Cyclooxygenase-2 and Prostaglandin E2 Signaling through Prostaglandin Receptor EP-2 Favor the Development of Myocarditis during Acute Trypanosoma cruzi Infection.
ABSTRACT: Inflammation plays an important role in the pathophysiology of Chagas disease, caused by Trypanosoma cruzi. Prostanoids are regulators of homeostasis and inflammation and are produced mainly by myeloid cells, being cyclooxygenases, COX-1 and COX-2, the key enzymes in their biosynthesis from arachidonic acid (AA). Here, we have investigated the expression of enzymes involved in AA metabolism during T. cruzi infection. Our results show an increase in the expression of several of these enzymes in acute T. cruzi infected heart. Interestingly, COX-2 was expressed by CD68+ myeloid heart-infiltrating cells. In addition, infiltrating myeloid CD11b+Ly6G- cells purified from infected heart tissue express COX-2 and produce prostaglandin E2 (PGE2) ex vivo. T. cruzi infections in COX-2 or PGE2-dependent prostaglandin receptor EP-2 deficient mice indicate that both, COX-2 and EP-2 signaling contribute significantly to the heart leukocyte infiltration and to the release of chemokines and inflammatory cytokines in the heart of T. cruzi infected mice. In conclusion, COX-2 plays a detrimental role in acute Chagas disease myocarditis and points to COX-2 as a potential target for immune intervention.
Project description:BACKGROUND:Accumulated evidence suggests that spinal cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) may be implicated in the development of opioid-induced hyperalgesia. METHODS:Rats received subcutaneous fentanyl injections at different doses (20-80 ?g kg-1), four separate times at 15-min intervals. Some rats only received fentanyl (60 ?g kg-1 × 4 doses) with or without surgical incision. Fentanyl-induced hyperalgesia was evaluated via a tail-pressure or paw-withdrawal test. The concentrations of spinal COX-2, EP-1 receptor (EP-1R) mRNA, and PGE2 were measured. The effects of the COX-2 inhibitor, parecoxib (intraperitoneal 10 mg kg-1), or the EP-1R antagonist, SC51089 (intraperitoneal 100 ?g kg-1), on hyperalgesia and spinal PGE2 were examined. RESULTS:Acute repeated injections of fentanyl dose-dependently induced mechanical hyperalgesia, which reached a peak at the 1st day and persisted for 1-4 days postinjection. This hyperalgesia could be partly or totally prevented by the pretreatment of either parecoxib or SC51089. Consistently, the levels of spinal COX-2 mRNA and PGE2 were also dose-dependently increased, reaching a peak at the first day and persisting for 2 days postinjection. Pretreatment with parecoxib could block the increase in spinal PGE2 and had no effects on spinal COX-2 and EP-1R mRNA. Fentanyl injection enhanced incision-induced mechanical and thermal hyperalgesia. CONCLUSIONS:Acute repeated fentanyl administration dose-dependently produced mechanical hyperalgesia and augmented surgery induced postoperative hyperalgesia. This behavioural change was paralleled with an increase in spinal COX-2 mRNA and PGE2 after fentanyl administration. Inhibition of COX-2 or blockade of EP-1R can partly or totally prevent hyperalgesia.
Project description:To investigate the effect of COX-2 polymorphism and its product, prostaglandin E2 (PGE2), on stroke risk in an endemic area for Chagas disease. In a separate cohort, to investigate the effect of COX-2 polymorphisms on the total burden of cerebral white matter disease.Cases were outpatients with ischemic stroke; controls were stroke-free subjects from 2 outpatient clinics (heart failure and caregivers of a movement disorders clinic). We extracted DNA from total blood to investigate the rs20417 COX-2 polymorphism. Serologic tests (Enzime-linked immunosorbent assay) were performed to confirm Trypanosoma cruzi infection and to quantify PGE2 levels. In the Boston cohort, white matter hyperintensity volume (WMHv) was quantified on the admission brain magnetic resonance images of subjects with ischemic stroke, who also donated DNA for the COX-2 gene region analysis.We studied 44 patients with stroke and 96 controls (46 with heart failure and 50 caregivers) in the Brazilian cohort; and 788 stroke patients (302 cardioembolic and 486 noncardioembolic) in the Boston cohort. In the Brazilian cohort, rs20417 polymorphism was associated with both stroke (P = 5 × 10(-6)) and decreased PGE2 levels (P = 4 × 10(-5)); similarly, Chagas was associated with stroke (P = 4 × 10(-3)) and decreased PGE2 levels (P = 7 × 10(-3)). In the Boston cohort, rs20417 polymorphism was associated with increased WMHv among noncardioembolic (P = .037), but not among cardioembolic stroke patients.Variation in COX-2 gene is associated with both symptomatic and silent brain cerebrovascular disease. This candidate gene region should be tested in population-based samples.
