Thrombospondin-1 promotes hemostasis through modulation of cAMP signaling in blood platelets.

Thrombospondin-1 (TSP-1) is released by platelets upon activation and can increase platelet activation, but its role in hemostasis in vivo is unclear. We show that TSP-1 is a critical mediator of hemostasis that promotes platelet activation by modulating inhibitory cyclic adenosine monophosphate (cAMP) signaling. Genetic deletion of TSP-1 did not affect platelet activation in vitro, but in vivo models of hemostasis and thrombosis showed that TSP-1-deficient mice had prolonged bleeding, defective thrombosis, and increased sensitivity to the prostacyclin mimetic iloprost. Adoptive transfer of wild-type (WT) but not TSP-1-/- platelets ameliorated the thrombotic phenotype, suggesting a key role for platelet-derived TSP-1. In functional assays, TSP-1-deficient platelets showed an increased sensitivity to cAMP signaling, inhibition of platelet aggregation, and arrest under flow by prostacyclin (PGI2). Plasma swap experiments showed that plasma TSP-1 did not correct PGI2 hypersensitivity in TSP-1-/- platelets. By contrast, incubation of TSP-1-/- platelets with releasates from WT platelets or purified TSP-1, but not releasates from TSP-1-/- platelets, reduced the inhibitory effects of PGI2. Activation of WT platelets resulted in diminished cAMP accumulation and downstream signaling, which was associated with increased activity of the cAMP hydrolyzing enzyme phosphodiesterase 3A (PDE3A). PDE3A activity and cAMP accumulation were unaffected in platelets from TSP-1-/- mice. Platelets deficient in CD36, a TSP-1 receptor, showed increased sensitivity to PGI2/cAMP signaling and diminished PDE3A activity, which was unaffected by platelet-derived or purified TSP-1. This scenario suggests that the release of TSP-1 regulates hemostasis in vivo through modulation of platelet cAMP signaling at sites of vascular injury.

Mechanism of vasorelaxation induced by 3'-hydroxy-5,6,7,4'-tetramethoxyflavone in the rats aortic ring assay.

Previous studies have demonstrated that 3'-hydroxy-5,6,7,4'-tetramethoxyflavone (TMF) content in Orthosiphon stamineus fractions correlate with its vasorelaxation activity. Even with the availability of previous studies, there is still very little information on the vasorelaxation effect of TMF, and few scientific studies have been carried out. Therefore, the present study was designed to investigate the vasorelaxation activity and mechanism of action of the TMF. The vasorelaxation activity and the underlying mechanisms of TMF were evaluated on thoracic aortic rings isolated from Sprague Dawley rats. TMF caused the relaxation of aortic rings with endothelium pre-contracted with phenylephrine. However, the vasorelaxant effect of TMF was significantly decreased in PE-primed endothelium-denuded and potassium chloride-primed endothelium-intact aortic rings. In the presence of Nω-nitro-L-arginine methyl ester, methylene blue, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, indomethacin, tetraethylammonium, 4-aminopyridine, barium chloride, atropine and propranolol, the relaxation stimulated by TMF was significantly reduced. TMF was also found to reduce Ca2+ release from sarcoplasmic reticulum (via IP3R) and block calcium channels (VOCC). The present study demonstrates the vasorelaxant effect of TMF involves NO/sGC/cGMP and prostacyclin pathways, calcium and potassium channels and muscarinic and beta-adrenergic receptors.

Inhibition of microsomal PGE synthase-1 reduces human vascular tone by increasing PGI2 : a safer alternative to COX-2 inhibition.

