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Inhibitors of microsomal prostaglandin E synthase 1 (mPGES-1) are in the early phase of clinical development. Deletion of mPges-1 in mice confers analgesia, restrains atherogenesis, and fails to accelerate thrombogenesis, while suppressing prostaglandin E2 (PGE2), but increasing the biosynthesis of prostacyclin (PGI2). In low-density lipoprotein receptor-deficient (Ldlr-/-) mice, this last effect represents the dominant mechanism by which mPges-1 deletion restrains thrombogenesis, while suppression of PGE2 accounts for its antiatherogenic effect. However, the effect of mPges-1 depletion on blood pressure (BP) in this setting remains unknown. Here, we show that mPges-1 depletion significantly increased the BP response to salt loading in male Ldlr-/- mice, whereas, despite the direct vasodilator properties of PGI2, deletion of the I prostanoid receptor (Ipr) suppressed this response. Furthermore, combined deletion of the Ipr abrogated the exaggerated BP response in male mPges-1-/- mice. Interestingly, these unexpected BP phenotypes were not observed in female mice fed a high-salt diet (HSD). This is attributable to the protective effect of estrogen in Ldlr-/- mice and in Ipr-/- Ldlr-/- mice. Thus, estrogen compensates for a deficiency in PGI2 to maintain BP homeostasis in response to high salt in hyperlipidemic female mice. In male mice, by contrast, the augmented formation of atrial natriuretic peptide (ANP) plays a similar compensatory role, restraining hypertension and oxidant stress in the setting of Ipr depletion. Hence, men with hyperlipidemia on a HSD might be at risk of a hypertensive response to mPGES-1 inhibitors.

Objective: Deletion of mPGES-1 (microsomal prostaglandin E synthase-1)-an anti-inflammatory target alternative to COX (cyclooxygenase)-2-attenuates injury-induced neointima formation in mice. This is attributable to the augmented levels of PGI 2 (prostacyclin)-a known restraint of the vascular response to injury, acting via IP (I prostanoid receptor). To examine the role of mPGES-1-derived PGE 2 (prostaglandin E 2 ) in vascular remodeling without the IP., Approach and Results: Mice deficient in both IP and mPGES-1 (DKO [double knockout] and littermate controls [IP KO (knockout)]) were subjected to angioplasty wire injury. Compared with the deletion of IP alone, coincident deletion of IP and mPGES-1 increased neointima formation, without affecting media area. Early pathological changes include impaired reendothelialization and increased leukocyte invasion in neointima. Endothelial cells (ECs), but not vascular smooth muscle cells, isolated from DKOs exhibited impaired cell proliferation. Activation of EP (E prostanoid receptor) 4 (and EP2, to a lesser extent), but not of EP1 or EP3, promoted EC proliferation. EP4 antagonism inhibited proliferation of mPGES-1-competent ECs, but not of mPGES-1-deficient ECs, which showed suppressed PGE 2 production. EP4 activation inhibited leukocyte adhesion to ECs in vitro, promoted reendothelialization, and limited neointima formation post-injury in the mouse. Endothelium-restricted deletion of EP4 in mice suppressed reendothelialization, increased neointimal leukocytes, and exacerbated neointimal formation., Conclusions: Removal of the IP receptors unmasks a protective role of mPGES-1-derived PGE 2 in limiting injury-induced vascular hyperplasia. EP4, in the endothelial compartment, is essential to promote reendothelialization and restrain neointimal formation after injury. Activating EP4 bears therapeutic potential to prevent restenosis after percutaneous coronary intervention., (© 2018 American Heart Association, Inc.)

