Differential effects of cyclo-oxygenase 1 and 2 inhibition on angiogenesis inhibitor-induced hypertension and kidney damage.

Vascular endothelial growth factor antagonism with angiogenesis inhibitors in cancer patients induces a 'preeclampsia-like' syndrome including hypertension, proteinuria and elevated endothelin (ET)-1. Cyclo-oxygenase (COX) inhibition with aspirin is known to prevent the onset of preeclampsia in high-risk patients. In the present study, we hypothesised that treatment with aspirin would prevent the development of angiogenesis inhibitor-induced hypertension and kidney damage. Our aims were to compare the effects of low-dose (COX-1 inhibition) and high-dose (dual COX-1 and COX-2 inhibition) aspirin on blood pressure, vascular function, oxidative stress, ET-1 and prostanoid levels and kidney damage during angiogenesis-inhibitor therapy in rodents. To this end, Wistar Kyoto rats were treated with vehicle, angiogenesis inhibitor (sunitinib) alone or in combination with low- or high-dose aspirin for 8 days (n=5-7/group). Our results demonstrated that prostacyclin (PGI2) and ET-1 were increased during angiogenesis-inhibitor therapy, while thromboxane (TXA2) was unchanged. Both low- and high-dose aspirin blunted angiogenesis inhibitor-induced hypertension and vascular superoxide production to a similar extent, whereas only high-dose aspirin prevented albuminuria. While circulating TXA2 and prostaglandin F2α levels were reduced by both low- and high-dose aspirin, circulating and urinary levels PGI2 were only reduced by high-dose aspirin. Lastly, treatment with aspirin did not significantly affect ET-1 or vascular function. Collectively our findings suggest that prostanoids contribute to the development of angiogenesis inhibitor-induced hypertension and renal damage and that targeting the prostanoid pathway could be an effective strategy to mitigate the unwanted cardiovascular and renal toxicities associated with angiogenesis inhibitors.

Panax Notoginseng Ameliorates Podocyte EMT by Targeting the Wnt/ß-Catenin Signaling Pathway in STZ-Induced Diabetic Rats.

INTRODUCTION: Epithelial-mesenchymal transition (EMT) may contribute to podocyte dysfunction in diabetic nephropathy (DN). Aiming to identify novel therapeutic options, we investigated the protective effects of Panax notoginseng (PN) on podocyte EMT in diabetic rats and explored its mechanisms. METHODS: Diabetes was induced in rats with streptozotocin (STZ) by intraperitoneal injection at 55 mg/kg. Diabetic rats were randomly divided into three groups, namely, diabetic rats, diabetic rats treated with beraprost sodium (BPS) at 0.6 mg/kg/d or PN at 0.4 g/kg/d p.o., for 12 weeks. Urinary albumin/creatinine ratio (ACR), biochemical parameters, renal histopathology, and podocyte morphological changes were evaluated. Protein expression of EMT markers (desmin, α-SMA, and nephrin) as well as components of the Wnt/ß-catenin pathway (wnt1, ß-catenin, and snail) was detected by immunohistochemistry and Western blot, respectively. RESULTS: In diabetic rats, severe hyperglycemia and albuminuria were detected. Moreover, mesangial expansion and podocyte foot process effacement were found markedly increased in diabetic kidneys. Increased protein expression of wnt1, ß-catenin, snail, desmin, and α-SMA, as well as decreased protein expression of nephrin was detected in diabetic kidneys. All these abnormalities found in DN rats were partially restored by PN treatment. CONCLUSION: PN ameliorated albuminuria and podocyte EMT in diabetic rats partly through inhibiting Wnt/ß-catenin signaling pathway. These findings provide experimental arguments for a novel therapeutic option in DN.

Results of an Expert Consensus Survey on the Treatment of Pulmonary Arterial Hypertension With Oral Prostacyclin Pathway Agents.