BACKGROUND AND PURPOSE: The side effects of cyclooxygenase-2 (COX-2) inhibitors on the cardiovascular system could be associated with reduced prostaglandin (PG)I2 synthesis. Microsomal PGE synthase-1 (mPGES-1) catalyses the formation of PGE2 from COX-derived PGH2 . This enzyme is induced under inflammatory conditions and constitutes an attractive target for novel anti-inflammatory drugs. However, it is not known whether mPGES-1 inhibitors could be devoid of cardiovascular side effects. The aim of this study was to compare, in vitro, the effects of mPGES-1 and COX-2 inhibitors on vascular tone in human blood vessels. EXPERIMENTAL APPROACH: The vascular tone and prostanoid release from internal mammary artery (IMA) and saphenous vein (SV) incubated for 30 min with inhibitors of mPGES-1 or COX-2 were investigated under normal and inflammatory conditions. KEY RESULTS: In inflammatory conditions, mPGES-1 and COX-2 proteins were more expressed, and increased levels of PGE2 and PGI2 were released. COX-2 and NOS inhibitors increased noradrenaline induced vascular contractions in IMA under inflammatory conditions while no effect was observed in SV. Interestingly, the mPGES-1 inhibitor significantly reduced (30-40%) noradrenaline-induced contractions in both vessels. This effect was reversed by an IP (PGI2 receptor) antagonist but not modified by NOS inhibition. Moreover, PGI2 release was increased with the mPGES-1 inhibitor and decreased with the COX-2 inhibitor, while both inhibitors reduced PGE2 release. CONCLUSIONS AND IMPLICATIONS: In contrast to COX-2 inhibition, inhibition of mPGES-1 reduced vasoconstriction by increasing PGI2 synthesis. Targeting mPGES-1 could provide a lower risk of cardiovascular side effects, compared with those of the COX-2 inhibitors. LINKED ARTICLES: This article is part of a themed section on Targeting Inflammation to Reduce Cardiovascular Disease Risk. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.22/issuetoc and http://onlinelibrary.wiley.com/doi/10.1111/bcp.v82.4/issuetoc.

Endothelial mechanotransduction proteins and vascular function are altered by dietary sucrose supplementation in healthy young male subjects.

KEY POINTS: Mechanotransduction in endothelial cells is a central mechanism in the regulation of vascular tone and vascular remodelling Mechanotransduction and vascular function may be affected by high sugar levels in plasma because of a resulting increase in oxidative stress and increased levels of advanced glycation end-products (AGE). In healthy young subjects, 2 weeks of daily supplementation with 3 × 75 g of sucrose was found to reduce blood flow in response to passive lower leg movement and in response to 12 W of knee extensor exercise. This vascular impairment was paralleled by up-regulation of platelet endothelial cell adhesion molecule (PECAM)-1, endothelial nitric oxide synthase, NADPH oxidase and Rho family GTPase Rac1 protein expression, an increased basal phosphorylation status of vascular endothelial growth factor receptor 2 and a reduced phosphorylation status of PECAM-1. There were no measurable changes in AGE levels. The findings of the present study demonstrate that daily high sucrose intake markedly affects mechanotransduction proteins and has a detrimental effect on vascular function. ABSTRACT: Endothelial mechanotransduction is important for vascular function but alterations and activation of vascular mechanosensory proteins have not been investigated in humans. In endothelial cell culture, simple sugars effectively impair mechanosensor proteins. To study mechanosensor- and vascular function in humans, 12 young healthy male subjects supplemented their diet with 3 × 75 g sucrose day-1 for 14 days in a randomized cross-over design. Before and after the intervention period, the hyperaemic response to passive lower leg movement and active knee extensor exercise was determined by ultrasound doppler. A muscle biopsy was obtained from the thigh muscle before and after acute passive leg movement to allow assessment of protein amounts and the phosphorylation status of mechanosensory proteins and NADPH oxidase. The sucrose intervention led to a reduced flow response to passive movement (by 17 ± 2%) and to 12 W of active exercise (by 9 ± 1%), indicating impaired vascular function. A reduced flow response to passive and active exercise was paralleled by a significant up-regulation of platelet endothelial cell adhesion molecule (PECAM-1), endothelial nitric oxide synthase, NADPH oxidase and the Rho family GTPase Rac1 protein expression in the muscle tissue, as well as an increased basal phosphorylation status of vascular endothelial growth factor receptor 2 and a reduced phosphorylation status of PECAM-1. The phosphorylation status was not acutely altered with passive leg movement. These findings indicate that a regular intake of high levels of sucrose can impair vascular mechanotransduction and increase the oxidative stress potential, and suggest that dietary excessive sugar intake may contribute to the development of vascular disease.

Role of prostacyclin signaling in endothelial production of soluble amyloid precursor protein-α in cerebral microvessels.

