Prostaglandin production selectively in brain endothelial cells is both necessary and sufficient for eliciting fever.

Fever is known to be elicited by prostaglandin E2 acting on the brain, but its origin has remained disputed. We show in mice that selective deletion of prostaglandin synthesis in brain endothelial cells, but not in neural cells or myeloid cells, abolished fever induced by intravenous administration of lipopolysaccharide and that selective rescue of prostaglandin synthesis in brain endothelial cells reinstated fever. These data demonstrate that prostaglandin production in brain endothelial cells is both necessary and sufficient for eliciting fever.

A phase 2 trial investigating the efficacy and safety of the mPGES-1 inhibitor vipoglanstat in systemic sclerosis-related Raynaud's.

OBJECTIVE: Our objective was to test the hypothesis, in a double-blind, placebo-controlled study that vipoglanstat, an inhibitor of microsomal prostaglandin E synthase-1 (mPGES-1) which decreases prostaglandin E2 (PGE2) and increases prostacyclin biosynthesis, improves RP. METHODS: Patients with systemic sclerosis (SSc) and ≥7 RP attacks during the last screening week prior to a baseline visit were randomised to four weeks treatment with vipoglanstat 120 mg or placebo. A daily electronic diary captured RP attacks (duration and pain) and Raynaud's Condition Score, with change in RP attacks/week as primary end point. Cold challenge assessments were performed at baseline and end of treatment. Exploratory endpoints included patients' and physicians' global impression of change, Assessment of Scleroderma-associated Raynaud's Phenomenon questionnaire, mPGES-1 activity, and urinary excretion of arachidonic acid metabolites. RESULTS: Sixty-nine subjects received vipoglanstat (n = 33) or placebo (n = 36). Mean weekly number of RP attacks (baseline; vipoglanstat 14.4[SD 6.7], placebo 18.2[12.6]) decreased by 3.4[95% CI -5.8;-1.0] and 4.2[-6.5;-2.0] attacks per week (p= 0.628) respectively. All patient reported outcomes improved, with no difference between the groups. Mean change in recovery of peripheral blood flow after cold challenge did not differ between the study groups. Vipoglanstat fully inhibited mPGES-1, resulting in 57% reduction of PGE2 and 50% increase of prostacyclin metabolites in urine. Vipoglanstat was safe and well tolerated. CONCLUSION: Although vipoglanstat was safe, and well tolerated in a dose achieving full inhibition of mPGES-1, it was ineffective in SSc-related RP. Further development and evaluation of vipoglanstat will therefore be in other diseases where mPGES-1 plays a pathogenetic role.

Targeting PTGES/PGE2 axis enhances sensitivity of colorectal cancer cells to 5-fluorouracil.

Insensitivity and resistance to 5-fluorouracil (5FU) remain as major hurdles for effective and durable 5FU-based chemotherapy in colorectal cancer (CRC) patients. In this study, we identified prostaglandin E synthase (PTGES)/prostaglandin E2 (PGE2) axis as an important regulator for 5FU sensitivity in CRC cells. We found that PTGES expression and PGE2 production are elevated in CRC cells in comparison to normal colorectal epithelial cells. Depletion of PTGES significantly enhanced the inhibitory effect of 5FU on CRC cell viability that was fully reverted by exogenous supplement of PGE2. Inhibition of PTGES enzymatic function, by either inducing loss-of-function mutant or treatment with selective inhibitors, phenocopied the PTGES depletion in terms of 5FU sensitization. Mechanistically, PTGES/PGE2 axis modulates glycolysis in CRC cells, thereby regulating the 5FU sensitivity. Importantly, high PTGES expression is correlated with poor prognosis in 5FU-treated CRC patients. Thus, our study defines PTGES/PGE2 axis as a novel therapeutic target for enhancing the efficacy of 5FU-based chemotherapy in CRC.

Neuroinflammatory mediators in acquired epilepsy: an update.

