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1.
PGE(2) is an important cyclooxygenase product that modulates airway inflammatory and smooth muscle responses. Signal transduction is mediated by four EP receptor subtypes that cause distinct effects on cell metabolism. To determine the role of EP(2) receptor activation, we produced a mouse lacking the EP(2) receptor by targeted gene disruption. The effect of aerosolized PGE(2) and other agonists was measured using barometric plethysmography and by measurements of lung resistance in mechanically ventilated mice. Inhalation of PGE(2) inhibited methacholine responses in wild-type but not in mice lacking the EP(2) receptor [EP(2)(-/-)]. After airway constriction was induced by methacholine aerosol, PGE(2) reduced the airway constriction enhanced pause in wild-type mice (from 0.88 +/- 0.15 to 0.55 +/- 0.06) but increased it in EP(2)(-/-) mice (from 0.73 +/- 0. 08 to 1.27 +/- 0.19). Similar results were obtained in mechanically ventilated mice. These data indicate that the EP(2) receptor mediates the bronchodilation effect of PGE(2).  相似文献   

2.
The effects of PGE(2) on longitudinal smooth muscle, the intracellular mechanisms involved, and the localization of EP receptors were investigated in rabbit small intestine. PGE(2) evoked contractions in small intestine that were reduced by tetrodotoxin and hexamethonium. 17-Phenyl trinor PGE(2), sulprostone, misoprostol and 16,16-dimethyl PGE(2) evoked contractions. Butaprost did not modify spontaneous motility. AH 6809 reduced PGE(2) and 17-phenyl trinor PGE(2)-induced contractions. Verapamil, Ca(2+) free medium, staurosporine, forskolin, theophylline, and rolipram diminished, while IP-20 and H-89 increased PGE(2)-induced contractions. Western blot analysis showed protein bands of 41kDa for EP(1), 71kDa for EP(2) and 62kDa for EP(3) receptors. EP(1), EP(2) and EP(3) receptors were detected in neurons of the myenteric and submucosal ganglia, but only EP(3) receptors were found in smooth muscle layers. This study did not detect EP(4) receptor. PGE(2)-induced contractions would be mediated through EP(1) and EP(3) receptors, and voltage-dependent Ca(2+) channels, protein kinase C, and cAMP would be implicated in these responses.  相似文献   

3.
Protease-activated receptors are G protein-coupled receptors activated by serine-proteases. Protease-activated receptor 2 is involved in the regulation of airway smooth muscle tone but its effects vary according to species and experimental conditions. We determined the effects of protease-activated receptor 2 activation on smooth muscle tone and airway reactivity to histamine in guinea pigs and smoking or non-smoking humans. The effects of trypsin and protease-activated receptor activating peptide on the isometric tension and response to histamine of guinea pig tracheal and human bronchial rings were studied. Human tissues were obtained from 6 smokers and 4 non-smokers. We assessed the effects of epithelial removal, inhibitors of cyclooxygenases, nitric oxide synthases, neutral endopeptidase and antagonists of acetylcholine, histamine, bradykinin and tachykinin receptors. Bronchomotor responses to protease-activated receptor 2 activation were variable in guinea pig, in half of animals PAR2 activation induced smooth muscle relaxation through the epithelial release of prostanoids but not of nitric oxide. In human airways, protease-activated receptor 2 activation reduced responsiveness to histamine in bronchial rings from smokers but increased responsiveness in bronchi from non-smokers. This study demonstrates an influence of tobacco smoking on the effect of protease-activated receptor 2 activation on airway responsiveness in humans, with an increased protection against histamine-induced contractions, probably through an increased epithelial release of prostanoids. The role of airway protease-activated receptor 2 may be to maintain smooth muscle tone homeostasis.  相似文献   