We tested hypothesis that activation of the prostacyclin (PGI2) receptor (IP receptor) signaling pathway in cerebral microvessels plays an important role in the metabolism of amyloid precursor protein (APP). In human brain microvascular endothelial cells activation of IP receptor with the stable analogue of PGI2, iloprost, stimulated expression of amyloid precursor protein and a disintegrin and metalloprotease 10 (ADAM10), resulting in an increased production of the neuroprotective and anticoagulant molecule, soluble APPα (sAPPα). Selective agonist of IP receptor, cicaprost, and adenylyl cyclase activator, forskolin, also enhanced expression of amyloid precursor protein and ADAM10. Notably, in cerebral microvessels of IP receptor knockout mice, protein levels of APP and ADAM10 were reduced. In addition, iloprost increased protein levels of peroxisome proliferator-activated receptor δ (PPARδ) in human brain microvascular endothelial cells. PPARδ-siRNA abolished iloprost-augmented protein expression of ADAM10. In contrast, GW501516 (a selective agonist of PPARδ) upregulated ADAM10 and increased production of sAPPα. Genetic deletion of endothelial PPARδ (ePPARδ-/-) in mice significantly reduced cerebral microvascular expression of ADAM10 and production of sAPPα. In vivo treatment with GW501516 increased sAPPα content in hippocampus of wild type mice but not in hippocampus of ePPARδ-/- mice. Our findings identified previously unrecognized role of IP-PPARδ signal transduction pathway in the production of sAPPα in cerebral microvasculature.

Comparisons between perivascular adipose tissue and the endothelium in their modulation of vascular tone.

The endothelium is an established modulator of vascular tone; however, the recent discovery of the anti-contractile nature of perivascular adipose tissue (PVAT) suggests that the fat, which surrounds many blood vessels, can also modulate vascular tone. Both the endothelium and PVAT secrete vasoactive substances, which regulate vascular function. Many of these factors are common to both the endothelium and PVAT; therefore, this review will highlight the potential shared mechanisms in the modulation of vascular tone. Endothelial dysfunction is a hallmark of many vascular diseases, including hypertension and obesity. Moreover, PVAT dysfunction is now being reported in several cardio-metabolic disorders. Thus, this review will also discuss the mechanistic insights into endothelial and PVAT dysfunction in order to evaluate whether PVAT modulation of vascular contractility is similar to that of the endothelium in health and disease. LINKED ARTICLES: This article is part of a themed section on Molecular Mechanisms Regulating Perivascular Adipose Tissue - Potential Pharmacological Targets? To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.20/issuetoc.

The vasorelaxant mechanisms of methanol on isolated rat aortic rings: Involvement of ion channels and signal transduction pathways.

It is reported that methanol is generally used as an industrial solvent, antifreeze, windshield washer fluid, cooking fuel and perfume. Methanol ingestion can lead to severe metabolic disturbances, blindness, or even death. So far, few studies about its negative effects on cardiovascular system have been reported. The purpose of this study was to determine the vasoactive effect of methanol and roles of ion channels and signal transduction pathways on isolated rat aorta. The results suggested that the mechanism of methanol-induced vasorelaxation at low concentrations (<500 mM) was mediated by ATP-sensitive K+ (KATP) and L-type Ca2+ channels, but the mechanism at high concentrations (>600 mM) was related to KATP, voltage-dependent K+, big-conductance Ca2+-activated K+, L-type Ca2+ channels as well as prostacyclin, protein kinase C, ß-adrenoceptors pathways. In addition, methanol induced a dose-dependent inhibition of vasoconstrictions caused by calcium chloride, potassium chloride, or norepinephrine. Further work is needed to investigate the relative contribution of each channel and pathway in methanol-induced vasoactive effect.

Oxymatrine attenuated isoproterenol-induced heart failure in rats via regulation of COX-2/PGI2 pathway.