Epilepsy is a group of chronic neurological disorders that have diverse etiologies but are commonly characterized by spontaneous seizures and behavioral comorbidities. Although the mechanisms underlying the epileptic seizures mostly remain poorly understood and the causes often can be idiopathic, a considerable portion of cases are known as acquired epilepsy. This form of epilepsy is typically associated with prior neurological insults, which lead to the initiation and progression of epileptogenesis, eventually resulting in unprovoked seizures. A convergence of evidence in the past two decades suggests that inflammation within the brain may be a major contributing factor to acquired epileptogenesis. As evidenced in mounting preclinical and human studies, neuroinflammatory processes, such as activation and proliferation of microglia and astrocytes, elevated production of pro-inflammatory cytokines and chemokines, blood-brain barrier breakdown, and upregulation of inflammatory signaling pathways, are commonly observed after seizure-precipitating events. An increased knowledge of these neuroinflammatory processes in the epileptic brain has led to a growing list of inflammatory mediators that can be leveraged as potential targets for new therapies of epilepsy and/or biomarkers that may provide valued information for the diagnosis and prognosis of the otherwise unpredictable seizures. In this review, we mainly focus on the most recent progress in understanding the roles of these inflammatory molecules in acquired epilepsy and highlight the emerging evidence supporting their candidacy as novel molecular targets for new pharmacotherapies of acquired epilepsy and the associated behavioral deficits.

Microsomal Prostaglandin E Synthase-1 and -2: Emerging Targets in Non-Alcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) affects a substantial proportion of the general population and is even more prevalent in obese and diabetic patients. NAFLD, and particularly the more advanced manifestation of the disease, nonalcoholic steatohepatitis (NASH), increases the risk for both liver-related and cardiovascular morbidity. The pathogenesis of NAFLD is complex and multifactorial, with many molecular pathways implicated. Emerging data suggest that microsomal prostaglandin E synthase-1 and -2 might participate in the development and progression of NAFLD. It also appears that targeting these enzymes might represent a novel therapeutic approach for NAFLD. In the present review, we discuss the association between microsomal prostaglandin E synthase-1 and -2 and NAFLD.

The molecular mechanisms of oxygen-sensing in human ductus arteriosus smooth muscle cells: A comprehensive transcriptome profile reveals a central role for mitochondria.

The ductus arteriosus (DA) connects the fetal pulmonary artery and aorta, diverting placentally oxygenated blood from the developing lungs to the systemic circulation. The DA constricts in response to increases in oxygen (O2) with the first breaths, resulting in functional DA closure, with anatomic closure occurring within the first days of life. Failure of DA closure results in persistent patent ductus arteriosus (PDA), a common complication of extreme preterm birth. The DA's response to O2, though modulated by the endothelium, is intrinsic to the DA smooth muscle cells (DASMC). DA constriction is mediated by mitochondrial-derived reactive oxygen species, which increase in proportion to arterial partial pressure of oxygen (PaO2). The resulting redox changes inhibit voltage-gated potassium channels (Kv) leading to cell depolarization, calcium influx and DASMC constriction. To date, there has not been an unbiased assessment of the human DA O2-sensors using transcriptomics, nor are there known molecular mechanisms which characterize DA closure. DASMCs were isolated from DAs obtained from 10 term infants at the time of congenital heart surgery. Cells were purified by flow cytometry, negatively sorting using CD90 and CD31 to eliminate fibroblasts or endothelial cells, respectively. The purity of the DASMC population was confirmed by positive staining for α-smooth muscle actin, smoothelin B and caldesmon. Cells were grown for 96 h in hypoxia (2.5% O2) or normoxia (19% O2) and confocal imaging with Cal-520 was used to determine oxygen responsiveness. An oxygen-induced increase in intracellular calcium of 18.1% ± 4.4% and SMC constriction (-27% ± 1.5% shortening) occurred in all cell lines within five minutes. RNA sequencing of the cells grown in hypoxia and normoxia revealed significant regulation of 1344 genes (corrected p < 0.05). We examined these genes using Gene Ontology (GO). This unbiased assessment of altered gene expression indicated significant enrichment of the following GOterms: mitochondria, cellular respiration and transcription. The top regulated biologic process was generation of precursor metabolites and energy. The top regulated cellular component was mitochondrial matrix. The top regulated molecular function was transcription coactivator activity. Multiple members of the NADH-ubiquinone oxidoreductase (NDUF) family are upregulated in human DASMC (hDASMC) following normoxia. Several of our differentially regulated transcripts are encoded by genes that have been associated with genetic syndromes that have an increased incidence of PDA (Crebb binding protein and Histone Acetyltransferase P300). This first examination of the effects of O2 on human DA transcriptomics supports a putative role for mitochondria as oxygen sensors.