Project description:Prostaglandin E2 (PGE2) plays an important role in bone development and metabolism. To interfere therapeutically in the PGE2 pathway, however, knowledge about the involved enzymes (cyclooxygenases) and receptors (PGE2 receptors) is essential. We therefore examined the production of PGE2 in cultured growth plate chondrocytes in vitro and the effects of exogenously added PGE2 on cell proliferation. Furthermore, we analysed the expression and spatial distribution of cyclooxygenase (COX)-1 and COX-2 and PGE2 receptor types EP1, EP2, EP3 and EP4 in the growth plate in situ and in vitro. PGE2 synthesis was determined by mass spectrometry, cell proliferation by DNA [3H]-thymidine incorporation, mRNA expression of cyclooxygenases and EP receptors by RT-PCR on cultured cells and in homogenized growth plates. To determine cellular expression, frozen sections of rat tibial growth plate and primary chondrocyte cultures were stained using immunohistochemistry with polyclonal antibodies directed towards COX-1, COX-2, EP1, EP2, EP3, and EP4. Cultured growth plate chondrocytes transiently secreted PGE2 into the culture medium. Although both enzymes were expressed in chondrocytes in vitro and in vivo, it appears that mainly COX-2 contributed to PGE2-dependent proliferation. Exogenously added PGE2 stimulated DNA synthesis in a dose-dependent fashion and gave a bell-shaped curve with a maximum at 10-8 M. The EP1/EP3 specific agonist sulprostone and the EP1-selective agonist ONO-D1-004 increased DNA synthesis. The effect of PGE2 was suppressed by ONO-8711. The expression of EP1, EP2, EP3, and EP4 receptors in situ and in vitro was observed; EP2 was homogenously expressed in all zones of the growth plate in situ, whereas EP1 expression was inhomogenous, with spared cells in the reserve zone. In cultured cells these four receptors were expressed in a subset of cells only. The most intense staining for the EP1 receptor was found in polygonal cells surrounded by matrix. Expression of receptor protein for EP3 and EP4 was observed also in rat growth plates. In cultured chrondrocytes, however, only weak expression of EP3 and EP4 receptor was detected. We suggest that in growth plate chondrocytes, COX-2 is responsible for PGE2 release, which stimulates cell proliferation via the EP1 receptor.
Project description:Prostaglandin (PG) E2, a major product of cyclooxygenase (COX)-2, acts as an immunomodulator at the maternal-fetal interface during pregnancy. It exerts biologic function through interaction with E-prostanoid (EP) receptors localized to the placenta. The activation of the COX-2/PGE2/EP signal pathway can alter the expression of the ATP-binding cassette (ABC) transporters, multidrug resistance protein 1 [P-glycoprotein (Pgp); gene: ABCB1], and breast cancer resistance protein (BCRP; gene: ABCG2), which function to extrude drugs and xenobiotics from cells. In the placenta, PGE2-mediated changes in ABC transporter expression could impact fetal drug exposure. Furthermore, understanding the signaling cascades involved could lead to strategies for the control of Pgp and BCRP expression levels. We sought to determine the impact of PGE2 signaling mechanisms on Pgp and BCRP in human placental cells. The treatment of placental cells with PGE2 up-regulated BCRP expression and resulted in decreased cellular accumulation of the fluorescent substrate Hoechst 33342. Inhibiting the EP1 and EP3 receptors with specific antagonists attenuated the increase in BCRP. EP receptor signaling results in activation of transcription factors, which can affect BCRP expression. Although PGE2 decreased nuclear factor ?-light chain-enhancer of activated B activation and increased activator protein 1, chemical inhibition of these inflammatory transcription factors did not blunt BCRP up-regulation by PGE2. Though PGE2 decreased Pgp mRNA, Pgp expression and function were not significantly altered. Overall, these findings suggest a possible role for PGE2 in the up-regulation of placental BCRP expression via EP1 and EP3 receptor signaling cascades.