Oxymatrine (OMT) is an active constituent of traditional Chinese herb Sophora japonica Ait which has been shown to exert potent anti-inflammatory,anti-oxidant and anti-fibrosis properties. Our previous studies have demonstrated that OMT has protective effects on isoproterenol-induced heart failure in rats through regulation of DDAH/ADMA metabolism pathway.In this study,we further investigated whether OMT could attenuate isoproterenol-induced heart failure through the regulation of COX-2/PGI2 pathway. Heart failure was induced in Sprague-Dawley rats by 5mg/kg isoproterenol subcutaneous injection for 7days. The rats were maintained on normal diet and randomly divided into five groups: control, isoproterenol, isoproterenol with OMT (50, 100mg/kg), and OMT alone groups (n=12 in each group). Serum brain natruretic peptide (BNP, a heart failure biomarker), histopathological variables, expression of Cytosolic phospholipase A2 (cPLA2), cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2) and Prostacyclin synthase (PGIS) were analysed. Administration of OMT significantly reduced the increased BNP in plasm of isoproterenol-induced rats, attenuated cardiac fibrosis,suppressed overexpression of myocardial COX-1 expression, up-regulated COX-2 and PGIS expression, but had no effects on isoproterenol-induced elevated protein cPLA2. And compared with control group, any indexes in sham rats treated with OMT (100mg/kg) alone were unaltered. These results demonstrated that OMT has cardioprotective effects on isoproterenol-induced heart failure in rats by regulating COX-2/PGI2 pathway.

Cardiovascular Consequences of Prostanoid I Receptor Deletion in Microsomal Prostaglandin E Synthase-1-Deficient Hyperlipidemic Mice.

BACKGROUND: Inhibitors of cyclooxygenase-2 alleviate pain and reduce fever and inflammation by suppressing the biosynthesis of prostacyclin (PGI2) and prostaglandin E2. However, suppression of these prostaglandins, particularly PGI2, by cyclooxygenase-2 inhibition or deletion of its I prostanoid receptor also predisposes to accelerated atherogenesis and thrombosis in mice. By contrast, deletion of microsomal prostaglandin E synthase 1 (mPGES-1) confers analgesia, attenuates atherogenesis, and fails to accelerate thrombogenesis, while suppressing prostaglandin E2, but increasing biosynthesis of PGI2. METHODS: To address the cardioprotective contribution of PGI2, we generated mice lacking the I prostanoid receptor together with mPges-1 on a hyperlipidemic background (low-density lipoprotein receptor knockouts). RESULTS: mPges-1 depletion modestly increased thrombogenesis, but this response was markedly further augmented by coincident deletion of the I prostanoid receptor (n=10-18). By contrast, deletion of the I prostanoid receptor had no effect on the attenuation of atherogenesis by mPGES-1 deletion in the low-density lipoprotein receptor knockout mice (n=17-21). CONCLUSIONS: Although suppression of prostaglandin E2 accounts for the protective effect of mPGES-1 deletion in atherosclerosis, augmentation of PGI2 is the dominant contributor to its favorable thrombogenic profile. The divergent effects on these prostaglandins suggest that inhibitors of mPGES-1 may be less likely to cause cardiovascular adverse effects than nonsteroidal anti-inflammatory drugs specific for inhibition of cyclooxygenase-2.

5-HT2 receptor blockade exhibits 5-HT vasodilator effects via nitric oxide, prostacyclin and ATP-sensitive potassium channels in rat renal vasculature.