A High Body Mass Index and the Vacuum Phenomenon Upregulate Pain-Related Molecules in Human Degenerated Intervertebral Discs.

Animal studies suggest that pain-related-molecule upregulation in degenerated intervertebral discs (IVDs) potentially leads to low back pain (LBP). We hypothesized that IVD mechanical stress and axial loading contribute to discogenic LBP's pathomechanism. This study aimed to elucidate the relationships among the clinical findings, radiographical findings, and pain-related-molecule expression in human degenerated IVDs. We harvested degenerated-IVD samples from 35 patients during spinal interbody fusion surgery. Pain-related molecules including tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-6, calcitonin gene-related peptide (CGRP), microsomal prostaglandin E synthase-1 (mPGES1), and nerve growth factor (NGF) were determined. We also recorded preoperative clinical findings including body mass index (BMI), Oswestry Disability Index (ODI), and radiographical findings including the vacuum phenomenon (VP) and spinal instability. Furthermore, we compared pain-related-molecule expression between the VP (-) and (+) groups. BMI was significantly correlated with the ODI, CGRP, and mPGES-1 levels. In the VP (+) group, mPGES-1 levels were significantly higher than in the VP (-) group. Additionally, CGRP and mPGES-1 were significantly correlated. Axial loading and mechanical stress correlated with CGRP and mPGES-1 expression and not with inflammatory cytokine or NGF expression. Therefore, axial loading and mechanical stress upregulate CGRP and mPGES-1 in human degenerated IVDs, potentially leading to chronic discogenic LBP.

Establishment of an in vitro 3D model for neuroblastoma enables preclinical investigation of combined tumor-stroma drug targeting.

The majority of anti-cancer therapies target the proliferating tumor cells, while the tumor stroma, principally unaffected, survives, and provide a niche for surviving tumor cells. Combining tumor cell and stroma-targeting therapies thus have a potential to improve patient outcome. The neuroblastoma stroma contains cancer-associated fibroblasts expressing microsomal prostaglandin E synthase-1 (mPGES-1). mPGES-1-derived prostaglandin E2 (PGE2 ) is known to promote tumor growth through increased proliferation and survival of tumor cells, immune suppression, angiogenesis, and therapy resistance, and we, therefore, hypothesize that mPGES-1 constitutes an interesting stromal target. Here, we aimed to develop a relevant in vitro model to study combination therapies. Co-culturing of neuroblastoma and fibroblast cells in 3D tumor spheroids mimic neuroblastoma tumors with regard to the cyclooxygenase/mPGES-1/PGE2 pathway. Using the spheroid model, we show that the inhibition of fibroblast-derived mPGES-1 enhanced the cytotoxic effect of doxorubicin and vincristine and significantly reduced tumor cell viability and spheroid growth. Cyclic treatment with vincristine in combination with an mPGES-1 inhibitor abrogated cell repopulation. Moreover, inhibition of mPGES-1 potentiated the cytotoxic effect of vincristine on established neuroblastoma allografts in mice. In conclusion, we established a 3D neuroblastoma model, highlighting the potential of combining stromal targeting of mPGES-1 with tumor cell targeting drugs like vincristine.

