho-kinase activation, Ca(2+) sensitization, and [Ca(2+)](i) responses are depressed by srcFK inhibitors and/or siRNA knockdown, suggesting a central role of srcFKs in HPV

2017 in Review, Perspectives for 2018

The articles in this edition focuses on 1) the efforts of the Kremlin to ensure a smooth run-up to the 2018 Russian Presidential election, noting that its major challenges still lie ahead; 2) Russia's slow pace of economic recovery in 2017, suggesting that the characterisation of the Russian economy as stagnant may not hold for some areas of the economy; and 3) how a key aim of Russian foreign policy in 2017 was to assert its role in several international crises, and that whilst this short-term aim has more or less been achieved, the success of Moscow’s long-term strategy to develop a broad-range of global relationships remains uncertain.ISSN:1863-042

2016 in Review, Looking Forward to 2017

ISSN:1863-042

Glabridin-induced vasorelaxation: Evidence for a role of BK_Ca channels and cyclic GMP

AbstractBackground and purposeGlabridin is a major flavonoid in Glycyrrhiza glabra (licorice) root, a traditional Asian medicine. Glabridin is reported to have anti-atherogenic, anti-inflammatory and anti-nephritic properties; however its effects on vascular tone remain unexplored.Experimental approachWe examined the effect of glabridin on rat main mesenteric artery using isometric myography and also ELISA to measure cGMP levels.Key resultsGlabridin (30μM) relaxed arteries pre-constricted with the thromboxane A2 analog U46619 (0.2μM) by ~60% in an endothelium-independent manner. Relaxation to 30μM glabridin was abolished by the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (1μM) and by the BKCa channel blocker tetraethyammonium (1mM) but was unaffected by the estrogen receptor antagonist ICI182780. The concentration-response curve to glabridin (0.1 to 30μM) was downshifted by the KATP channel blocker glibenclamide (10μM), the KV channel blocker 4-aminopyridine (300μM), and the KIR blocker BaCl2 (30μM). In U46619-contracted arteries partially relaxed by 0.1μM sodium nitroprusside, application of 10 and 30nM glabridin caused additional vasorelaxation. Glabridin (30μM) approximately doubled tissue [cyclic GMP]. Application of the phosphodiesterase inhibitor isobutylmethylxanthine caused a much larger rise in [cyclic GMP], and glabridin failed to cause vasorelaxation or a further rise in [cGMP] when co-applied with IBMX.Conclusions and implicationsVasorelaxation to glabridin is dependent on the opening of K+ channels, particularly BKCa, probably caused by a rise in cellular [cyclic GMP] owing to phosphodiesterase inhibition. In the presence of sodium nitroprusside an effect of glabridin is observed at nM concentrations, similar those measured in plasma following human ingestion of licorice flavonoid oil

Sphingosylphosphorylcholine potentiates vasoreactivity and voltage-gated Ca²⁺entry via NOX1 and reactive oxygen species

AIMS: Sphingosylphosphorylcholine (SPC) elicits vasoconstriction at micromolar concentrations. At lower concentrations (≤1 µmol/L), however, it does not constrict intrapulmonary arteries (IPAs), but strongly potentiates vasoreactivity. Our aim was to determine whether this also occurs in a systemic artery and to delineate the signalling pathway. METHODS AND RESULTS: Rat mesenteric arteries and IPAs mounted on a myograph were challenged with ∼25 mmol/L [K(+)] to induce a small vasoconstriction. SPC (1 µmol/L) dramatically potentiated this constriction in all arteries by ∼400%. The potentiation was greatly suppressed or abolished by inhibition of phospholipase C (PLC; U73122), PKCε (inhibitory peptide), Src (PP2), and NADPH oxidase (VAS2870), and also by Tempol (superoxide scavenger), but not by inhibition of Rho kinase (Y27632). Potentiation was lost in mesenteric arteries from p47(phox–/–), but not NOX2(−/–), mice. The intracellular superoxide generator LY83583 mimicked the effect of SPC. SPC elevated reactive oxygen species (ROS) in vascular smooth muscle cells, and this was blocked by PP2, VAS2870, and siRNA knockdown of PKCε. SPC (1 µmol/L) significantly reduced the EC(50) for U46619-induced vasoconstriction, an action ablated by Tempol. In patch-clamped mesenteric artery cells, SPC (200 nmol/L) enhanced Ba(2+) current through L-type Ca(2+) channels, an action abolished by Tempol but mimicked by LY83583. CONCLUSION: Our results suggest that low concentrations of SPC activate a PLC-coupled and NOX1-mediated increase in ROS, with consequent enhancement of voltage-gated Ca(2+) entry and thus vasoreactivity. We speculate that this pathway is not specific for SPC, but may also contribute to vasoconstriction elicited by other G-protein coupled receptor and PLC-coupled agonists