The aim of this study was to determine whether orally sarpogrelate (selective 5-HT2 antagonist) treatment (30 mg/kg/day; 14 days) could modify 5-HT renal vasoconstrictor responses, characterizing 5-HT receptors and mediator mechanisms involved in serotonergic responses in the in situ autoperfused rat kidney. Intra-arterial (i.a.) injections of 5-HT (0.00000125 to 0.1 µg/kg) decreased renal perfusion pressure (RPP) but did not affect the mean blood pressure (MBP). i.a. agonists 5-CT (5-HT1/7), CGS-12066B (5-HT1B), L-694,247 (5-HT1D) or AS-19 (5-HT7) mimicked renal 5-HT vasodilator effect. However, neither 8-OH-DPAT (5-HT1A) nor 1-phenylbiguanide (5-HT3) modified RPP. Moreover: (i) GR-55562 (5-HT1B antagonist) and L-NAME (nitric oxide synthase [NOS] inhibitor) blocked CGS-12066B-induced vasodilator response, (ii) LY310762 (5-HT1D antagonist) and indomethacin (non-selective cyclooxygenase inhibitor) blocked L-694,247-induced vasodilator response; (iii) SB-258719 (5-HT7 antagonist) and glibenclamide (ATP-sensitive K+ channel blocker) blocked AS-19-induced vasodilator response; and (iv) 5-HT- or 5-CT-elicited renal vasodilation was significantly blocked by the mixture of GR-55562 + LY310762 + SB-258719. Furthermore, eNOS and iNOS proteins and prostacyclin levels are overexpressed in sarpogrelate-treated rats. Our data suggest that 5-HT exerts renal vasodilator effect in the in situ autoperfused sarpogrelate-treated rat kidney, mediated by 5-HT1D, 5-HT1B and 5-HT7 receptors, involving cyclooxygenase-derived prostacyclin, nitric oxide synthesis/release and ATP-sensitive K+ channels, respectively.

Prostaglandin I2 Signaling and Inhibition of Group 2 Innate Lymphoid Cell Responses.

RATIONALE: Group 2 innate lymphoid cells (ILC2s) robustly produce IL-5 and IL-13, cytokines central to the asthma phenotype; however, the effect of prostaglandin (PG) I2 on ILC2 function is unknown. OBJECTIVES: To determine the effect of PGI2 on mouse and human ILC2 cytokine expression in vitro and the effect of endogenous PGI2 and the PGI2 analog cicaprost on lung ILC2s in vivo. METHODS: Flow-sorted bone marrow ILC2s of wild-type (WT) and PGI2 receptor-deficient (IP(-/-)) mice were cultured with IL-33 and treated with the PGI2 analog cicaprost. WT and IP(-/-) mice were challenged intranasally with Alternaria alternata extract for 4 consecutive days to induce ILC2 responses, and these were quantified. Prior to A. alternata extract, challenged WT mice were treated with cicaprost. Human flow-sorted peripheral blood ILC2s were cultured with IL-33 and IL-2 and treated with the PGI2 analog cicaprost. MEASUREMENT AND MAIN RESULTS: We demonstrate that PGI2 inhibits IL-5 and IL-13 protein expression by IL-33-stimulated ILC2s purified from mouse bone marrow in a manner that was dependent on signaling through the PGI2 receptor IP. In a mouse model of 4 consecutive days of airway challenge with an extract of A. alternata, a fungal aeroallergen associated with severe asthma exacerbations, endogenous PGI2 signaling significantly inhibited lung IL-5 and IL-13 protein expression, and reduced the number of lung IL-5- and IL-13-expressing ILC2s, as well as the mean fluorescence intensity of IL-5 and IL-13 staining. In addition, exogenous administration of a PGI2 analog inhibited Alternaria extract-induced lung IL-5 and IL-13 protein expression, and reduced the number of lung IL-5- and IL-13-expressing ILC2s and the mean fluorescence intensity of IL-5 and IL-13 staining. Finally, a PGI2 analog inhibited IL-5 and IL-13 expression by human ILC2s that were stimulated with IL-2 and IL-33. CONCLUSIONS: These results suggest that PGI2 may be a potential therapy to reduce the ILC2 response to protease-containing aeroallergens, such as Alternaria.

Time-dependent activation of prostacyclin and nitric oxide pathways during continuous i.v. infusion of serelaxin (recombinant human H2 relaxin).