Diosgenin suppresses COX-2 and mPGES-1 via GR and improves LPS-induced liver injury in mouse.

Using a wild yam (Dioscorea japonica), we previously found novel anti-inflammatory and anti-carcinogenic effects via the downregulation of cyclooxygenase (COX)-2 and microsomal prostaglandin E synthase (mPGES)-1. One of the substances in wild yam is a steroidal saponin, diosgenin. We demonstrated that diosgenin suppressed COX-2 in human non-small-cell lung carcinoma A549 cells via nuclear factor-kappa B (NF-κB) translocation and the effects were reversed by a glucocorticoid receptor antagonist, RU486. In lipopolysaccharide (LPS)-induced mouse liver injury, COX-2 and mPGES-1 were induced and localized in sinusoidal macrophages and endothelial cells; however, diosgenin administration significantly suppressed Ptgs2 and Ptges expression and decreased COX-2 and mPGES-1 immunopositive cells in the sinusoids. Multiple immunohistochemical analyses showed that diosgenin had an effect on COX-2 and mPGES-1, particularly in the macrophages. Thus, we showed that diosgenin downregulated COX-2 and mPGES-1 via the glucocorticoid receptor and suppressed COX-2 and mPGES-1 in the macrophages of LPS-induced acute mouse liver injury.

Pathophysiological role of prostaglandin E synthases in liver diseases.

Prostaglandin E synthases (PGESs) convert cyclooxygenase (COX)-derived prostaglandin H2 (PGH2) into prostaglandin E2 (PGE2) and comprise at least three types of structurally and biologically distinct enzymes. Two of these, namely microsomal prostaglandin E synthase-1 (mPGES-1) and mPGES-2, are membrane-bound enzymes. mPGES-1 is an inflammation-inducible enzyme that converts PGH2 into PGE2. mPGES-2 is a bifunctional enzyme that generally forms a complex with haem in the presence of glutathione. This enzyme can metabolise PGH2 into malondialdehyde and can produce PGE2 after its separation from haem. In this review, we discuss the role of PGESs, particularly mPGES-1 and mPGES-2, in the pathogenesis of liver diseases. A better understanding of the roles of PGESs in liver disease may aid in the development of treatments for patients with liver diseases.

A defective prostaglandin E synthase could affect egg formation in the silkworm Bombyx mori.

We had previously reported a prostaglandin E synthase (bmPGES) in the silkworm Bombyx mori that catalyzes the isomerization of PGH2 to PGE2. The present study aimed to provide a genome-editing characterization of bmPGES in B. mori. Results showed bmPGES gene disruption to result in a reduced content of PGE2. The change affected the expression of chorion genes and egg formation in silkworms. Collectively, the results indicated that bmPGES could be involved in reproduction of B. mori. Therefore, this study provides insights into the physiological role of bmPGES and PGE2 in silkworms.

Ellagitannins from Punica granatum leaves suppress microsomal prostaglandin E synthase-1 expression and induce lung cancer cells to undergo apoptosis.

Prostaglandin E2 (PGE2), which is a potent pro-inflammatory lipid mediator, is biosynthesized from arachidonic acid by cyclooxygenase-2 (COX-2) and microsomal PGE synthase-1 (mPGES-1). Non-steroidal anti-inflammatory drugs (NSAIDs) are used clinically as COX inhibitors, but they have gastrointestinal and cardiovascular side-effects. Thus, the terminal enzyme mPGES-1 holds promise as the next therapeutic target. In this study, we found that the ellagitannins granatin A and granatin B isolated from pomegranate leaves, and geraniin, which is their structural analog, selectively suppressed mPGES-1 expression without affecting COX-2 in non-small cell lung carcinoma A549 cells. The ellagitannins also down-regulated tumor necrosis factor α, inducible nitric oxide synthase, and anti-apoptotic factor B-cell chronic lymphocytic leukemia/lymphoma 2, and induced A549 cells to undergo apoptosis. These findings indicate that the ellagitannins have anti-inflammatory and anti-carcinogenic effects, due to their specific suppression of mPGES-1.Abbreviations: Bcl-2: B-cell chronic lymphocytic leukemia/lymphoma 2; COX: cyclooxygenase; CRE: cAMP response element; DHHDP: dehydrohexahydroxydiphenoyl; Et2O: diethyl ether; EtOAc: ethyl acetate; GAPDH: glyceraldehyde 3-phosphate dehydrogenase; iNOS: inducible nitric oxide synthase; mPGES-1: microsomal prostaglandin E synthase-1; n-BuOH: water-saturated n-butanol; NSAIDs: non-steroidal anti-inflammatory drugs; NF-κB: nuclear factor-κB; PG: prostaglandin; TNF: tumor necrosis factor; TUNEL: terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling.