BACKGROUND: Treatment of pulmonary arterial hypertension (PAH) has evolved substantially over the past two decades and varies according to etiology, functional class (FC), hemodynamic parameters, and other clinical factors. Current guidelines do not provide definitive recommendations regarding the use of oral prostacyclin pathway agents (PPAs) in PAH. To provide guidance on the use of these agents, an expert panel was convened to develop consensus statements for the initiation of oral PPAs in adults with PAH. METHODS: A systematic literature search was conducted using MEDLINE. The established RAND/University of California Los Angeles appropriateness method, which incorporates the Delphi method and the nominal group technique, was used to create consensus statements. Idiopathic, heritable, repaired congenital heart defect, and drug- or toxin-induced PAH (IPAH+) was considered as one etiologic grouping. The process was focused on the use of oral treprostinil or selexipag in patients with IPAH+ or connective tissue disease-associated PAH and FC II or III symptoms receiving background dual endothelin receptor antagonist/phosphodiesterase type 5 inhibitor therapy. RESULTS: The panel developed 14 consensus statements regarding the appropriate use of oral PPAs in the target population. The panel identified 13 clinical scenarios in which selexipag may be considered as a treatment option. CONCLUSIONS: The paucity of clinical evidence overall, and particularly from randomized trials in this setting, creates a gap in knowledge. These consensus statements are intended to aid physicians in navigating treatment options and using oral PPAs in the most appropriate manner in patients with PAH.

Effects of arachidonic acid and its major prostaglandin derivatives on bovine myoblast proliferation, differentiation, and fusion.

Many studies have shown positive effects of prostaglandins (PGs) on various steps of skeletal muscle formation such as myoblast proliferation and myotube hypertrophy. In animals, PGs are synthesized through the action of the rate-limiting enzymes cyclooxygenase (COX) -1 and COX-2 from arachidonic acid (AA), a conditionally essential fatty acid. As a step toward exploring the possibility of using dietary supplementation of AA to improve skeletal muscle growth in cattle, which are major meat-producing animals, we determined the effects of AA and its major PG derivatives PGE2, PGF2α, and PGI2 on proliferation, differentiation, and fusion of primary bovine myoblasts in vitro. In the proliferation experiment, myoblasts were cultured in a growth medium to which was added 10 µM AA, 1 µM PGE2, 1 µM PGF2α, 1 µM PGI2, or vehicle control for 24 h, and the proliferating cells were identified by 5-ethynyl-2'-deoxyuridine (EdU) labeling. This experiment revealed that AA, PGE2, PGF2α, and PGI2 each increased the number of proliferating cells by 13%, 24%, 16%, and 16%, respectively, compared to the control (n = 7, P < 0.05). In the differentiation and fusion test, myoblasts were induced to differentiate and fuse into myotubes in the presence of the aforementioned treatments for 0, 24, 48, and 72 h. Based on quantitative reverse transcription PCR analyses of mRNAs of myoblast differentiation and fusion markers (myogenin; myosin heavy chain 3; creatine kinase, muscle; myomaker) at 0, 24, and 48 h of differentiation, AA, PGE2, and PGF2α promoted myoblast differentiation (n = 6, P < 0.05). Based on Giemsa staining and counting the number of myotubes at 72 h of differentiation, PGE2 enhanced the number of formed myotubes by 14% (P < 0.05) compared to the control. Treating the myoblasts with AA and either the COX-1 and COX-2 common inhibitor indomethacin or the COX-2-specific inhibitor NS-398 reversed the stimulatory effect of AA on myoblast proliferation (n = 4, P < 0.05). Overall, this study demonstrates that exogenous AA stimulates bovine myoblast proliferation and differentiation in culture. The results of this study suggest that AA stimulates myoblast proliferation through its metabolites PGE2, PGF2α, or PGI2, and that AA stimulates myoblast differentiation through PGE2.

Beneficial Effect of Berberis amurensis Rupr. on Penile Erection.

OBJECTIVE: To investigate whether the methanol extract of Berberis amurensis Rupr. (BAR) augments penile erection using in vitro and in vivo experiments. METHODS: The ex vivo study used corpus cavernosum strips prepared from adult male New Zealand White rabbits. In in vivo studies for intracavernous pressure (ICP), blood pressure, mean arterial pressure (MAP), and increase of peak ICP were continuously monitored during electrical stimulation of Sprague-Dawley rats. RESULTS: Preconstricted with phenylephrine (PE) in isolated endotheliumintact rabbit corus cavernosum, BAR relaxed penile smooth muscle in a dose-dependent manner, which was inhibited by pretreatment with NG-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase inhibitor, and 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin-1-one, a soluble guanylyl cclase inhibitor. BAR significantly relaxed penile smooth muscles dose-dependently in ex vivo, and this was inhibited by pretreatment with L-NAME 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin-1-one. BAR-induced relaxation was significantly attenuated by pretreatment with tetraethylammonium (TEA, P<0.01), a nonselective K+ channel blocker, 4-aminopyridine (4-AP, P<0.01), a voltage-dependent K+ channel blocker, and charybdotoxin (P<0.01), a large and intermediate conductance Ca2+ sensitive-K+ channel blocker, respectively. BAR induced an increase in peak ICP, ICP/MAP ratio and area under the curve dose dependently. CONCLUSION: BAR augments penile erection via the nitric oxide/cyclic guanosine monophosphate system and Ca2+ sensitive-K+ (BKCa and IKCa) channels in the corpus cavernosum.