4.
Isoprostanes are generated during periods of oxidative stress, which characterize diseases such as asthma and cystic fibrosis. They also elicit functional responses and may therefore contribute to the pathology of these diseases. We set out to examine the effects of isoprostanes on airway responsiveness to cholinergic stimulation. Muscle bath techniques were employed using isolated bovine tracheal smooth muscle. 8-Isoprostaglandin E2 (8-iso-PGE2) increased tone directly on its own, although the magnitude of this response, even at the highest concentration tested, was only a fraction of that evoked by KCl or carbachol. More importantly, though, pretreatment of the tissues with 8-iso-PGE2 (10 microM) markedly augmented responses to submaximal and even subthreshold concentrations of KCl, carbachol, or histamine, whereas maximal responses to these agents were unaffected by the isoprostane. The augmentative effect on cholinergic responsiveness was mimicked by PGE2 (0.1 microM) and by the FP agonists PGF2 (0.1 microM) and fluprostenol (0.1 microM), but not by the EP3 agonist sulprostone (0.1 microM) or the TP agonist U-46619 (0.1 microM). Antagonists of EP1 receptors (AH-6809 and SC-19920, 10 microM) and TP receptors (ICI-192605, 1 microM) had no effect on 8-iso-PGE2-induced augmentation of cholinergic responsiveness. We conclude that 8-iso-PGE2 induces nonspecific airway smooth muscle hyperresponsiveness through a non-TP non-EP prostanoid receptor.  相似文献   

5.
Although G protein-coupled receptor (GPCR) kinases (GRKs) have been shown to mediate desensitization of numerous GPCRs in studies using cellular expression systems, their function under physiological conditions is less well understood. In the current study, we employed various strategies to assess the effect of inhibiting endogenous GRK2/3 on signaling and function of endogenously expressed G s-coupled receptors in human airway smooth muscle (ASM) cells. GRK2/3 inhibition by expression of a Gbetagamma sequestrant, a GRK2/3 dominant-negative mutant, or siRNA-mediated knockdown increased intracellular cAMP accumulation mediated via beta-agonist stimulation of the beta-2-adrenergic receptor (beta 2AR). Conversely, neither 5'-( N-ethylcarboxamido)-adenosine (NECA; activating the A2b adenosine receptor) nor prostaglandin E2 (PGE 2; activating EP2 or EP4 receptors)-stimulated cAMP was significantly increased by GRK2/3 inhibition. Selective knockdown using siRNA suggested the majority of PGE 2-stimulated cAMP in ASM was mediated by the EP2 receptor. Although a minor role for EP3 receptors in influencing PGE 2-mediated cAMP was determined, the GRK2/3-resistant nature of EP2 receptor signaling in ASM was confirmed using the EP2-selective agonist butaprost. Somewhat surprisingly, GRK2/3 inhibition did not augment the inhibitory effect of the beta-agonist on mitogen-stimulated increases in ASM growth. These findings demonstrate that with respect to G s-coupled receptors in ASM, GRK2/3 selectively attenuates beta 2AR signaling, yet relief of GRK2/3-dependent beta 2AR desensitization does not influence at least one important physiological function of the receptor.  相似文献   

6.
The ductus arteriosus (DA) is a fetal shunt that directs right ventricular outflow away from pulmonary circulation and into the aorta. Critical roles for prostaglandin E(2) (PGE(2)) and the EP4 receptor (EP4) have been established in maintaining both the patency of the vessel in utero and in its closure at birth. Here we have generated mice in which loss of EP4 expression is limited to either the smooth muscle (SMC) or endothelial cells and demonstrated that SMC, but not endothelial cell expression of EP4 is required for DA closure. The genome wide expression analysis of full term wild type and EP4(-/-) DA indicates that PGE(2)/EP4 signaling modulates expression of a number of unique pathways, including those involved in SMC proliferation, cell migration, and vascular tone. Together this supports a mechanism by which maturation and increased contractility of the vessel is coupled to the potent smooth muscle dilatory actions of PGE(2).  相似文献   