Aminothiazoles inhibit osteoclastogenesis and PGE2 production in LPS-stimulated co-cultures of periodontal ligament and RAW 264.7 cells, and RANKL-mediated osteoclastogenesis and bone resorption in PBMCs.

Inflammatory mediator prostaglandin E2 (PGE2 ) contributes to bone resorption in several inflammatory conditions including periodontitis. The terminal enzyme, microsomal prostaglandin E synthase-1 (mPGES-1) regulating PGE2 synthesis is a promising therapeutic target to reduce inflammatory bone loss. The aim of this study was to investigate effects of mPGES-1 inhibitors, aminothiazoles TH-848 and TH-644, on PGE2 production and osteoclastogenesis in co-cultures of periodontal ligament (PDL) and osteoclast progenitor cells RAW 264.7, stimulated by lipopolysaccharide (LPS), and bone resorption in RANKL-mediated peripheral blood mononuclear cells (PBMCs). PDL and RAW 264.7 cells were cultured separately or co-cultured and treated with LPS alone or in combination with aminothiazoles. Multinucleated cells stained positively for tartrate-resistant acid phosphatase (TRAP) were scored as osteoclast-like cells. Levels of PGE2 , osteoprotegerin (OPG) and interleukin-6, as well as mRNA expression of mPGES-1, OPG and RANKL were analysed in PDL cells. PBMCs were treated with RANKL alone or in combination with aminothiazoles. TRAP-positive multinucleated cells were analysed and bone resorption was measured by the CTX-I assay. Aminothiazoles reduced LPS-stimulated osteoclast-like cell formation both in co-cultures and in RAW 264.7 cells. Additionally, aminothiazoles inhibited PGE2 production in LPS-stimulated cultures, but did not affect LPS-induced mPGES-1, OPG or RANKL mRNA expression in PDL cells. In PBMCs, inhibitors decreased both osteoclast differentiation and bone resorption. In conclusion, aminothiazoles reduced the formation of osteoclast-like cells and decreased the production of PGE2 in co-cultures as well as single-cell cultures. Furthermore, these compounds inhibited RANKL-induced bone resorption and differentiation of PBMCs, suggesting these inhibitors for future treatment of inflammatory bone loss such as periodontitis.

Microsomal prostaglandin E synthase-1 is a critical factor in dopaminergic neurodegeneration in Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disorder of uncertain pathogenesis characterized by the loss of nigrostriatal dopaminergic neurons. Although increased production of prostaglandin E2 (PGE2) has been implicated in tissue damage in several pathological settings, the role of microsomal prostaglandin E synthase-1 (mPGES-1), an inducible terminal enzyme for PGE2 synthesis, in dopaminergic neurodegeneration remains unclear. Here we show that mPGES-1 is up-regulated in the dopaminergic neurons of the substantia nigra of postmortem brain tissue from PD patients and in neurotoxin 6-hydroxydopamine (6-OHDA)-induced PD mice. The expression of mPGES-1 was also up-regulated in cultured dopaminergic neurons stimulated with 6-OHDA. The genetic deletion of mPGES-1 not only abolished 6-OHDA-induced PGE2 production but also inhibited 6-OHDA-induced dopaminergic neurodegeneration both in vitro and in vivo. Nigrostriatal projections, striatal dopamine content, and neurological functions were significantly impaired by 6-OHDA administration in wild-type (WT) mice, but not in mPGES-1 knockout (KO) mice. Furthermore, in cultured primary mesencephalic neurons, addition of PGE2 to compensate for the deficiency of 6-OHDA-induced PGE2 production in mPGES-1 KO neurons recovered 6-OHDA toxicity to almost the same extent as that seen in WT neurons. These results suggest that induction of mPGES-1 enhances 6-OHDA-induced dopaminergic neuronal death through excessive PGE2 production. Thus, mPGES-1 may be a valuable therapeutic target for treatment of PD.