Project description:Pancreatic stellate cells are source of dense fibrotic stroma, a constant pathological feature of chronic pancreatitis and pancreatic adenocarcinoma. We observed correlation between levels of cyclooxygenase 2 (COX-2) and its product prostaglandin E2 (PGE2) and the extent of pancreatic fibrosis. The aims of this study were to delineate the effects of PGE2 on immortalized human pancreatic stellate cells (HPSCs) and to identify the receptor involved.Immunohistochemistry, reverse transcription-polymerase chain reaction and quantitative reverse transcription-polymerase chain reaction were used to assess COX-2, extracellular matrix, and matrix metalloproteinase gene expression. Eicosanoid profile was determined by liquid chromatography-tandem mass spectrometry. Human pancreatic stellate cell proliferation was assessed by MTS assay, migration by Boyden chamber assay, and invasion using an invasion chamber. Transient silencing was obtained by small interfering RNA.Human pancreatic stellate cells express COX-2 and synthesize PGE2. Prostaglandin E2 stimulated HPSC proliferation, migration, and invasion and stimulated expression of both extracellular matrix and matrix metalloproteinase genes. Human pancreatic stellate cells expressed all 4 EP receptors. Only blocking the EP4 receptor resulted in abrogation of PGE2-mediated HPSC activation. Specificity of EP4 for the effects of PGE2 on stellate cells was confirmed using specific antagonists.Our data indicate that PGE2 regulates pancreatic stellate cell profibrotic activities via EP4 receptor, thus suggesting EP4 receptor as useful therapeutic target for pancreatic cancer to reduce desmoplasia.
Project description:Cyclooxygenase-2 (COX-2) triggers pro-inflammatory processes that can aggravate neuronal degeneration and functional impairments in many neurological conditions, mainly via producing prostaglandin E2 (PGE2) that activates four membrane receptors, EP1-EP4. However, which EP receptor is the culprit of COX-2/PGE2-mediated neuronal inflammation and degeneration remains largely unclear and presumably depends on the insult types and responding components. Herein, we demonstrated that COX-2 was induced and showed nuclear translocation in two neuronal cell lines - mouse Neuro-2a and human SH-SY5Y - after treatment with neurotoxin 6-hydroxydopamine (6-OHDA), leading to the biosynthesis of PGE2 and upregulation of pro-inflammatory cytokine interleukin-1?. Inhibiting COX-2 or microsomal prostaglandin E synthase-1 suppressed the 6-OHDA-triggered PGE2 production in these cells. Treatment with PGE2 or EP2 selective agonist butaprost, but not EP4 agonist CAY10598, increased cAMP response in both cell lines. PGE2-initiated cAMP production in these cells was blocked by our recently developed novel selective EP2 antagonists - TG4-155 and TG6-10-1, but not by EP4 selective antagonist GW627368X. The 6-OHDA-promoted cytotoxicity was largely blocked by TG4-155, TG6-10-1 or COX-2 selective inhibitor celecoxib, but not by GW627368X. Our results suggest that PGE2 receptor EP2 is a key mediator of COX-2 activity-initiated cAMP signaling in Neuro-2a and SH-SY5Y cells following 6-OHDA treatment, and contributes to oxidopamine-mediated neurotoxicity.
Project description:The cyclooxygenase-2 (COX-2) enzyme is frequently overexpressed in epithelial malignancies including those of the breast, prostate, lung, kidney, ovary, and liver and elevated expression is associated with worse outcomes. COX-2 catalyzes the metabolism of arachidonic acid to prostaglandins. The COX-2 product prostaglandin E2 (PGE2) binds to four G-protein-coupled EP receptors designated EP1-EP4. EP4 is commonly upregulated in cancer and supports cell proliferation, migration, invasion, and metastasis through activation of multiple signaling pathways including ERK, cAMP/PKA, PI3K/AKT, and NF-?B. EP4 antagonists inhibit metastasis in preclinical models. Cancer stem cells, that underlie therapy resistance and disease relapse, are driven by the expression of EP4. Resistance to several chemotherapies is reversed in the presence of EP4 antagonists. In addition to tumor cell-autonomous roles of EP4, many EP4-positive host cells play a role in tumor behavior. Endothelial cell-EP4 supports tumor angiogenesis and lymphangiogenesis. Natural Killer (NK) cells are critical to the mechanism by which systemically administered EP4 antagonists inhibit metastasis. PGE2 acts on EP4 expressed on the NK cell to inhibit tumor target cell killing, cytokine production, and chemotactic activity. Myeloid-derived suppressor cells (MDSCs), that inhibit the development of cytotoxic T cells, are induced by PGE2 acting on myeloid-expressed EP2 and EP4 receptors. Inhibition of MDSC-EP4 leads to maturation of effector T cells and suppresses the induction of T regulatory cells. A number of EP4 antagonists have proven useful in dissecting these mechanisms. There is growing evidence that EP4 antagonism, particularly in combination with either chemotherapy, endocrine therapy, or immune-based therapies, should be investigated further as a promising novel approach to cancer therapy. Several EP4 antagonists have now progressed to early phase clinical trials and we eagerly await the results of those studies.