7.
Using muscle bath techniques, we examined the inhibitory activities of several E- and F-ring isoprostanes in canine and porcine airway smooth muscle. 8-Isoprostaglandin E1 and 8-isoprostaglandin E2 (8-iso PGE2) reversed cholinergic tone in a concentration-dependent manner, whereas the F-ring isoprostanes were ineffective. Desensitization with 8-iso-PGE2 and PGE2 implicated isoprostane activity at the PGE2 receptor (EP). We found that the inhibitory E-ring isoprostane responses were significantly augmented by rolipram (a type IV phosphodiesterase inhibitor), while 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (a guanylate cyclase inhibitor) had no effect, suggesting a role for cAMP in isoprostane-mediated relaxations. 8-Iso-PGE2 did not reverse KCl tone, suggesting that voltage-dependent Ca2+ influx and myosin light chain kinase are not suppressed by isoprostanes. Patch-clamp studies showed marked suppression of K+ currents by 8-iso-PGE2. We conclude that E-ring isoprostanes exert PGE2 receptor-directed, cAMP-dependent relaxations in canine and porcine airway smooth muscle. This activity is not dependent on K+ channel activation or the direct inhibition of voltage-operated Ca2+ influx or myosin light chain kinase.  相似文献   

8.
Beta2AR desensitization in airway smooth muscle (ASM) mediated by airway inflammation has been proposed to contribute to asthma pathogenesis and diminished efficacy of beta-agonist therapy. Mechanistic insight into this phenomenon is largely conceptual and lacks direct empirical evidence. Here, we employ molecular and genetic strategies to reveal mechanisms mediating cytokine effects on ASM beta2AR responsiveness. Ectopic expression of inhibitory peptide (PKI-GFP) or a mutant regulatory subunit of PKA (RevAB-GFP) effectively inhibited intracellular PKA activity in cultured human ASM cells and enhanced beta2AR responsiveness by mitigating both agonist-specific (beta-agonist-mediated) desensitization and cytokine (IL-1beta and TNF-alpha)-induced heterologous desensitization via actions on multiple targets. In the absence of cytokine treatment, PKA inhibition increased beta2AR-mediated signaling by increasing both beta2AR-G protein coupling and intrinsic adenylyl cyclase activity. PKI-GFP and RevAB-GFP expression also conferred resistance to cytokine-promoted beta2AR-G protein uncoupling and disrupted feed-forward mechanisms of PKA activation by attenuating the induction of COX-2 and PGE2. Cytokine treatment of tracheal ring preparations from wild-type mice resulted in a profound loss of beta-agonist-mediated relaxation of methacholine-contracted rings, whereas rings from EP2 receptor knockout mice were largely resistant to cytokine-mediated beta2AR desensitization. These findings identify EP2 receptor- and PKA-dependent mechanisms as the principal effectors of cytokine-mediated beta2AR desensitization in ASM.  相似文献   

9.
Mast cells are implicated in the pathogenesis of a broad spectrum of immunological disorders. These cells release inflammatory mediators in response to a number of stimuli, including IgE-Ag complexes. The degranulation of mast cells is modified by PGs. To begin to delineate the pathway(s) used by PGs to regulate mast cell function, we examined bone marrow-derived mast cells (BMMC) cultured from mice deficient in the EP(1), EP(2), EP(3), and EP(4) receptors for PGE(2). Although BMMCs express all four of these PGE(2) receptors, potentiation of Ag-stimulated degranulation and IL-6 cytokine production by PGE(2) is dependent on the EP(3) receptor. Consistent with the coupling of this receptor to G(alphai), PGE(2) activation of the EP(3) receptor leads to both inhibition of adenylate cyclase and increased intracellular Ca(2+). The magnitude of increase in intracellular Ca(2+) induced by EP(3) activation is similar to that observed after activation of cells with IgE and Ag. Although PGE alone is not sufficient to initiate BMMC degranulation, stimulation of cells with PGE along with PMA induces degranulation. These actions are mediated by the EP(3) receptor through signals involving Ca(2+) mobilization and/or decreased cAMP levels. Accordingly, these studies identify PGE(2)/EP(3) as a proinflammatory signaling pathway that promotes mast cell activation.  相似文献   

10.
We found that centrally administered prostaglandin (PG) E(2) exhibited anxiolytic-like activity in the elevated plus-maze and open field test in mice. Agonists selective for EP(1) and EP(4) receptors, among four receptor subtypes for PGE(2), mimicked the anxiolytic-like activity of PGE(2). The anxiolytic-like activity of PGE(2) was blocked by an EP(1) or EP(4) antagonist, as well as in EP(4) but not EP(1) knockout mice. Central activation of either EP(1) or EP(4) receptors resulted in anxiolytic-like activity. The PGE(2)-induced anxiolytic-like activity was inhibited by antagonists for serotonin 5-HT(1A), dopamine D(1) and GABA(A) receptors. Taken together, PGE(2) exhibits anxiolytic-like activity via EP(1) and EP(4) receptors, with downstream involvement of 5-HT(1A), D(1) and GABA(A) receptor systems.  相似文献   