Preclinical Pharmacology and Pharmacokinetics of Inhaled Hexadecyl-Treprostinil (C16TR), a Pulmonary Vasodilator Prodrug.

This article describes the preclinical pharmacology and pharmacokinetics (PK) of hexadecyl-treprostinil (C16TR), a prodrug of treprostinil (TRE), formulated in a lipid nanoparticle (LNP) for inhalation as a pulmonary vasodilator. C16TR showed no activity (>10 µM) in receptor binding and enzyme inhibition assays, including binding to prostaglandin E2 receptor 2, prostaglandin D2 receptor 1, prostaglandin I2 receptor, and prostaglandin E2 receptor 4; TRE potently bound to each of these prostanoid receptors. C16TR had no effect (up to 200 nM) on platelet aggregation induced by ADP in rat blood. In hypoxia-challenged rats, inhaled C16TR-LNP produced dose-dependent (0.06-6 µg/kg), sustained pulmonary vasodilation over 3 hours; inhaled TRE (6 µg/kg) was active at earlier times but lost its effect by 3 hours. Single- and multiple-dose PK studies of inhaled C16TR-LNP in rats showed proportionate dose-dependent increases in TRE Cmax and area under the curve (AUC) for both plasma and lung; similar results were observed for dog plasma levels in single-dose PK studies. In both species, inhaled C16TR-LNP yielded prolonged plasma TRE levels and a lower plasma TRE Cmax compared with inhaled TRE. Inhaled C16TR-LNP was well tolerated in rats and dogs; TRE-related side effects included cough, respiratory tract irritation, and emesis and were seen only after high inhaled doses of C16TR-LNP in dogs. In guinea pigs, inhaled TRE (30 µg/ml) consistently produced cough, but C16TR-LNP (30 µg/ml) elicited no effect. These results demonstrate that C16TR-LNP provides long-acting pulmonary vasodilation, is well tolerated in animal studies, and may necessitate less frequent dosing than inhaled TRE with possibly fewer side effects.

The Second-Generation PGI2 Analogue Treprostinil Fails to Chemoprevent Tumors in a Murine Lung Adenocarcinoma Model.

Prostacyclin (prostaglandin I2, PGI2) overproduction in FVB/N mice prevents the formation of carcinogen and tobacco smoke-induced adenomas, and administration of the oral prostacyclin analogue iloprost to wild-type mice also prevented carcinogen-induced mouse lung adenoma formation. Former smokers taking oral iloprost showed improved bronchial dysplasia histology compared with placebo. Next-generation oral prostacyclin analogues, like treprostinil, were developed for the treatment of pulmonary arterial hypertension (PAH). On the basis of our prior studies with iloprost, we performed preclinical studies examining the ability of treprostinil to chemoprevent urethane-induced murine lung adenocarcinoma. We determined the MTD in chow (prior studies had delivered treprostinil by gavage), and this dose produced serum levels in the experimental animals similar to those found in PAH patients treated with treprostinil. We then examined the chemopreventive efficacy of treprostinil exposure initiated both before (1 week) and after (6 weeks) urethane exposure to better model chemoprevention studies conducted in former smokers. Neither of these dosing strategies prevented murine lung cancer; however, we did detect changes in pulmonary inflammatory cell infiltrate and expression of CXCR4 (a chemokine receptor previously shown to increase in response to treprostinil exposure) in tumor-bearing, treprostinil-treated animals, indicating that the drug was bioavailable. One potential explanation stems from iloprost and treprostinil differentially activating cell surface prostaglandin receptors and intracellular peroxisome proliferator-activated receptors. When murine lung tumor cells were treated with treprostinil, their proliferation rate increased; in contrast, iloprost had no effect on proliferation. Future investigations comparing these two agents will provide insight into iloprost's chemopreventive mechanisms. Cancer Prev Res; 10(11); 671-9. ©2017 AACR.