BACKGROUND AND PURPOSE: In the RELAX-AHF trial, a 48 h i.v. serelaxin infusion reduced systemic vascular resistance in patients with acute heart failure. Consistent with preclinical studies, serelaxin augments endothelial vasodilator function in rat mesenteric arteries. Little is known about the contribution of endothelium-derived relaxing factors after a longer duration of continuous serelaxin treatment. Here we have assessed vascular reactivity and mechanistic pathways in mesenteric arteries and veins and the aorta after 48 or 72 h continuous i.v. infusion of serelaxin. EXPERIMENTAL APPROACH: Male rats were infused with either placebo or serelaxin (13.3 µg·kg(-1) ·h(-1) ) via the jugular vein using osmotic minipumps. Vascular function was assessed using wire myography. Changes in gene and protein expression and 6-keto PGF1α levels were determined by quantitative PCR, Western blot and ELISA respectively. KEY RESULTS: Continuous i.v. serelaxin infusion augmented endothelium-dependent relaxation in arteries (mesenteric and aorta) but not in mesenteric veins. In mesenteric arteries, 48 h i.v. serelaxin infusion increased basal NOS activity, associated with increased endothelial NOS (eNOS) expression. Interestingly, phosphorylated-eNOS(Ser1177) , eNOS and basal NOS activity were reduced in mesenteric arteries following 72 h serelaxin treatment. At 72 h, serelaxin treatment improved bradykinin-mediated relaxation through COX2-derived PGI2 production. CONCLUSIONS AND IMPLICATIONS: Continuous i.v. serelaxin infusion enhanced endothelial vasodilator function in arteries but not in veins. The underlying mediator at 48 h was NO but there was a transition to PGI2 by 72 h. Activation of the PGI2 -dependent pathway is key to the prolonged vascular response to serelaxin treatment.

Potassium channels and prostacyclin contribute to vasorelaxant activities of Tridax procumbens crude aqueous leaf extract in rat superior mesenteric arteries.

BACKGROUND: Previous studies have shown that aqueous extract of the leaf of Tridax procuinbens is capable of lowering blood pressure through its vasodilatory effects. In the present study attempt was made to examine the biological active components of T procuinbens leaf using GC-MS methods. We further investigated the role of K+ channels in the vasorelaxation effects of Tridax procumbens using rat isolated mesenteric artery. METHODS: The superior mesenteric artery isolated from healthy, young adult Wistar rats (250-300 g) were precontracted with phenylephrine (PE) (10(-7) M) and potassium chloride (KCl) (60 mM) and were treated with Various concentrations of aqueous extract ofT procumbens (0.9.0 mg/ml). The changes in arterial tension were recorded using a force-displacement transducer (Model 7004; Ugo Basil Varese, Italy) coupled to data capsule acquisition system. RESULTS: The results of GG-MS revealed the presence of linoleic acid. The T. procumbens extract (TPE) ranging from 0.5-9.0 mg/mI significantly (p<0.05) reduced the, contraction induced by (PE) and (KCl) in a concentration-dependent manner. The extract also antagonised the calcium-induced vasoconstriction (1(-9) - 10(-5)) in calcium-free with high concentration of potassium as well as. in calcium- and potassium free physiological solutions. The vasorelaxing effect caused by TPE was significantly (p<0.05) attenuated with preincubation of potassium channels blockers (Barium chloride and apamin), NO synthaseinhibitor (L-NAME), prostacyclin inhibitor (indomethacin), atropine; propranolol, and methylene blue while it was not affected by preincubation with glibenclamide and tetra ethyl ammonium, 4-aminopyridine (4-AP) and oxadiazolo quinoxalin (ODQ). CONCLUSION: The results of this study demonstrate that T procumbens extract causes vasodilatory effects by blocking calcium channels and the vasodilatory effect of the extract may also be due to stimulation of prostacyclin production and opening of small-conductance Ga2+ activated potassium channels. The observed effect of this extract may be probably due to the presence of linoleic acid in this extract.

[Impacts of high-volume hemofiltration on hemodynamics and vascular endothelial permeability in children with septic shock].

OBJECTIVE: To investigate the effects of high-volume hemofiltration (HVHF) on hemodynamics, vasoactive factors, and vascular endothelial permeability in children with septic shock by a comparative analysis. METHODS: Thirty-six children who were diagnosed with septic shock between January 2013 and September 2014 were randomly divided into control and observation groups (n=18 each). Children in the control group were treated with the standard-volume hemofiltration (SVHF), while children in the observation group were treated with HVHF. The hemodynamic indices and levels of vasoactive factors including 6-keto-prostaglandin F1α (6-keto-PGF1α), thromboxane B2 (TXB2), soluble E-selectin (sE-selectin), and endothelium-derived relaxing factor (EDRF) were determined before and after treatment. In addition, the effects of ultrafiltrate on endothelial cell permeability were assessed. RESULTS: Compared with the control group, the observation group had significantly higher mean arterial pressure, significantly higher blood oxygen saturation, and a significantly lower heart rate after treatment (P<0.05). The levels of TXB2 and sE-selectin were significantly lower in the observation group than in the control group (P<0.05), while the levels of 6-keto-PGF1α and EDRF were significantly higher in the observation group than in the control group (P<0.05). Compared with the control group, the ultrafiltrate significantly attenuated the transepithelial electrical resistance in the observation group (P<0.05). CONCLUSIONS: Compared with SVHF, HVHF is a more effective approach for improving the hemodynamics and levels of vasoactive factors and reducing the vascular endothelial permeability in children with septic shock.