11.
Prostaglandin E receptors   总被引:14,自引:0,他引:14  
Prostaglandin (PG) E(2) exerts its actions by acting on a group of G-protein-coupled receptors (GPCRs). There are four GPCRs responding to PGE(2) designated subtypes EP1, EP2, EP3, and EP4 and multiple splicing isoforms of the subtype EP3. The EP subtypes exhibit differences in signal transduction, tissue localization, and regulation of expression. This molecular and biochemical heterogeneity of PGE receptors leads to PGE(2) being the most versatile prostanoid. Studies on knock-out mice deficient in each EP subtype have defined PGE(2) actions mediated by each subtype and identified the role each EP subtype plays in various physiological and pathophysiological responses. Here we review recent advances in PGE receptor research.  相似文献   

12.
In this study, we examined effects of chemical stimulation of the ventrolateral region of the midbrain periaqueductal gray (vl PAG) on airway smooth muscle tone. We observed that in anesthetized, paralyzed, and artificially ventilated ferrets, vl PAG stimulation elicited airway smooth muscle relaxation. To clarify the mechanisms underlying this observation, we examined the GABA-GABA(A) receptor signaling pathway by 1) examining the expression of GABA(A) receptors on airway-related vagal preganglionic neurons (AVPNs) located in the rostral nucleus ambiguus region (rNA), by use of receptor immunochemistry and confocal microscopy; 2) measuring GABA release within the rNA by using microdialysis; and 3) performing physiological experiments to determine the effects of selective blockade of GABA(A) receptors expressed by AVPNs in the rNA region on vl PAG-induced airway relaxation, thereby defining the role of the GABA(A) receptor subtype in this process. We observed that AVPNs located in the rNA region do express the GABA(A) receptor beta-subtype. In addition, we demonstrated that activation of vl PAG induced GABA release within the rNA region, and this release was associated with airway smooth muscle relaxation. Blockade of the GABA(A) receptor subtype expressed by AVPNs in the rNA by bicuculline diminished the inhibitory effects of vl PAG stimulation on airway smooth muscle tone. These data indicate, for the first time, that activation of vl PAG dilates the airways by a release of GABA and activation of GABA(A) receptors expressed by AVPNs.  相似文献   

13.
Prostaglandin E2 (PGE2) mediates the organization of male rat sexual behavior and medial preoptic area (MPOA) neuroanatomy during a sensitive perinatal window. PGE2 is up-regulated in response to estradiol, and initiates a two-fold increase in dendritic spines densities on neurons. All the four receptors for PGE2 and EP1-4 are present in developing POA, a critical region controlling male sexual behavior. Previous studies explored that EP receptors are involved in PGE2-induction of neonatal levels of spinophilin protein, a surrogate marker for dendritic spine formation, but did not assess behavioral masculinization. Here, we used two approaches, suppression of EP receptor expression with antisense oligonucleotides and activation of EP receptors with selective agonists, to test which receptors are necessary and sufficient, respectively, for the effects of PGE2 on behavior and neuronal morphology. In female rats, neonatal treatment with antisense oligonucleotides against EP2 or EP4 but not EP1 or EP3 completely prevented the expression of adult behavior organized by PGE2 exposure. The effects of ONO-DI-004, ONO-AE-259-01, ONO-AE-248, and ONO-AE1-329 (EP1-4 agonists respectively) were equivalent to PGE2 treatment, which suggests activating any EP receptor neonatally suffices in masculinizing sex behavior. When given alone, not all EP agonists increased neonatal POA spinophilin levels; yet giving each agonist neonatally increased adult levels. Moreover, adult spinophilin levels significantly correlated with two measures of male sexual behavior. The body of evidence suggests that EP2 and EP4 are both necessary and sufficient for PGE2-induced masculinization of sex behavior, whereas EP1 and EP3 provide redundant roles.  相似文献   