Notable alkaline tolerance of Kocuria marina isolate from blood of a pediatric patient with continuous intravenous epoprostenol therapy.

This study was the first to describe the hitherto deficiently evaluated alkaline tolerance of Kocuria marina isolate from a pediatric patient with continuous intravenous epoprostenol dosing therapy. Our isolate from blood of a 7-year-old Japanese boy was finally identified as K. marina by the morphological, cultural, and biochemical properties together with the comparative sequence analyses of the 16S rRNA genes. The K. marina isolate, the causative agent of catheter-related blood-stream infection, was not only revealed to be salt tolerant (NaCl 15%), but also demonstrated to be stably survived with no apparent decrease of cell counts for long periods (120 h) in an alkaline environment (pH 8, 9, 10, and 11) at 35 °C. Its remarkable tolerance to the stresses of high alkalinity compared with a clinical Staphylococcus aureus strain should provide consistent interpretation that the environment of high alkalinity (pH 10.2-10.8) measures should be insufficient to inactivate almost all the causative agents including K. marina strains in the solution of epoprostenol (pH 10.4) (Flolan(®), GlaxoSmithKline, Ltd., Tokyo, Japan.). To the best of our knowledge, the first description of the property of being tolerant to high alkalinity that the K. marina isolate exhibited was noteworthy and a useful piece of information. In conclusion, we believe that the present study should be a notification regarding the potential risk of catheter-related blood-stream infections due to K. marina, suggestive of an alkalophile, especially in patients receiving continuous intravenous epoprostenol dosing therapy.

The prostaglandin agonist beraprost aggravates doxorubicin-mediated apoptosis by increasing iNOS expression in cardiomyocytes.

Doxorubicin (DOX) is widely used as an anti-cancer agent although it causes irreversible cardiomyopathy by increasing oxidative stress and deregulating nitric oxide production. Beraprost (BPS), a stable prostacyclin (PGI2) analog, is a potent vasodilator that has beneficial effects on myocardial ischemia. The objectives of the present study were to delineate the uncertain effects of prostcyclin therapy on DOX induced cardiomyopathy and to explore the mechanisms underlying PGI2 and DOX interaction. For this reason, we stimulated endogenous PGI2 production using bicistronic COX-1/PGIS gene transfer and BPS supplementation, and investigated the effects on DOX-induced cardiomyopathy. Caspase-dependent protein content, lactate dehydrogenase (LDH), DNA fragmentation, and TUNEL positive cells were elevated in DOX-treated cardiomyocytes. These indicators were further elevated by adenovirus-COX- 1/PGIS transfection or BPS supplementation. In addition, PGI2 overexpression further increased iNOS expression and superoxide accumulation in cardiomyocytes compared with DOX alone, which may be the reason for aggravated cytotoxicity. Moreover, BPS can induce cAMP response elements (CRE) binding to the iNOS promoter and phospho- cAMP response element binding protein (CREB) expression in a cyclic AMP-dependent manner. Our in vivo studies show that MnTBAP and aminoguanidine treatment of DOX and BPS co-administered in mice can attenuate caspase-3 and PARP-1 protein expression, and improve mouse survival, as observed in the iNOS gene-deleted mice. In conclusion, we demonstrated that BPS or adv-COX-1/PGIS increases PGI2 levels through iNOS expression and peroxynitrite production, via CREB protein phosphorylation; thereby aggravating DOX-mediated cardiotoxicity.

A comparative study of PGI2 mimetics used clinically on the vasorelaxation of human pulmonary arteries and veins, role of the DP-receptor.

Prostacyclin (PGI2) and its mimetics (iloprost, treprostinil, beraprost and MRE-269) are potent vasodilators (via IP-receptor activation) and a major therapeutic intervention for pulmonary hypertension (PH). These PGI2 mimetics have anti-proliferative and potent vasodilator effects on pulmonary vessels. We compared the relaxant effects induced by these recognized IP-agonists in isolated human pulmonary arteries (HPA) and veins (HPV). In addition, using selective antagonists, the possible activation of other prostanoid relaxant receptors (DP, EP4) was investigated. Iloprost and treprostinil were the more potent relaxant agonists when both vessels were analyzed. HPA were significantly more sensitive to iloprost than to treprostinil, pEC50 values: 7.94±0.06 (n=23) and 6.73±0.08 (n=33), respectively. In contrast, in HPV these agonists were equipotent. The relaxations induced by treprostinil were completely or partially inhibited by IP-antagonists in HPA or HPV, respectively. The effects of the IP-agonists were not significantly modified by the EP4 antagonist. Finally, DP-antagonists inhibited the relaxations induced by treprostinil in HPV, suggesting that the DP-receptor plays a role in treprostinil-induced relaxation in the HPV. These data suggest that iloprost and treprostinil should be the most effective clinically available agonists to decrease pulmonary vascular resistance and to prevent oedema formation (by similar decrease in HPA and HPV resistance) in PH patients.