Involvement of prostaglandin I(2) in nitric oxide-induced vasodilation of retinal arterioles in rats.

The soluble guanylyl cyclase/cGMP system plays an important role in the vasodilator response to nitric oxide (NO) in various vascular beds. However, in rat retinal arterioles, the cyclooxygenase-1/cAMP-mediated pathway contributes to the vasodilator effects of NO, although the specific prostanoid involved remains to be elucidated. In the present study, we investigated the role of prostaglandin I2 and its receptor (prostanoid IP receptor) system in NO-induced vasodilation of rat retinal arterioles in vivo. Fundus images were captured using a digital camera that was equipped with a special objective lens. Changes in diameter of retinal arterioles were assessed. The NO donor (±)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR3) increased the diameter of retinal arterioles but decreased systemic blood pressure in a dose-dependent manner. Treatment of rats with indomethacin, a non-selective cyclooxygenase inhibitor, markedly attenuated the retinal vasodilator, but not depressor responses to NOR3. The prostanoid IP receptor antagonist 4,5-dihydro-N-[4-[[4-(1-methylethoxy)phenyl]methyl]phenyl]-1H-imadazol-2-amine (CAY10441), and the prostaglandin I2 synthase inhibitor 9α,11α-azoprosta-5Z,13E-dien-1-oic acid (U-51605), both showed similar preventive effects against the NOR3-induced retinal vasodilator response. Neither CAY10441 nor U-51605 showed any significant effects on the depressor response to NOR3. NOR3 enhanced the release of prostaglandin I2 from cultured human retinal microvascular endothelial cells and the NOR3-induced prostaglandin I2 release was almost completely abolished by the cyclooxygenase-1 inhibitor SC-560, but not by the cyclooxygenase-2 inhibitor NS-398. However, NOR3 did not increase the release of prostaglandin I2 from human intestinal microvascular endothelial cells. These results suggest that NO exerts its dilatory effect via cyclooxygenase-1/prostaglandin I2/prostanoid IP receptor signaling mechanisms in the retinal vasculature.

Epoprostenol sodium for treatment of pulmonary arterial hypertension.

The release of endogenous prostacyclin (PGI2) is depressed in patients with pulmonary arterial hypertension (PAH). PGI2 replacement therapy by epoprostenol infusion is one of the best treatments available for PAH. Here, we provide an overview of the current clinical data for epoprostenol. Epoprostenol treatment improves symptoms, exercise capacity, and hemodynamics, and is the only treatment that has been shown to reduce mortality in patients with idiopathic PAH (IPAH) in randomized clinical trials. We have reported that high-dose epoprostenol therapy (>40 ng/kg/min) also results in marked hemodynamic improvement in some patients with IPAH. High-dose epoprostenol has a pro-apoptotic effect on PAH-PASMCs via the IP receptor and upregulation of Fas ligand (FasL) in vitro. However, long-term intravenous administration of epoprostenol is sometimes associated with catheter-related infections and leads to considerable inconvenience for the patient. In the future, the development of new routes of administration or the development of powerful PGI2 analogs, IP-receptor agonists, and gene and cell-based therapy enhancing PGI2 production with new routes of administration is required.

Inhibition of cyclooxygenase-2 causes a decrease in coronary flow in diabetic mice. The possible role of PGE2 and dysfunctional vasodilation mediated by prostacyclin receptor.