14.
PGs are derived from arachidonic acid by PG-endoperoxide synthase (PTGS)-1 and PTGS2. Although enhanced levels of PGs are present during acute and chronic inflammation, a functional role for prostanoids in inflammation has not been clearly defined. Using a series of genetically engineered mice, we find that PTGS1 has the capacity to induce acute inflammation, but PTGS2 has negligible effects on the initiation of this response. Furthermore, we show that the contribution of PTGS1 is mediated by PGE(2) acting through the E-prostanoid (EP)3 receptor. Moreover, in the absence of EP3 receptors, inflammation is markedly attenuated, and the addition of nonsteroidal anti-inflammatory agents does not further impair the response. These studies demonstrate that PGE(2) promotes acute inflammation by activating EP3 receptors and suggest that EP3 receptors may be useful targets for anti-inflammatory therapy.  相似文献   

15.
The prostanoid receptors on human airway smooth muscle cells (HASMC) that augment the release by IL-1beta of granulocyte colony-stimulating factor (G-CSF) have been characterized and the signaling pathway elucidated. PCR of HASM cDNA identified products corresponding to EP(2), EP(3), and EP(4) receptor subtypes. These findings were corroborated at the protein level by immunocytochemistry. IL-1beta promoted the elaboration of G-CSF, which was augmented by PGE(2). Cicaprost (IP receptor agonist) was approximately equiactive with PGE(2), whereas PGD(2), PGF(2alpha), and U-46619 (TP receptor agonist) were over 10-fold less potent. Neither SQ 29,548 nor BW A868C (TP and DP(1) receptor antagonists, respectively) attenuated the enhancement of G-CSF release evoking any of the prostanoids studied. With respect to PGE(2), the EP receptor agonists 16,16-dimethyl PGE(2) (nonselective), misoprostol (EP(2)/EP(3) selective), 17-phenyl-omega-trinor PGE(2) (EP(1) selective), ONO-AE1-259, and butaprost (both EP(2) selective) were full agonists at enhancing G-CSF release. AH 6809 (10 microM) and L-161,982 (2 microM), which can be used in HASMC as selective EP(2) and EP(4) receptor antagonists, respectively, failed to displace to the right the PGE(2) concentration-response curve that described the augmented G-CSF release. In contrast, AH 6809 and L-161,982 in combination competitively antagonized PGE(2)-induced G-CSF release. Augmentation of G-CSF release by PGE(2) was mimicked by 8-BrcAMP and abolished in cells infected with an adenovirus vector encoding an inhibitor protein of cAMP-dependent protein kinase (PKA). These data demonstrate that PGE(2) facilitates G-CSF secretion from HASMC through a PKA-dependent mechanism by acting through EP(2) and EP(4) prostanoid receptors and that effective antagonism is realized only when both subtypes are blocked concurrently.  相似文献   

16.
17.
18.
The expression of cyclooxygenase-2 (COX-2) and the synthesis of prostaglandin E2 (PGE2) as well as of cytokines such as interleukin-6 (IL-6) have all been suggested to propagate neuropathology in different brain disorders such as HIV-dementia, prion diseases, stroke and Alzheimer's disease. In this report, we show that PGE2-stimulated IL-6 release in U373 MG human astroglioma cells and primary rat astrocytes. PGE2-induced intracellular cAMP formation was mediated via prostaglandin E receptor 2 (EP2), but inhibition of cAMP formation and protein kinase A or blockade of EP1/EP2 receptors did not affect PGE2-induced IL-6 synthesis. This indicates that the cAMP pathway is not part of PGE2-induced signal transduction cascade leading to IL-6 release. The EP3/EP1-receptor agonist sulprostone failed to induce IL-6 release, suggesting an involvement of EP4-like receptors. PGE2-activated p38 mitogen-activated kinase (p38 MAPK) and protein kinase C (PKC). PGE2-induced IL-6 synthesis was inhibited by specific inhibitors of p38 MAPK (SB202190) and PKC (GF203190X). Although, up to now, EP receptors have only rarely been linked to p38 MAPK or PKC activation, these results suggest that PGE2 induces IL-6 via an EP4-like receptor by the activation of PKC and p38 MAPK via an EP4-like receptor independently of cAMP.  相似文献   