Allergic airway diseases are immune disorders associated with heightened type 2 immune responses and IL-5 and IL-13 production at the site of inflammation. We have previously reported that cyclooxygenase (COX) inhibition by indomethacin augmented allergic airway inflammation in a STAT6-independent manner. However, the key COX product(s) responsible for restraining indomethacin-mediated STAT6-independent allergic inflammation is unknown. In this study, using the mouse model of OVA-induced allergic airway inflammation, we identified that PGI2 receptor (IP) signaling was critical for indomethacin-induced, STAT6-independent proallergic effects. We demonstrated that IP deficiency increased inflammatory cell infiltration, eosinophilia, and IL-5 and IL-13 expression in the lung in a STAT6-independent manner. The augmented STAT6-independent allergic inflammation correlated with enhanced primary immune responses to allergic sensitization and elevated production of multiple inflammatory chemokines (CCL11, CCL17, CCL22, and CXCL12) in the lung after allergen challenge. We also showed that the PGI2 analogue cicaprost inhibited CD4 T cell proliferation and IL-5 and IL-13 expression in vitro, and IP deficiency diminished the stimulatory effect of indomethacin on STAT6-independent IL-5 and IL-13 responses in vivo. The inhibitory effects of PGI2 and the IP signaling pathway on CD4 T cell activation, inflammatory chemokine production, and allergic sensitization and airway inflammation suggest that PGI2 and its analogue iloprost, both Food and Drug Administration-approved drugs, may be useful in treating allergic diseases and asthma. In addition, inhibiting PGI2 signaling by drugs that either block PGI2 production or restrain IP signaling may augment STAT6-independent pathways of allergic inflammation., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Background: Pharmacologic studies with cyclooxygenase-2 (COX-2) inhibitors suggest that the late phase of ischemic preconditioning (PC) is mediated by COX-2. However, nonspecific effects of COX-2 inhibitors cannot be ruled out, and the selectivity of these inhibitors for COX-2 vs. COX-1 is only relative. Furthermore, the specific prostaglandin (PG) receptors responsible for the salubrious actions of COX-2-derived prostanoids remain unclear., Objective: To determine the role of COX-2 and prostacyclin receptor (IP) in late PC by gene deletion., Methods: COX-2 knockout (KO) mice (COX-2(-/-)), prostacyclin receptor KO (IP(-/-)) mice, and respective wildtype (WT, COX-2(+/+) and IP(+/+)) mice underwent sham surgery or PC with six 4-min coronary occlusion (O)/4-min R cycles 24 h before a 30-min O/24 h R., Results: There were no significant differences in infarct size (IS) between non-preconditioned (non-PC) COX-2(+/+), COX-2(-/-), IP(+/+), and IP(-/-) mice, indicating that neither COX-2 nor IP modulates IS in the absence of PC. When COX-2(-/-) or IP(-/-) mice were preconditioned, IS was not reduced, indicating that the protection of late PC was completely abrogated by deletion of either the COX-2 or the IP gene. Administration of the IP selective antagonist, RO3244794 to C57BL6/J (B6) mice 30 min prior to the 30-min O had no effect on IS. When B6 mice were preconditioned 24 h prior to the 30-min O, IS was markedly reduced; however, the protection of late PC was completely abrogated by pretreatment of RO3244794., Conclusions: This is the first study to demonstrate that targeted disruption of the COX-2 gene completely abrogates the infarct-sparing effect of late PC, and that the IP, downstream of the COX-2/prostanoid pathway, is a key mediator of the late PC. These results provide unequivocal molecular genetic evidence for an essential role of the COX-2/PGI2 receptor axis in the cardioprotection afforded by the late PC.

Cardiac fibrosis is a consequence of many cardiovascular diseases and contributes to impaired ventricular function. Activation of the prostacyclin receptor (IP) protects against cardiac fibrosis, but the molecular mechanisms are not totally understood. Using mouse cardiac fibroblasts, we found that IP activation with cicaprost suppressed expression of collagen I and other target genes of transforming growth factor-beta. This effect of cicaprost was unlikely to be mediated by inhibition of the Smad2/3 or mitogen-activated protein kinase (MAPK) activities, but was associated with cAMP elevation and phosphorylation of the transcription factor cAMP response element binding protein (CREB). Expression of a non-phosphorylated CREB mutant suppressed the inhibitory effect of cicaprost. It appears that phosphorylated CREB binds to and sequestrates the transcription coactivator CBP/p300 from binding to Smad. Inhibition of the intrinsic histone acetyl-transferase activity of CBP/p300 with garcinol significantly suppressed collagen I expression in fibroblasts. Using apolipoprotein E and IP double knockout mouse, we demonstrated that endogenous prostacyclin/IP signaling had an inhibitory effect on angiotensin II-induced cardiac fibrosis under hypercholesterolemic conditions. Taken together, our results suggest that the prostacyclin/IP pathway suppresses cardiac fibrosis, at least partly, by inducing CREB phosphorylation.