Endothelium-dependent vascular relaxation induced by Globularia alypum extract is mediated by EDHF in perfused rat mesenteric arterial bed.

The vasodilatory effect of Globularia alypum L. (GA) extract was evaluated in rat mesenteric arterial bed pre-contracted by continuous infusion of phenylephrine (2-4 ng/mL). Bolus injections of GA elicited dose-response vasodilation, which was abolished after endothelium removal. Addition of a nitric oxide synthase inhibitor, N(G)-nitro-l-arginine methyl ester (100 µmol/L), alone or in the presence of a cyclooxygenase inhibitor, indomethacin (10 µmol/L), did not significantly affect the vasodilation of the mesenteric arterial bed in response to GA extract. These results suggest that GA-induced vasodilation is endothelium dependent but nitric oxide and prostacyclin independent. In the presence of high K(+) (60 mmol/L), the GA vasodilatory effect was completely abolished, suggesting that the vasodilation effect is mediated by hyperpolarization of the vascular cells. Also, pre-treatment with atropine (a muscarinic receptors antagonist) antagonized the GA-induced vasodilation, suggesting that the vasodilatory effect is mainly mediated by the endothelium-derived hyperpolarizing factor through activation of endothelial muscarinic receptors.

Improvement of pulmonary microcirculation after lung transplantation using phosphodiesterase-5 inhibitor modified preservation solution.

OBJECTIVE: Optimising the preservation modality and thus maintaining the post-transplanted organ function remains a point of interest in research in order to prevent deleterious ischaemia/reperfusion injury. Microcirculation allows the assessment of initial graft function before obvious functional parameters. It was the aim of our study to compare the effects of epoprostenol and sildenafil on the pulmonary microcirculation and haemodynamics, when used in the preservation solution in lung transplantation. METHODS: Twenty-one pigs underwent single LuTx after 24h graft-ischaemia, preserved with buffered low potassium-dextran solution (I, control); with addition of 0.66 microg/kg/bw epoprostenol (II) or with 0.15 mg/kg/bw sildenafil (III). The pulmonary microcirculation, alveolar capillary diameter (ACD), red blood cell (RBC) velocity and functional capillary density (FCD), were assessed by intravital microscopy (OPS-imaging) hourly until 6h after reperfusion. Haemodynamics and blood gas exchange were monitored at all timepoints. RESULTS: ACD was increased in group III directly after reperfusion (132+/-4.4% vs 121+/-3.1%, in % of baseline, III vs I; mean+/-SEM; p<0.05) and decreased during the experiment. RBC velocity did not reach statistical significance (256+/-93 vs 263+/-85 and 283+/-66 microm/s, III vs II and I; mean+/-SD). FCD in group III was higher than in I and II beginning 3h after reperfusion (10.1+/-1.4 vs 6.1+/-1.9 microm/microm(2), III vs I; mean+/-SEM; p<0.05). CONCLUSIONS: Our study demonstrated a significantly improved microcirculation after application of PDF V during organ procurement, probably because of better distribution of the preservation solution. Further studies are necessary, to prove the long-term effects of this observation.

Prostacyclin prevents murine lung cancer independent of the membrane receptor by activation of peroxisomal proliferator--activated receptor gamma.