Several lines of evidence suggest that cyclooxygenase-2 (COX-2) activity can have a beneficial role in the maintenance of vascular tone of the blood vessels in diabetes. Specifically, the increased production of prostacyclin (PGI2) and prostaglandin E2 (PGE2), mediated by COX-2, has been suggested to compensate for decreased synthesis of nitric oxide (NO). The study investigates whether inhibition of COX-2 may reduce the coronary flow in diabetic animals and may also lead to decreased synthesis of prostaglandins. Mice aged 18-20 weeks were used for the study: those with leptin receptor deficiency (db/db) served as a model of diabetes while heterozygous (db/+) mice served as controls. Coronary flow was measured by the Langendorff method, and prostaglandin synthesis by myocardia was assayed in heart perfusates. COX-2 inhibition was found to reduce basal coronary flow in db/db mice but had no effect in db/+ mice. Secretion of PGE2 was found to be higher in db/db mice, while prostacyclin synthesis did not differ. COX-2 inhibition decreased production of both prostaglandins to similar levels in both groups. The use of ONO-1301, a specific agonist for the prostacyclin receptor revealed that vasodilating responses mediated by the receptor were impaired in db/db mice. The expression levels of the receptor in cardiac tissue did not differ between the groups. It is concluded that the increased COX-2 contribution to vasodilation in diabetic animals appears to be partially a result of increased COX-2-dependent synthesis of PGE2 and also may be caused by impaired vasodilation mediated by the prostacyclin receptor.

Prostacyclin protects vascular integrity via PPAR/14-3-3 pathway.

Vascular integrity is protected by the lining endothelial cells (ECs) through structural and molecular protective mechanisms. In response to external stresses, ECs are dynamic in producing protective molecules such as prostacyclin (PGI2). PGI2 is known to inhibit platelet aggregation and controls smooth muscle cell contraction via IP receptors. Recent studies indicate that PGI2 defends endothelial survival and protects vascular smooth muscle cell from apoptosis via peroxisome-proliferator activated receptors (PPAR). PPAR activation results in 14-3-3 upregulation. Increase in cytosolic 14-3-3ɛ or 14-3-3ß enhances binding and sequestration of Akt-mediated phosphorylated Bad and reduces Bad-mediated apoptosis via the mitochondrial pathway. Experimental data indicate that administration of PGI2 analogs or augmentation of PGI2 production by gene transfer attenuates endothelial damage and organ infarction caused by ischemia-reperfusion injury. The protective effect of PGI2 is attributed in part to preserving endothelial integrity.

Importance of cyclooxygenase-1/prostacyclin in modulating gastric mucosal integrity under stress conditions.

BACKGROUND AND AIM: We investigated the roles of cyclooxygenase (COX) isozymes and prostaglandins (PGs) and their receptors in mucosal defense against cold-restraint stress (CRS)-induced gastric lesions. METHODS: Male C57BL/6 wild-type (WT) mice and those lacking COX-1 or COX-2 as well as those lacking EP1, EP3, or IP receptors were used after 18 h fasting. Animals were restrained in Bollman cages and kept in a cold room at 10°C for 90 min. RESULTS: CRS induced multiple hemorrhagic lesions in WT mouse stomachs. The severity of these lesions was significantly worsened by pretreatment with the nonselective COX inhibitors (indomethacin, loxoprofen) or selective COX-1 inhibitor (SC-560), while neither of the selective COX-2 inhibitors (rofecoxib and celecoxib) had any effect. These lesions were also aggravated in animals lacking COX-1, but not COX-2. The expression of COX-2 mRNA was not detected in the stomach after CRS, while COX-1 expression was observed under normal and stressed conditions. The gastric ulcerogenic response to CRS was similar between EP1 or EP3 knockout mice and WT mice, but was markedly worsened in animals lacking IP receptors. Pretreating WT mice with iloprost (the PGI2 analog) significantly prevented CRS-induced gastric lesions in the presence of indomethacin. PGE2 also reduced the severity of these lesions, and the effect was mimicked by the EP4 agonist, AE1-329. CONCLUSIONS: These results suggest that endogenous PGs derived from COX-1 play a crucial role in gastric mucosal defense during CRS, and this action is mainly mediated by PGI2 /IP receptors and partly by PGE2 /EP4 receptors.