Transient global cerebral ischemia causes delayed neuronal death in the hippocampal CA1 region. It also induces an increase in cyclooxygenase 2 (COX-2), which generates several metabolites of arachidonic acid, known as prostanoids, including prostacyclin (PGI(2)). To determine the role of the PGI(2) receptor (IP) in post-ischemic delayed cell death, wild-type and IP knockout (IP(-/-)) C57Bl/6 mice were subjected to 12-min bilateral common carotid artery occlusion or sham surgery, followed by 7 days of reperfusion. In the sham-operated mice, no statistical difference in CA1 hippocampal neuronal density was observed between the wild-type (2836+/-18/mm(2)) and IP(-/-) (2793+/-43/mm(2)) mice. Interestingly, in animals subjected to ischemia, surviving neuronal density in wild-type mice decreased to 50.5+/-7.9% and that of IP(-/-) mice decreased to 23.0+/-4.5% of their respective sham-operated controls (P<0.05). The results establish a role for the IP receptor in protecting pyramidal hippocampal neurons after this global ischemic model and suggest that IP receptor agonists could be developed to prevent delayed pyramidal neuronal cell death.

PGI(2) plays a key role in limiting Th2-mediated airway inflammation. In studies to investigate the mechanism underlying such regulation, we found that the PGI(2) receptor, IP, is preferentially expressed by effector CD4(+) Th2 cells, when compared with Th1 cells. Adoptive transfer of DO11.10 Th2 cells pretreated with PGI(2) resulted in considerably attenuated pulmonary inflammation and airway hyperreactivity in BALB/c recipient mice in response to OVA inhalation. This suppression was independent of increased cAMP levels, because pretreatment of Th2 cells with dibutyryl cAMP before transfer had no effect on airway inflammation. Moreover, PGI(2) pretreatment of Th2 cells suppressed the ability of the cells to infiltrate the lungs but not the spleen. In vitro studies showed that PGI(2) did not affect IL-4 and IL-5 production or the level of IFN-gamma by the T cells. However, the prostanoid strongly inhibited CCL17-induced chemotaxis of CD4(+) Th2 but not Th1 cells. The IP was implicated in this process since migration of wild-type Th2 cells in response to CCL17 was markedly reduced following treatment with PGI(2), whereas IP-deficient Th2 cells were unaffected and migrated effectively. Collectively, these experiments suggest that PGI(2), which is generated by endothelial cells during lung inflammatory response, serves to limit the influx of Th2 cells to the airways. Our results identify PGI(2)-IP as an important pathway for inhibiting allergic pulmonary inflammation by controlling recruitment of CD4(+) Th2 cells into the inflammatory site.

Prostacyclin plays an important cardioprotective role, which has been increasingly appreciated in recent years in light of adverse effects of COX-2 inhibitors in clinical trials. This cardioprotection is thought to be mediated, in part, by prostacyclin inhibition of platelet aggregation. Multiple lines of evidence suggest that prostacyclin additionally protects from cardiovascular disease by pleiotropic effects on vascular smooth muscle. Genetic deletion of the prostacyclin receptor in mice revealed an important role for prostacyclin in preventing the development of atherosclerosis, intimal hyperplasia, and restenosis. In vitro studies have shown these effects may be due to prostacyclin inhibition of vascular smooth muscle cell proliferation and migration. Prostacyclin has also been shown to promote vascular smooth muscle cell differentiation at the level of gene expression through the Gs/cAMP/PKA pathway. Recently identified single nucleotide polymorphisms in the prostacyclin receptor that compromise receptor function suggest that some genetic variations may predispose individuals to increased cardiovascular disease. Herein, we review the literature on the cardioprotective effects of prostacyclin on vascular smooth muscle, and the underlying molecular signaling mechanisms. Understanding the role of prostacyclin and other eicosanoid mediators in the vasculature may lead to improved therapeutic and preventative options for cardiovascular disease.

