16,830 research outputs found

    Furosemide Enhances the Release of Endothelial Kinsis, Nitric Oxide and Prostacyclin

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    Despite a wealth of data, the mechanism of the direct dilator effect of furosemide on the systemic arterial and venous systems is far from being satisfactorily understood. Therefore, we investigated whether furosemide is capable of stimulating the production of the endogenous vasodilators nitric oxide and prostacyclin in primary cultured bovine aortic endothelial cells by an enhanced synthesis and release of endothelium-derived kinins. Nitric oxide production was assessed in terms of intracellular guanosine cyclic-3',5' monophosphate accumulation; kinin and prostacyclin release were determined by specific radioimmunoassays. Furosemide concentration- and time- dependently increased the formation of nitric oxide and prostacyclin. Maximal increases of both autacoids were already obtained after a 5-min incubation with 3 x 10(-7) to 10(-6) mol/l of furosemide. In the same concentration range, furosemide led to an enhanced release of kinins into the supernatant of the cells. This observation was supported by the inhibitory effect of the specific B2 kinin receptor antagonist icatibant (Hoe 140) on the furosemide-induced increase of nitric oxide and prostacyclin. Thus the hemodynamic effects, and in particular the direct early dilator effect, of furosemide may be explained in part by an enhanced endothelial synthesis and release of bradykinin and related kinins, which in turn stimulates endothelial autacoid formation via B2 kinin receptor activation

    BCP crystals increase prostacyclin production and upregulate the prostacyclin receptor in OA synovial fibroblasts: potential effects on mPGES1 and MMP-13

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    SummaryObjectiveTo investigate the potential involvement of prostacyclin in basic calcium phosphate (BCP) crystal-induced responses in osteoarthritic synovial fibroblasts (OASF).MethodsOASF grown in culture were stimulated with BCP crystals. Prostacyclin production was measured by enzyme immunoassay. Expression of messenger RNA (mRNA) transcripts was assessed by real-time polymerase chain reaction (PCR). Expression of prostacyclin synthase (PGIS) and the prostacyclin (IP) receptor was measured. The effects of iloprost, a prostacyclin analogue, on expression of genes implicated in osteoarthritis such as microsomal prostaglandin E2 synthase 1 (mPGES1) and matrix metalloproteinases (MMPs) were also studied. FPT inhibitor II, a farnesyl transferase inhibitor, was used to antagonize iloprost-induced responses.ResultsBCP crystal stimulation led to a five-fold increase in prostacyclin production in OASF compared to untreated cells. This induction was attenuated by cyclooxygenase (COX)-2 and COX-1 inhibition at 4 and 32h, respectively. PGIS and IP receptor transcripts were constitutively expressed in OASF. BCP crystals upregulated IP receptor expression two-fold. While iloprost diminished BCP crystal-stimulated IP receptor upregulation, the inhibitory effect of iloprost was blocked by the farnesyl transferase inhibitor. In addition, iloprost upregulated mPGES1 and downregulated MMP-13 expression in BCP crystal-stimulated OASF, effects that were not influenced by the farnesyl transferase inhibitor.ConclusionsThese data showed for the first time that BCP crystals can increase prostacyclin production and upregulate expression of the IP receptor in OASF. The potential of prostacyclin to influence BCP crystal-stimulated responses was supported by the effects of iloprost on the expression of the IP receptor, mPGES1 and MMP-13. These data demonstrate the potential involvement of prostacyclin in BCP crystal-associated osteoarthritis (OA) and suggest that inhibition of PG synthesis with non-steroidal anti-inflammatory drugs may have both deleterious and beneficial effects in BCP crystal-associated OA

    Role of prostacyclin in pulmonary hypertension

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    Date of Acceptance: 11/12/2014 This is an open access article distributed under the terms of the Creative Commons Attribution license CC BY-4.0, which permits unrestricted use, distribution and reproduction in any medium, provided the original work is properly cited.Prostacyclin is a powerful cardioprotective hormone released by the endothelium of all blood vessels. Prostacyclin exists in equilibrium with other vasoactive hormones and a disturbance in the balance of these factors leads to cardiovascular disease including pulmonary arterial hypertension. Since it’s discovery in the 1980s concerted efforts have been made to make the best therapeutic utility of prostacyclin, particularly in the treatment of pulmonary arterial hypertension. This has centred on working out the detailed pharmacology of prostacyclin and then synthesising new molecules based on its structure that are more stable or more easily tolerated. In addition, newer molecules have been developed that are not analogues of prostacyclin but that target the receptors that prostacyclin activates. Prostacyclin and related drugs have without doubt revolutionised the treatment and management of pulmonary arterial hypertension but are seriously limited by side effects within the systemic circulation. With the dawn of nanomedicine and targeted drug or stem cell delivery systems it will, in the very near future, be possible to make new formulations of prostacyclin that can evade the systemic circulation allowing for safe delivery to the pulmonary vessels. In this way, the full therapeutic potential of prostacyclin can be realised opening the possibility that pulmonary arterial hypertension will become, if not curable, a chronic manageable disease that is no longer fatal. This review discusses these and other issues relating to prostacyclin and its use in pulmonary arterial hypertensionPeer reviewedFinal Published versio