Project description:Lymphangiogenesis plays an important role in homeostasis, metabolism, and immunity, and also occurs during wound-healing. Here, we examined the roles of prostaglandin E2 (PGE2) receptor (EP) signaling in enhancement of lymphangiogenesis in wound healing processes. The hole-punch was made in the ears of male C57BL/6 mice using a metal ear punch. Healing process and lymphangiogenesis together with macrophage recruitment were analyzed in EP knockout mice. Lymphangiogenesis was up-regulated in the granulation tissues at the margins of punched-hole wounds in mouse ears, and this increase was accompanied by increased expression levels of COX-2 and microsomal prostaglandin E synthase-1. Administration of celecoxib, a COX-2 inhibitor, suppressed lymphangiogenesis in the granulation tissues and reduced the induction of the pro-lymphangiogenic factors, vascular endothelial growth factor (VEGF) -C and VEGF-D. Topical applications of selective EP receptor agonists enhanced the expressions of lymphatic vessel endothelial hyaluronan receptor-1 and VEGF receptor-3. The wound-healing processes and recruitment of CD11b-positive macrophages, which produced VEGF-C and VEGF-D, were suppressed under COX-2 inhibition. Mice lacking either EP3 or EP4 exhibited reduced wound-healing, lymphangiogenesis and recruitment of M2 macrophages, compared with wild type mice. Proliferation of cultured human lymphatic endothelial cells was not detected under PGE2 stimulation. Lymphangiogenesis and recruitment of M2 macrophages that produced VEGF-C/D were suppressed in mice treated with a COX-2 inhibitor or lacking either EP3 or EP4 during wound healing. COX-2 and EP3/EP4 signaling may be novel targets to control lymphangiogenesis in vivo.
Project description:Kaposi's sarcoma-associated herpes virus (KSHV) is implicated in the pathogenesis of KS, a chronic inflammation-associated malignancy. Cyclooxygenase-2 (COX-2) and its metabolite prostaglandin E2 (PGE2), two pivotal proinflammatory/oncogeneic molecules, are proposed to play roles in the expression of major KSHV latency-associated nuclear antigen-1 (LANA-1). Microsomal PGE2 synthase, PGE2, and its receptors (EP1, EP2, EP3, and EP4) were detected in KS lesions with the distinct staining of EP2/EP4 in KS lesions. In latently infected endothelial TIVE-LTC cells, EP receptor antagonists downregulated LANA-1 expression as well as Ca(2+), p-Src, p-PI3K, p-PKCzeta/lambda, and p-NF-kappaB, which are also some of the signal molecules proposed to be important in KS pathogenesis. Exogenous PGE2 and EP receptor agonists induced the LANA-1 promoter in 293 cells, and YY1, Sp1, Oct-1, Oct-6, C/EBP, and c-Jun transcription factors seem to be involved in this induction. PGE2/EP receptor-induced LANA-1 promoter activity was downregulated significantly by the inhibition of Ca(2+), p-Src, p-PI3K, p-PKCzeta/lambda, and p-NF-kappaB. These findings implicate the inflammatory PGE2/EP receptors and the associated signal molecules in herpes virus latency and uncover a novel paradigm that shows the evolution of KSHV genome plasticity to use inflammatory response for its survival advantage of maintaining latent gene expression. These data also suggest that potential use of anti-COX-2 and anti-EP receptor therapy may not only ameliorate the chronic inflammation associated with KS but could also lead to elimination of the KSHV latent infection and the associated KS lesions.
Project description:The number of colorectal cancer (CRC) patients is increasing worldwide. Accumulating evidence has shown that the tumor microenvironment (TME), including macrophages, neutrophils, and fibroblasts, plays an important role in the development and progression of CRC. Although targeting the TME could be a promising therapeutic approach, the mechanisms by which inflammatory cells promote CRC tumorigenesis are not well understood. When inflammation occurs in tissues, prostaglandin E2 (PGE2) is generated from arachidonic acid by the enzyme cyclooxygenase-2 (COX-2). PGE2 regulates multiple functions in various immune cells by binding to the downstream receptors EP1, EP2, EP3, and EP4, and plays an important role in the development of CRC. The current therapies targeting PGE2 using non-steroidal anti-inflammatory drugs (NSAIDs) or COX-2 inhibitors have failed due to the global prostanoid suppression resulting in the severe adverse effects despite the fact they could prevent tumorigenesis. Therefore, therapies targeting the specific downstream molecules of PGE2 signaling could be a promising approach. This review highlights the role of each EP receptor in the TME of CRC tumorigenesis and their therapeutic potential.