Microsomal prostaglandin E synthase 2 deficiency is resistant to acetaminophen-induced liver injury.

Acetaminophen (APAP)-induced liver injury is the main cause of acute liver failure. This study investigated the role of microsomal prostaglandin E synthase 2 (mPGES-2), discovered as one of the prostaglandin E2 (PGE2) synthases, in mediating APAP-induced liver injury. Using mPGES-2 wild-type (WT) and knockout (KO) mice, marked resistance to APAP-induced liver damage was found in mPGES-2 KO, as indicated by robust improvement of liver histology, changes in liver enzyme release, and marked decrease in APAP-cysteine adducts (APAP-CYS) and inflammatory markers. Moreover, the results confirmed that increase in liver PGE2 content in KO mice under basal conditions was not critical for the protection from APAP-induced liver injury. Importantly, mPGES-2 deletion inhibited the production of malondialdehyde (MDA), increasing glutathione (GSH) level. Enhanced GSH level may contribute to the inhibition of APAP toxicity in mPGES-2 KO mice. To further elucidate the role of mPGES-2 in the liver injury induced by APAP, adeno-associated viruses (AAV) were used to overexpress mPGES-2 in the liver. The results showed that mPGES-2 overexpression aggravates liver injury associated with an increase in inflammatory markers and chemokines after APAP treatment. Moreover, a lower level of GSH was detected in the mPGES-2 overexpression group compared to the control group. Collectively, our findings indicate that mPGES-2 plays a critical role in regulating APAP-induced liver injury, possibly by regulating GSH and APAP-CYS level, which may provide a potential therapeutic strategy for the prevention and treatment of APAP-induced liver injury.

Targeting the COX/mPGES-1/PGE2 Pathway in Neuroblastoma.

The importance of prostaglandin E2 in cancer progression is well established, but research on its role in cancer has so far mostly been focused on epithelial cancer in adults while the knowledge about the contribution of prostaglandin E2 to childhood malignancies is limited. Neuroblastoma, an extracranial solid tumor of the sympathetic nervous system, mainly affects young children. Patients with tumors classified as high-risk have poor survival despite receiving intensive treatment, illustrating a need for new treatments complimenting existing ones. The basis of neuroblastoma treatment e.g. chemotherapy and radiation therapy, target the proliferating genetically unstable tumor cells leading to treatment resistance and relapses. The tumor microenvironment is an avenue, still to a great extent, unexplored and lacking effective targeted therapies. Cancer-associated fibroblasts is the main source of prostaglandin E2 in neuroblastoma contributing to angiogenesis, immunosuppression and tumor growth. Prostaglandin E2 is formed from its precursor arachidonic acid in a two-step enzymatic reaction. Arachidonic acid is first converted by cyclooxygenases into prostaglandin H2 and then further converted by microsomal prostaglandin E synthase-1 into prostaglandin E2. We believe targeting of microsomal prostaglandin E synthase-1 in cancer-associated fibroblasts will be an effective future therapeutic strategy in fighting neuroblastoma.

A pilot, randomized, double-blind, placebo-controlled trial to assess the safety and efficacy of a novel Boswellia serrata extract in the management of osteoarthritis of the knee.