Overexpression of prostacyclin synthase (PGIS) decreases lung tumor multiplicity in chemical- and cigarette-smoke-induced murine lung cancer models. Prostacyclin signals through a single G-protein-coupled receptor (IP), which signals through cyclic AMP. To determine the role of this receptor in lung cancer chemoprevention by prostacyclin, PGIS-overexpressing mice were crossed to mice that lack the IP receptor [IP(-/-)]. Carcinogen-induced lung tumor incidence was similar in IP(+/+), IP(+/-), and IP(-/-) mice, and overexpression of PGIS gave equal protection in all three groups, indicating that the protective effects of prostacyclin are not mediated through activation of IP. Because prostacyclin can activate members of the peroxisomal proliferator-activated receptor (PPAR) family of nuclear receptors, we examined the role of PPARgamma in the protection of prostacyclin against lung tumorigenesis. Iloprost, a stable prostacyclin analogue, activated PPARgamma in nontransformed bronchial epithelial cells and in a subset of human non-small-cell lung cancer cell lines. Iloprost-impregnated chow fed to wild-type mice resulted in elevated lung macrophages and decreased lung tumor formation. Transgenic animals with lung-specific PPARgamma overexpression also developed fewer lung tumors. This reduction was not enhanced by administration of supplemental iloprost. These studies indicate that PPARgamma is a critical target for prostacyclin-mediated lung cancer chemoprevention and may also have therapeutic activity.

Negative mesenteric effects of lung recruitment maneuvers in oleic acid lung injury are transient and short lasting.

OBJECTIVE: To test the hypothesis that repeated recruitment maneuvers (RMs) have sustained negative effects on mesenteric circulation, metabolism, and oxygenation 60 mins after RMs in pigs with oleic acid lung injury. Further, we aimed to test the hypothesis that an infusion of prostacyclin (PC) at 33 ng.kg.min would attenuate such possible negative mesenteric effects. DESIGN: Randomized, experimental, controlled study. SETTING: University hospital animal laboratory. SUBJECTS: A total of 31 anesthetized, fluid-resuscitated pigs with oleic acid lung injury. INTERVENTIONS: : Animals were randomized to one of the following four groups: a control group (n = 7) that received no intervention, recruitment group (n = 8) that underwent the RM sequence, a prostacyclin group (n = 8) that received an infusion of PC, and a recruitment-prostacyclin group (n = 8) that received an infusion of PC and concomitant RM sequence. MEASUREMENTS AND MAIN RESULTS: We measured systemic and mesenteric hemodynamic variables, jejunal mucosal perfusion, mesenteric lactate flux, jejunal tissue oxygen tension, and mesenteric oxygen delivery, uptake, and extraction ratio. Five minutes after RMs, mesenteric oxygen extraction ratio and mesenteric lactate flux were more prominently increased in the recruitment group, giving evidence of worsened mesenteric conditions after RMs. These signs of worsened conditions were further supported by more decreased jejunal tissue oxygen tension and portal vein oxygen saturation in the recruitment group. PC preserved mesenteric oxygenation, as indicated by less of a decrease in portal vein oxygen saturation at the time corresponding to 5 mins after RM and less of a decrease in mesenteric oxygen delivery at the time corresponding to 15 mins after RM. PC preserved mesenteric oxygenation as indicated by less of a decrease in portal vein oxygen saturation at 5 mins after RM and an attenuated increase in mesenteric oxygen extraction ratio at 5 mins after RM. There was a trend toward worsened jejunal mucosal perfusion, although not significant. CONCLUSIONS: In an oleic acid lung injury model, three repeated RMs did not improve systemic oxygenation or lung mechanics. Negative effects on mesenteric oxygenation and metabolism were transient and short lasting. The intestinal effects of PC during RMs were minor and opposing, showing preserved oxygenation but a trend toward worsened mucosal perfusion.

Overview of treprostinil sodium for the treatment of pulmonary arterial hypertension.

Pulmonary arterial hypertension is a life-threatening disorder that refers to a group of diseases characterized by an abnormal elevation of the blood pressure within the pulmonary circulation due to a vasculopathy of the pulmonary microcirculation (1). If left untreated, the overall prognosis of pulmonary arterial hypertension is poor, with a 5-year survival rate of 34%. The most common cause of death is progressive right-sided heart failure (2). There is no pharmacologic cure for pulmonary arterial hypertension, and a team approach is required for the proper care of these patients (3). Treatment is directed at improving clinical symptoms, increasing exercise tolerance and extending survival. Until just a few years ago, standard treatment options for the treatment of pulmonary arterial hypertension were limited and included coumarin derivatives, calcium channel blockers, diuretics, digoxin and oxygen supplementation (4). In the last decade, the therapeutic options for pulmonary arterial hypertension have made considerable advances. In at least three major randomized controlled trials, the continuous intravenous infusion of epoprostenol, a synthetic salt of prostacyclin with potent vasodilatory ability, has been shown to improve exercise tolerance, hemodynamics, survival and quality of life in patients with New York Heart Association (NYHA) functional classes III and IV with either primary pulmonary hypertension or pulmonary arterial hypertension associated with scleroderma (5-9). Epoprostenol has now become a mainstay therapy in the long-term treatment of primary pulmonary hypertension or pulmonary arterial hypertension associated with scleroderma. Nevertheless, epoprostenol is far from an ideal form of therapy. Issues such as the short half-life of the drug, need for continuous central intravenous access and significant side effects have led to the development of more stable prostacyclin analogues as alternative therapies for primary pulmonary hypertension. These alternative therapies include iloprost (inhaled delivery) and treprostinil (subcutaneous delivery). As a result, the Food and Drug Administration (FDA) recently approved treprostinil for the treatment of pulmonary arterial hypertension in patients with NYHA classes II-IV. This review will offer a discussion of the basic pharmacology of treprostinil, its similarities to and differences from epoprostenol, the animal studies as well as the initial investigational studies leading to its FDA approval, and clinical uses of the drug alone or in combination with newer therapies directed at pulmonary arterial hypertension developed over the last decade.