    Induction of prostacyclin receptor expression in human erythroleukemia cells

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    AbstractWe have identified both high-affinity (KD = 36±3 nM) and low-affinity (KD = 2.1±0.8 μM) prostacyclin (PGI2)-receptor sites on human erythroleukemia (HEL) cells using the radiolabelled prostacyclin analogue, [3H]iloprost. The addition of the phorbol ester, TPA, to the culture medium caused a 5–10-fold increase in the number of both the low- and the high-affinity sites, without any change in their affinity constants. Iloprost stimulated HEL cell membrane adenylate cyclase activity 5-fold. This stimulation was potentiated in the presence of GTP, indicating a conventional PGI2 receptor-Gs-adenylate cyclase system. HEL cells represent a source of prostacyclin receptor mRNA which may be of value in expression cloning of this receptor

    High throughput mutagenesis for identification of residues regulating human prostacyclin (hIP) receptor

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    The human prostacyclin receptor (hIP receptor) is a seven-transmembrane G protein-coupled receptor (GPCR) that plays a critical role in vascular smooth muscle relaxation and platelet aggregation. hIP receptor dysfunction has been implicated in numerous cardiovascular abnormalities, including myocardial infarction, hypertension, thrombosis and atherosclerosis. Genomic sequencing has discovered several genetic variations in the PTGIR gene coding for hIP receptor, however, its structure-function relationship has not been sufficiently explored. Here we set out to investigate the applicability of high throughput random mutagenesis to study the structure-function relationship of hIP receptor. While chemical mutagenesis was not suitable to generate a mutagenesis library with sufficient coverage, our data demonstrate error-prone PCR (epPCR) mediated mutagenesis as a valuable method for the unbiased screening of residues regulating hIP receptor function and expression. Here we describe the generation and functional characterization of an epPCR derived mutagenesis library compromising >4000 mutants of the hIP receptor. We introduce next generation sequencing as a useful tool to validate the quality of mutagenesis libraries by providing information about the coverage, mutation rate and mutational bias. We identified 18 mutants of the hIP receptor that were expressed at the cell surface, but demonstrated impaired receptor function. A total of 38 non-synonymous mutations were identified within the coding region of the hIP receptor, mapping to 36 distinct residues, including several mutations previously reported to affect the signaling of the hIP receptor. Thus, our data demonstrates epPCR mediated random mutagenesis as a valuable and practical method to study the structurefunction relationship of GPCRs. © 2014 Bill et al

    Diverse pharmacology of prostacyclin mimetics: Implications for pulmonary hypertension

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    Pulmonary arterial hypertension (PAH) is a progressive vascular remodelling disease where patients ultimately die from heart failure. Increased production of vasoconstrictors (endothelin-1 and thromboxane A2) accompanied by loss of prostacyclin, nitric oxide (NO), bone morphogenetic protein receptor type 2 (BMPR2) and TASK-1 combine to cause endothelial apoptosis, smooth muscle hyperactivity and thickening of the blood vessel wall. Prostacyclin remains the most efficacious treatment for PAH, and several prostacyclin analogues are approved for use via different administration routes. They act as vasodilators but potently inhibit platelet aggregation, cell proliferation and inflammation. The pharmacology of each prostacyclin (IP) receptor agonist is distinct, with other targets contributing to their therapeutic and side-effect profile, including prostanoid EP1, EP3, EP2 and DP1 receptors, alongside peroxisome proliferator-activated receptors (PPARs), to which prostacyclin and some analogues directly bind. To improve selectivity, selexipag, a non-prostanoid was developed, whose only significant biological target is the IP receptor, but is a partial agonist in cyclic AMP assays and has no anti-aggregatory properties in vivo. Prostanoid receptor expression profiles in the normal and diseased lung demonstrate loss of the IP receptor and upregulation of EP2 and EP3 receptors in PAH, affecting the action of prostacyclin mimetics in different ways. We discuss how prostacyclins might rescue BMPR2 and TASK-1 dysfunction and the importance of EP2 receptors as negative modulators of vascular tone, proliferation and fibrosis. Alongside DP1 and EP4 receptors, they have specific roles in veins and airways. Whether drugs selective for the IP receptor confer a superior or reduced therapeutic benefit remains an important clinical question as do the role of platelets in PAH

    The role of endothelin-1 in pulmonary arterial hypertension.