19.
G protein-coupled receptors (GPCRs) transduce extracellular signals into intracellular events. The waning responsiveness of GPCRs in the face of persistent agonist stimulation, or desensitization, is a necessary event that ensures physiological homeostasis. GPCR kinases (GRKs) are important regulators of GPCR desensitization. GRK5, one member of the GRK family, desensitizes central M(2) muscarinic receptors in mice. We questioned whether GRK5 might also be an important regulator of peripheral muscarinic receptor responsiveness in the cardiopulmonary system. Specifically, we wanted to determine the role of GRK5 in regulating muscarinic receptor-mediated control of airway smooth muscle tone or regulation of cholinergic-induced bradycardia. Tracheal pressure, heart rate, and tracheal smooth muscle tension were measured in mice having a targeted deletion of the GRK5 gene (GRK5(-/-)) and littermate wild-type (WT) control mice. Both in vivo and in vitro results showed that the airway contractile response to a muscarinic receptor agonist was not different between GRK5(-/-) and WT mice. However, the relaxation component of bilateral vagal stimulation and the airway smooth muscle relaxation resulting from beta(2)-adrenergic receptor activation were diminished in GRK5(-/-) mice. These data suggest that M(2) muscarinic receptor-mediated opposition of airway smooth muscle relaxation is regulated by GRK5 and is, therefore, excessive in GRK5(-/-) mice. In addition, this study shows that GRK5 regulates pulmonary responses in a tissue- and receptor-specific manner but does not regulate peripheral cardiac muscarinic receptors. GRK5 regulation of airway responses may have implications in obstructive airway diseases such as asthma or chronic obstructive pulmonary disease.  相似文献   

20.
The kidney plays a central role in long-term regulation of arterial blood pressure and salt and water homeostasis. This is achieved in part by the local actions of paracrine and autacoid mediators such as the arachidonic acid-prostanoid system. The present study tested the role of specific PGE(2) E-prostanoid (EP) receptors in the regulation of renal hemodynamics and vascular reactivity to PGE(2). Specifically, we determined the extent to which the EP(2) and EP(3) receptor subtypes mediate the actions of PGE(2) on renal vascular tone. Renal blood flow (RBF) was measured by ultrasonic flowmetry, whereas vasoactive agents were injected directly into the renal artery of male mice. Studies were performed on two independent mouse lines lacking either EP(2) or EP(3) (-/-) receptors and the results were compared with wild-type controls (+/+). Our results do not support a unique role of the EP(2) receptor in regulating overall renal hemodynamics. Baseline renal hemodynamics in EP(2)-/- mice [RBF EP(2)-/-: 5.3 +/- 0.8 ml. min(-1). 100 g kidney wt(-1); renal vascular resistance (RVR) 19.7 +/- 3.6 mmHg. ml(-1). min. g kidney wt] did not differ statistically from control mice (RBF +/+: 4.0 +/- 0.5 ml. min(-1). 100 g kidney wt(-1); RVR +/+: 25.4 +/- 4.9 mmHg. ml(-1). min. 100 g kidney wt(-1)). This was also the case for the peak RBF increase after local PGE(2) (500 ng) injection into the renal artery (EP(2)-/-: 116 +/- 4 vs. +/+: 112 +/- 2% baseline RBF). In contrast, we found that the absence of EP(3) receptors in EP(3)-/- mice caused a significant increase (43%) in basal RBF (7.9 +/- 0.8 ml. min(-1). g kidney wt(-1), P < 0.05 vs. +/+) and a significant decrease (41%) in resting RVR (11.6 +/- 1.4 mmHg. ml(-1). min. g kidney wt(-1), P < 0.05 vs. +/+). Local administration of 500 ng of PGE(2) into the renal artery caused more pronounced renal vasodilation in EP(3)-/- mice (128 +/- 2% of basal RBF, P < 0.05 vs. +/+). We conclude that EP(3 )receptors mediate vasoconstriction in the kidney of male mice and its actions are tonically active in the basal state. Furthermore, EP(3) receptors are capable of buffering PGE(2)-mediated renal vasodilation.  相似文献   

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