Daily prickly pear consumption improves platelet function.

Prickly pear is traditionally used by Pima Indians as a dietary nutrient against diabetes mellitus. We examined the effect of daily consumption of 250 g in 8 healthy volunteers and 8 patients with mild familial heterozygous hypercholesterolemia on various parameters of platelet function. Beside its action on lipids and lipoproteins, prickly pear consumption significantly reduced the platelet proteins (platelet factor 4 and beta-thromboglobulin), ADP-induced platelet aggregation and improved platelet sensitivity (against PGI2 and PGE1) in volunteers as well as in patients. Also plasma 11-DH-TXB2 and the WU-test showed a significant improvement in both patients and volunteers. In contrast, collagen-induced platelet aggregation and the number of circulating endothelial cells showed a significant response in patients only. No influence of prickly pear ingestion on peripheral platelet count was monitored. The dietary run-in period did not influence any of the parameters of haemostasis examined. No sex difference was seen. Prickly pear may induce at least part of its beneficial actions on the cardiovascular system via decreasing platelet activity and thereby improving haemostatic balance.

Dietary zinc modifies the characteristics of endothelial dilation in normozincemic rats.

Because zinc attenuates endothelial cell dysfunction that proceeds atherosclerosis, depressed zinc status may be involved in the initiation of endothelial dysfunction. However, before recommending a zinc-enriched diet to reduce the risks for atherosclerosis, the effect of excess zinc on endothelial cell functions in normozincemic status should be known. Therefore, in this study, the effect of dietary zinc on normal endothelial cell functions in animals subjected to a diet containing 334 +/- 58 ppm zinc for 30 d was studied to see whether supplemented zinc has an effect on endothelial cells. Despite a slight increase in blood zinc, unaltered aortic and kidney zinc contents were associated with unchanged blood pressure in rats subjected to a zinc-enriched diet. Increased basal nitric oxide and prostacyclin were accompanied by a normal response to phenylephrine. Dietary zinc influenced neither endothelial-dependent nor endothelial-independent relaxations significantly. However, it elevated the share of M1-type cholinoceptor response as well as dilator prostaglandin release, which seems to be nitric oxide dependent. There was a strong correlation (r=0.826, p<0.05) between M1-type cholinoceptor response and prostacyclin release in zinc-treated rings. These results suggested that zinc ions increases M1-mediated prostacyclin release in normal endothelial cells without altering intracellular pathways.

Facilitation by endogenous prostaglandins of capsaicin-induced gastric protection in rodents through EP2 and IP receptors.

We investigated the role that prostaglandins (PGs) and EP receptors play in facilitating the gastroprotective action of capsaicin against HCl/ethanol in rats and mice. Male Sprague-Dawley rats and C57BL/6 mice were used after 18 h of fasting. The animals were given HCl/ethanol (60% in 150 mM HCl) p.o. and killed 1 h later. Capsaicin or various EP agonists were given p.o. 30 min or i.v. 10 min before HCl/ethanol. In some cases, indomethacin or various EP agonists were given s.c. 30 min or i.v 10 min before capsaicin, respectively. Gastric lesions induced by HCl/ethanol were significantly inhibited by PGE(2) as well as capsaicin. The effect of PGE(2) was antagonized by ONO-AE-829 (EP1 antagonist), whereas the capsaicin action was mitigated by indomethacin as well as sensory deafferentation but not by ONO-AE-829. The generation of mucosal PGE(2) was not affected by either capsaicin or sensory deafferentation, but was significantly inhibited by indomethacin. Although neither butaprost (EP2), ONO-NT-012 (EP3), nor 11-deoxy PGE1 (EP4) alone had any effect on HCl/ethanol-induced gastric lesions, only butaprost restored the protective action of capsaicin in the presence of indomethacin. Capsaicin provided a protective action against HCl/ethanol-induced gastric lesions in wild-type (+/+) mice in an indomethacin-sensitive manner, and this action was similarly observed in EP1 (-/-) and EP3 (-/-) mice but not in the animals lacking IP receptors. These results suggest that capsaicin exhibits gastric cytoprotection, essentially by stimulating sensory neurons, and this action is facilitated by endogenous PGs through EP2/IP receptors, probably sensitizing the sensory neurons to capsaicin.