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    Pulmonary arterial hypertension (PAH) is a rare but debilitating disease, which if left untreated rapidly progresses to right ventricular failure and eventually death. In the quest to understand the pathogenesis of this disease differences in the profile, expression and action of vasoactive substances released by the endothelium have been identified in patients with PAH. Of these, endothelin-1 (ET-1) is of particular interest since it is known to be an extremely powerful vasoconstrictor and also involved in vascular remodelling. Identification of ET-1 as a target for pharmacological intervention has lead to the discovery of a number of compounds that can block the receptors via which ET-1 mediates its effects. This review sets out the evidence in support of a role for ET-1 in the onset and progression of the disease and reviews the data from the various clinical trials of ET-1 receptor antagonists for the treatment of PAH

    The molecular genetics and cellular mechanisms underlying pulmonary arterial hypertension

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    Pulmonary arterial hypertension (PAH) is an incurable disorder clinically characterised by a sustained elevation of mean arterial pressure in the absence of systemic involvement. As the adult circulation is a low pressure, low resistance system, PAH represents a reversal to a foetal state. The small pulmonary arteries of patients exhibit luminal occlusion resultant from the uncontrolled growth of endothelial and smooth muscle cells. This vascular remodelling is comprised of hallmark defects, most notably the plexiform lesion. PAH may be familial in nature but the majority of patients present with spontaneous disease or PAH associated with other complications. In this paper, the molecular genetic basis of the disorder is discussed in detail ranging from the original identification of the major genetic contributant to PAH and moving on to current next-generation technologies that have led to the rapid identification of additional genetic risk factors. The impact of identified mutations on the cell is examined, particularly, the determination of pathways disrupted in disease and critical to pulmonary vascular maintenance. Finally, the application of research in this area to the design and development of novel treatment options for patients is addressed along with the future directions PAH research is progressing towards

    Consequences of altered eicosanoid patterns for nociceptive processing in mPGES-1-deficient mice

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    Cyclooxygenase-2 (COX-2)-dependent prostaglandin (PG) E2 synthesis in the spinal cord plays a major role in the development of inflammatory hyperalgesia and allodynia. Microsomal PGE2 synthase-1 (mPGES-1) isomerizes COX-2-derived PGH2 to PGE2. Here, we evaluated the effect of mPGES-1-deficiency on the noci-ceptive behavior in various models of nociception that depend on PGE2 synthesis. Surprisingly, in the COX-2-dependent zymosan-evoked hyperalgesia model, the nociceptive behavior was not reduced in mPGES-1-deficient mice despite a marked decrease of the spinal PGE2 synthesis. Similarly, the nociceptive behavior was unaltered in mPGES-1-deficient mice in the formalin test. Importantly, spinal cords and primary spinal cord cells derived from mPGES-1-deficient mice showed a redirection of the PGE2 synthesis to PGD2, PGF2α and 6-keto-PGF1α (stable metabolite of PGI2). Since the latter prostaglandins serve also as mediators of noci-ception they may compensate the loss of PGE2 synthesis in mPGES-1-deficient mice

    Estrogen, angiogenesis, immunity and cell metabolism: Solving the puzzle

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    Estrogen plays an important role in the regulation of cardiovascular physiology and the immune system by inducing direct effects on multiple cell types including immune and vascular cells. Sex steroid hormones are implicated in cardiovascular protection, including endothelial healing in case of arterial injury and collateral vessel formation in ischemic tissue. Estrogen can exert potent modulation effects at all levels of the innate and adaptive immune systems. Their action is mediated by interaction with classical estrogen receptors (ERs), ER\u3b1 and ER\u3b2, as well as the more recently identified G-protein coupled receptor 30/G-protein estrogen receptor 1 (GPER1), via both genomic and non-genomic mechanisms. Emerging data from the literature suggest that estrogen deficiency in menopause is associated with an increased potential for an unresolved inflammatory status. In this review, we provide an overview through the puzzle pieces of how 17\u3b2-estradiol can influence the cardiovascular and immune systems
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