Background: In the heart, the expressions of several types of prostanoid receptors have been reported. However, their roles in cardiac hypertrophy in vivo remain unknown. We intended to clarify the roles of these receptors in pressure overload-induced cardiac hypertrophy using mice lacking each of their receptors., Methods and Results: We used a model of pressure overload-induced cardiac hypertrophy produced by banding of the transverse aorta in female mice. In wild-type mice subjected to the banding, cardiac hypertrophy developed during the observation period of 8 weeks. In mice lacking the prostaglandin (PG) I2 receptor (IP(-/-)), however, cardiac hypertrophy and cardiomyocyte hypertrophy were significantly greater than in wild-type mice at 2 and 4 weeks but not at 8 weeks, whereas there was no such augmentation in mice lacking the prostanoid receptors other than IP. In addition, cardiac fibrosis observed in wild-type hearts was augmented in IP(-/-) hearts, which persisted for up to 8 weeks. In IP(-/-) hearts, the expression level of mRNA for atrial natriuretic peptide, a representative marker of cardiac hypertrophy, was significantly higher than in wild-type hearts. In vitro, cicaprost, an IP agonist, reduced platelet-derived growth factor-induced proliferation of wild-type noncardiomyocytes, although it could not inhibit cardiotrophin-1-induced hypertrophy of cardiomyocytes. Accordingly, cicaprost increased cAMP concentration efficiently in noncardiomyocytes., Conclusions: IP plays a suppressive role in the development of pressure overload-induced cardiac hypertrophy via the inhibition of both cardiomyocyte hypertrophy and cardiac fibrosis. Both effects have been suggested as originating from the action on noncardiomyocytes rather than cardiomyocytes.

Background: This study examined whether targeted disruption of the genes for the prostacyclin receptor (IP) or the thromboxane A2 receptor (TP) confers a susceptibility to salt-dependent hypertension., Methods and Results: Eight female IP- or TP-deficient mice were examined. Baseline systolic blood pressure (SBP) did not differ between TP(-/-) and TP(+/+), but was significantly lower in the IP(-/-) group than in the IP(+/+). With a high salt diet, SBP in IP(-/-) gradually increased. In contrast, SBP in the IP(+/+), TP(-/-), or TP(+/+) groups remained unchanged., Conclusions: The prostacyclin receptor may participate in the maintenance of baseline BP. With salt loading, BP adaptation may take place, at least in part, via IP mediated signals.

Production of thromboxane (TX) A2 and PG I2/prostacyclin (PGI2) is increased in patients with atherosclerosis. However, their roles in atherogenesis have not been critically defined. To examine this issue, we cross-bred atherosclerosis-prone apoE-deficient mice with mice deficient in either the TXA receptor (TP) or the PGI receptor (IP). Although they showed levels of serum cholesterol and triglyceride similar to those of apoE-deficient mice, apoE-/-TP-/- mice exhibited a significant delay in atherogenesis, and apoE-/-IP-/- mice exhibited a significant acceleration in atherogenesis compared with mice deficient in apoE alone. The plaques in apoE-/-IP-/- mice showed partial endothelial disruption and exhibited enhanced expression of ICAM-1 and decreased expression of platelet endothelial cell adhesion molecule 1 (PECAM-1) in the overlying endothelial cells compared with those of apoE-/-TP-/- mice. Platelet activation with thrombin ex vivo revealed higher and lower sensitivity for surface P-selectin expression in platelets of apoE-/-IP-/- and apoE-/-TP-/- mice, respectively, than in those of apoE-/- mice. Intravital microscopy of the common carotid artery revealed a significantly greater number of leukocytes rolling on the vessel walls in apoE-/-IP-/- mice than in either apoE-/-TP-/- or apoE-/- mice. We conclude that TXA2 promotes and PGI2 prevents the initiation and progression of atherogenesis through control of platelet activation and leukocyte-endothelial cell interaction.

Persistent reduction of renal perfusion pressure induces renovascular hypertension by activating the renin-angiotensin-aldosterone system; however, the sensing mechanism remains elusive. Here we investigated the role of PGI2 in renovascular hypertension in vivo, employing mice lacking the PGI2 receptor (IP-/- mice). In WT mice with a two-kidney, one-clip model of renovascular hypertension, the BP was significantly elevated. The increase in BP in IP-/- mice, however, was significantly lower than that in WT mice. Similarly, the increases in plasma renin activity, renal renin mRNA, and plasma aldosterone in response to renal artery stenosis were all significantly lower in IP-/- mice than in WT mice. All these parameters were measured in mice lacking the four PGE2 receptor subtypes individually, and we found that these mice had similar responses to WT mice. PGI2 is produced by COX-2 and a selective inhibitor of this enzyme, SC-58125, also significantly reduced the increases in plasma renin activity and renin mRNA expression in WT mice with renal artery stenosis, but these effects were absent in IP-/- mice. When the renin-angiotensin-aldosterone system was activated by salt depletion, SC-58125 blunted the response in WT mice but not in IP-/- mice. These results indicate that PGI2 derived from COX-2 plays a critical role in regulating the release of renin and consequently renovascular hypertension in vivo.