Endothelial secretogogues and deformability of erythrocytes.

Many diseases of the heart and circulatory system have been linked with both dysfunction of vascular endothelium and insufficient deformability of erythrocytes. Using shear stress laser diffractometry we investigated whether deformability of erythrocytes would be regulated endogenously by generation of two endothelial secretogogues: prostacyclin and nitric oxide. Experiments were performed in rats ex vivo and with whole blood or isolated erythrocytes in vitro. Iloprost--a stable analogue of prostacyclin (10 microg/kg i.v.) and SIN-1 (NO-donor) at a dose of 2 mg/kg/min i.v induced a significant improvement of deformability of erythrocytes ex vivo. Improvements of deformability by these two compounds were also evident in vitro when they were applied at a range of concentrations of 1 microM and 3 microM, respectively. Cyclooxygenase (indomethacin 20 mg.kg i.v.) and nitric oxide synthase (L-NAME 10 mg/kg i.v.) inhibitors while worsening deformability ex vivo, they did not affect (3 mM and 10 microM, respectively) rheological functions of erythrocytes in vitro. Aggravating effects of these inhibitors on erythrocyte deformability ex vivo were reversed by prostacyclin and nitric oxide supplemented exogenously. Aspirin at a low (1 mg/kg i.v.) and high dose (50 mg/kg i.v.), contrary to indomethacin and L-NAME, aggravated erythrocyte deformability either ex vivo or in vitro. It is concluded that autocrine function of vascular endothelium plays an important role in regulation of rheology of red blood cells in flowing blood. The mechanism of this phenomenon is unclear but some possible explanations are discussed. In addition, in our experiments aspirin revealed unique erythrocyte damaging properties, possibly independent of inhibition of cyclooxygenase, but related to a direct protein acetylation.

Effects of selective cyclooxygenase-2 inhibition on vascular responses and thrombosis in canine coronary arteries.

BACKGROUND: Prostanoid synthesis via the action of cyclooxygenase-2 (COX-2) is a component of the inflammatory response. Prostacyclin, a product of COX-2 in vascular endothelium, has important physiological roles, such as increasing blood flow to injured tissues, reducing leukocyte adherence, and inhibiting platelet aggregation. We examined the possibility that selective COX-2 inhibition could suppress the protective effects of prostacyclin, resulting in an alteration of the hemostatic balance and vascular tone. METHODS AND RESULTS: Circumflex coronary artery thrombosis was induced in dogs by vascular electrolytic injury. Orally administered celecoxib (COX-2 inhibition) or high-dose aspirin (HDA) (COX-1 and COX-2 inhibition) did not alter time to occlusive thrombus formation compared with controls (celecoxib 77.7+/-7.2 minutes, HDA 72.0+/-18.5 minutes, control 93.0+/-21.8 minutes). Oral HDA with an endothelial recovery period (HDA-ER) (COX-1 inhibition) produced a significant increase in time to vessel occlusion (257.0+/-41.6 minutes). The observed increase in time to occlusion was abolished when celecoxib was administered to animals dosed with HDA-ER (80.7+/-20.6 minutes). The vasomotor effect of endothelium-derived prostacyclin was examined by monitoring coronary flow during intracoronary administration of arachidonic acid or acetylcholine. In celecoxib-treated animals, vasodilation in response to arachidonic acid was reduced significantly compared with controls. CONCLUSIONS: The results indicate important physiological roles for COX-2-derived prostacyclin and raise concerns regarding an increased risk of acute vascular events in patients receiving COX-2 inhibitors. The risk may be increased in individuals with underlying inflammatory disorders, including coronary artery disease.