A double-blind, placebo-controlled human trial was conducted to evaluate the safety and efficacy of a standardized oral supplementation of Boswellin®, a novel extract of Boswellia serrata extract (BSE) containing 3-acetyl-11-keto-ß-boswellic acid (AKBBA) with ß-boswellic acid (BBA). A total of 48 patients with osteoarthritis (OA) of the knee were randomized and allocated to the BSE and placebo groups for intervention. Patients were administered BSE or placebo for a period of 120 days. The trial results revealed that BSE treatment significantly improved the physical function of the patients by reducing pain and stiffness compared with placebo. Radiographic assessments showed improved knee joint gap and reduced osteophytes (spur) confirming the efficacy of BSE treatment. BSE also significantly reduced the serum levels of high-sensitive C-reactive protein, a potential inflammatory marker associated with OA of the knee. No serious adverse events were reported. This is the first study with BSE conducted for a period of 120 days, longer than any other previous clinical trial on patients with OA of the knee. The findings provide evidence that biologically active constituents of BSE, namely, AKBBA and BBA, act synergistically to exert anti-inflammatory/anti-arthritic activity showing improvement in physical and functional ability and reducing the pain and stiffness.

Immune-Induced Fever Is Dependent on Local But Not Generalized Prostaglandin E2 Synthesis in the Brain.

Fever occurs upon binding of prostaglandin E2 (PGE2) to EP3 receptors in the median preoptic nucleus of the hypothalamus, but the origin of the pyrogenic PGE2 has not been clearly determined. Here, using mice of both sexes, we examined the role of local versus generalized PGE2 production in the brain for the febrile response. In wild-type mice and in mice with genetic deletion of the prostaglandin synthesizing enzyme cyclooxygenase-2 in the brain endothelium, generated with an inducible CreERT2 under the Slco1c1 promoter, PGE2 levels in the CSF were only weakly related to the magnitude of the febrile response, whereas the PGE2 synthesizing capacity in the hypothalamus, as reflected in the levels of cyclooxygenase-2 mRNA, showed strong correlation with the immune-induced fever. Histological analysis showed that the deletion of cyclooxygenase-2 in brain endothelial cells occurred preferentially in small- and medium-sized vessels deep in the brain parenchyma, such as in the hypothalamus, whereas larger vessels, and particularly those close to the neocortical surface and in the meninges, were left unaffected, hence leaving PGE2 synthesis largely intact in major parts of the brain while significantly reducing it in the region critical for the febrile response. Furthermore, injection of a virus vector expressing microsomal prostaglandin E synthase-1 (mPGES-1) into the median preoptic nucleus of fever-refractive mPGES-1 knock-out mice, resulted in a temperature elevation in response to LPS. We conclude that the febrile response is dependent on local release of PGE2 onto its target neurons and not on the overall PGE2 production in the brain.SIGNIFICANCE STATEMENT By using mice with selective deletion of prostaglandin synthesis in brain endothelial cells, we demonstrate that local prostaglandin E2 (PGE2) production in deep brain areas, such as the hypothalamus, which is the site of thermoregulatory neurons, is critical for the febrile response to peripheral inflammation. In contrast, PGE2 production in other brain areas and the overall PGE2 level in the brain do not influence the febrile response. Furthermore, partly restoring the PGE2 synthesizing capacity in the anterior hypothalamus of mice lacking such capacity with a lentiviral vector resulted in a temperature elevation in response to LPS. These data imply that the febrile response is dependent on the local release of PGE2 onto its target neurons, possibly by a paracrine mechanism.

WWL70 protects against chronic constriction injury-induced neuropathic pain in mice by cannabinoid receptor-independent mechanisms.

BACKGROUND: Targeting the endocannabinoid system has emerged as an effective strategy for the treatment of inflammatory and neurological diseases. Unlike the inhibition of the principal 2-arachidonyl glycerol (2-AG) hydrolytic enzyme monoacylglycerol lipase (MAGL), which leads to 2-AG overload and cannabinoid receptor desensitization, selective inhibition of the minor 2-AG hydrolytic enzyme alpha, beta-hydrolase domain 6 (ABHD6) can provide therapeutic benefits without producing cannabimimetic side effects. We have shown that inhibition of ABHD6 significantly reduces neuroinflammation and exerts neuroprotection in animal models of traumatic brain injury and multiple sclerosis. However, the role of ABHD6 inhibition on neuropathic pain has not been explored. METHODS: Neuropathic pain was induced by chronic constriction injury (CCI) of the mouse sciatic nerve and examined by Hargreaves and Von Frey tests. Activation of inflammatory cells and the production of cytokines and chemokines in the spinal cord dorsal horn, dorsal root ganglion (DRG), and sciatic nerve were assessed by qRT-PCR, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry. The levels of 2-AG and arachidonic acid (AA) in sciatic nerve were quantified by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). RESULTS: Treatment with the selective ABHD6 inhibitor WWL70 significantly alleviated CCI-induced thermal hyperalgesia and mechanical allodynia. Microglia activation, macrophage infiltration, and the production of nociceptive mediators were reduced in the ipsilateral lumbar spinal cord dorsal horn, DRG, and sciatic nerve of WWL70-treated animals. The diminished cytokine and chemokine production is likely due to the inhibitory effect of WWL70 on NF-κB phosphorylation. Surprisingly, the anti-nociceptive and anti-inflammatory effects of WWL70 were not reversed by addition of the cannabinoid receptor antagonists. Treatment with WWL70 did not alter the levels of 2-AG, AA, and the phosphorylation of cytosolic phospholipase A2 (cPLA2), but significantly reduced the production of prostaglandin E2 (PGE2) and the expression of cyclooxygenase-2 (COX-2) and prostaglandin E synthase-2 (PGES2) in the injured sciatic nerve. CONCLUSIONS: This study reveals a novel mechanism for the antinociceptive effect of the 2-AG catabolic enzyme ABHD6 inhibitor WWL70. Understanding the interaction between endocannabinoid and eicosanoid pathways might provide a new avenue for the treatment of inflammatory and neuropathic pain.

Sulforaphane, a Chemopreventive Compound, Inhibits Cyclooxygenase-2 and Microsomal Prostaglandin E Synthase-1 Expression in Human HT-29 Colon Cancer Cells.

BACKGROUND: A high expression of prostaglandin E2 (PGE2) is found in colorectal cancer. Therefore, blocking of PGE2 generation has been identified as a promising approach for anticancer therapy. Sulforaphane (SFN), an isothiocyanate derived from glucosinolate, is used as the antioxidant and anticancer agents. METHODS: HT-29 cells were treated with various concentrations of SFN and compared to untreated cells for the expression of microsomal prostaglandin E synthase-1 (mPGES-1), cyclooxygenase 2 (COX-2), hypoxia-inducible factor-1 (HIF-1), C-X-C chemokine receptor type 4 (CXCR4), vascular endothelial growth factor (VEGF), and matrix metalloproteinase (MMP)-2 and MMP-9 at the mRNA level. The PGE2 level was measured by ELISA assay. Apoptosis was evaluated by the proportion of sub-G1 cells. The activity of caspase-3 was determined using an enzymatic assay. HT-29 cell migration was assessed using a scratch test. RESULTS: SFN preconditioning decreased the expression of COX-2, mPGES-1, HIF-1, VEGF, CXCR4, MMP-2, and MMP-9. An apoptotic effect of SFN was preceded by the activation of caspase-3 as well as accumulation of cells in the sub-G1 phase of the cell cycle. SFN decreased PGE2 generation and inhibited the in vitro motility/wound-healing activity of HT-29 cells. CONCLUSIONS: SFN anticancer effects are associated with antiproliferative, antiangiogenic, and antimetastatic activities arising from the downregulation of the COX-2/ mPGES-1 axis.