RESUMEN
Oligodendrocyte-lineage cells, including NG2 glia, undergo prominent changes in various neurodegenerative disorders. Here, we identify a neuroprotective role for NG2 glia against prion toxicity. NG2 glia were activated after prion infection in cerebellar organotypic cultured slices (COCS) and in brains of prion-inoculated mice. In both model systems, depletion of NG2 glia exacerbated prion-induced neurodegeneration and accelerated prion pathology. Loss of NG2 glia enhanced the biosynthesis of prostaglandin E2 (PGE2) by microglia, which augmented prion neurotoxicity through binding to the EP4 receptor. Pharmacological or genetic inhibition of PGE2 biosynthesis attenuated prion-induced neurodegeneration in COCS and mice, reduced the enhanced neurodegeneration in NG2-glia-depleted COCS after prion infection, and dampened the acceleration of prion disease in NG2-glia-depleted mice. These data unveil a non-cell-autonomous interaction between NG2 glia and microglia in prion disease and suggest that PGE2 signaling may represent an actionable target against prion diseases.
Asunto(s)
Dinoprostona , Microglía , Neuroglía , Neuronas , Enfermedades por Prión , Transducción de Señal , Animales , Microglía/metabolismo , Dinoprostona/metabolismo , Ratones , Transducción de Señal/fisiología , Neuronas/metabolismo , Neuronas/efectos de los fármacos , Neuronas/patología , Neuroglía/metabolismo , Enfermedades por Prión/metabolismo , Enfermedades por Prión/patología , Ratones Endogámicos C57BL , Antígenos/metabolismo , Proteoglicanos/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Priones/metabolismo , Priones/toxicidad , Cerebelo/metabolismo , Cerebelo/patologíaRESUMEN
The parasitic helminth Schistosoma mansoni is a potent inducer of type 2 immune responses by stimulating dendritic cells (DCs) to prime T helper 2 (Th2) responses. We previously found that S. mansoni soluble egg antigens (SEA) promote the synthesis of Prostaglandin E2 (PGE2) by DCs through ERK-dependent signaling via Dectin-1 and Dectin-2 that subsequently induces OX40L expression, licensing them for Th2 priming, yet the ligands present in SEA involved in driving this response and whether specific targeting of PGE2 synthesis by DCs could affect Th2 polarization are unknown. We here show that the ability of SEA to bind Dectin-2 and drive ERK phosphorylation, PGE2 synthesis, OX40L expression, and Th2 polarization is impaired upon cleavage of high-mannose glycans by Endoglycosidase H treatment. This identifies high-mannose glycans present on glycoproteins in SEA as important drivers of this signaling axis. Moreover, we find that OX40L expression and Th2 induction are abrogated when microsomal prostaglandin E synthase-1 (mPGES) is selectively inhibited, but not when a general COX-1/2 inhibitor is used. This shows that the de novo synthesis of PGE2 is vital for the Th2 priming function of SEA-stimulated DCs as well as points to the potential existence of other COX-dependent lipid mediators that antagonize PGE2-driven Th2 polarization. Lastly, specific PGE2 inhibition following immunization with S. mansoni eggs dampened the egg-specific Th cell response. In summary, our findings provide new insights in the molecular mechanisms underpinning Th2 induction by S. mansoni and identify druggable targets for potential control of helminth driven-Th2 responses.
Asunto(s)
Dinoprostona , Lectinas Tipo C , Manosa , Polisacáridos , Schistosoma mansoni , Células Th2 , Animales , Ratones , Antígenos Helmínticos/inmunología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Dinoprostona/metabolismo , Lectinas Tipo C/metabolismo , Lectinas Tipo C/inmunología , Manosa/metabolismo , Manosa/inmunología , Ratones Endogámicos C57BL , Óvulo/inmunología , Óvulo/metabolismo , Ligando OX40/metabolismo , Polisacáridos/inmunología , Polisacáridos/metabolismo , Schistosoma mansoni/inmunología , Esquistosomiasis mansoni/inmunología , Esquistosomiasis mansoni/metabolismo , Esquistosomiasis mansoni/parasitología , Células Th2/inmunología , Células Th2/metabolismoRESUMEN
BACKGROUND: Disease-modifying antirheumatic drugs (DMARDs) are widely used for treating rheumatoid arthritis (RA). However, there are no established biomarkers to predict a patient's response to these therapies. Prostanoids, encompassing prostaglandins, prostacyclins, and thromboxanes, are potent lipid mediators implicated in RA progression. Nevertheless, the influence of DMARDs on prostanoid biosynthesis in RA patients remains poorly understood. This study aims to assess the impact of various DMARDs on urinary prostanoids levels and to explore whether urinary prostanoid profiles correlate with disease activity or response to therapy. METHODS: This study included 152 Swedish female patients with early RA, all rheumatoid factor (RF) positive, enrolled in the NORD-STAR trial (registration number: NCT01491815). Participants were randomized into four therapeutic regimes: methotrexate (MTX) combined with (i) prednisolone (arm ACT), (ii) TNF-α blocker certolizumab pegol (arm CZP), (iii) CTLA-4Ig abatacept (arm ABA), or (iv) IL-6R blocker tocilizumab (arm TCZ). Urine samples, collected before start of treatment and at 24 weeks post-treatment, were analyzed for tetranor-prostaglandin E metabolite (tPGEM), tetranor-prostaglandin D metabolite (tPGDM), 2,3-dinor thromboxane B2 (TXBM), 2,3-dinor-6-keto prostaglandin F1a (PGIM), leukotriene E4 (LTE4) and 12-hydroxyeicosatetraenoic acid (12-HETE) using liquid chromatography-mass spectrometry (LC-MS). Generalized estimating equation (GEE) models were used to analyze the change in urinary eicosanoids and their correlations to clinical outcomes. RESULTS: Patients receiving MTX combined with CZP or TCZ exhibited significant elevations in urinary tPGEM and TXBM levels after 24 weeks of treatment. Other eicosanoids did not show significant alterations in response to any treatment. Baseline urinary eicosanoid levels did not correlate with baseline clinical disease activity index (CDAI) levels, nor with changes in CDAI from baseline to week 24. Their levels were also similar between patients who achieved CDAI remission and those with active disease at week 24. CONCLUSIONS: Treatment with anti-TNF or anti-IL6R agents in early RA patients leads to an increased systemic production of proinflammatory and prothrombotic prostanoids. However, urinary eicosanoid levels do not appear to be predictive of the response to DMARDs therapy.
Asunto(s)
Antirreumáticos , Artritis Reumatoide , Dimaprit/análogos & derivados , Humanos , Femenino , Prostaglandinas , Antirreumáticos/uso terapéutico , Inhibidores del Factor de Necrosis Tumoral , Artritis Reumatoide/tratamiento farmacológico , Metotrexato , Certolizumab PegolRESUMEN
High-throughput drug screening enables the discovery of new anticancer drugs. Although monolayer cell cultures are commonly used for screening, their limited complexity and translational efficiency require alternative models. Three-dimensional cell cultures, such as multicellular tumor spheroids (MCTS), mimic tumor architecture and offer promising opportunities for drug discovery. In this study, we developed a neuroblastoma MCTS model for high-content drug screening. We also aimed to decipher the mechanisms underlying synergistic drug combinations in this disease model. Several agents from different therapeutic categories and with different mechanisms of action were tested alone or in combination with selective inhibition of prostaglandin E2 by pharmacological inhibition of microsomal prostaglandin E synthase-1 (mPGES-1). After a systematic investigation of the sensitivity of individual agents and the effects of pairwise combinations, GFP-transfected MCTS were used in a confirmatory screen to validate the hits. Finally, inhibitory effects on multidrug resistance proteins were examined. In summary, we demonstrate how MCTS-based high-throughput drug screening has the potential to uncover effective drug combinations and provide insights into the mechanism of synergy between an mPGES-1 inhibitor and chemotherapeutic agents.
Asunto(s)
Resistencia a Antineoplásicos , Neuroblastoma , Humanos , Prostaglandina-E Sintasas , Esferoides Celulares , Neuroblastoma/tratamiento farmacológico , Descubrimiento de Drogas/métodosRESUMEN
Microsomal Prostaglandin E Synthase 1 (mPGES-1) is the key enzyme for the generation of the pro-inflammatory lipid mediator prostaglandin E2 (PGE2), which contributes to several pathological features of many diseases. Inhibition of mPGES-1 has been shown to be a safe and effective therapeutic strategy in various pre-clinical studies. In addition to reduced PGE2 formation, it is also suggested that the potential shunting into other protective and pro-resolving prostanoids may play an important role in resolution of inflammation. In the present study, we analysed the eicosanoid profiles in four in vitro inflammation models and compared the effects of mPGES-1 inhibition with those of cyclooxygenase-2 (Cox-2) inhibition. Our results showed a marked shift to the PGD2 pathway under mPGES-1 inhibition in A549 cells, RAW264.7 cells and mouse bone marrow-derived macrophages (BMDMs), whereas enhanced prostacyclin production was observed in rheumatoid arthritis synovial fibroblasts (RASFs) treated with an mPGES-1 inhibitor. As expected, Cox-2 inhibition completely suppressed all prostanoids. This study suggests that the therapeutic effects of mPGES-1 inhibition may be mediated by modulation of other prostanoids in addition to PGE2 reduction.
Asunto(s)
Inflamación , Prostaglandinas , Ratones , Animales , Prostaglandina-E Sintasas/metabolismo , Ciclooxigenasa 2/metabolismo , Ácido Araquidónico , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Dinoprostona/metabolismo , EicosanoidesRESUMEN
BACKGROUND AND PURPOSE: Heart failure with reduced ejection fraction (HFrEF) is a major consequence of myocardial infarction (MI). The microsomal prostaglandin E synthase-1 (mPGES-1)/PGE2 pathway has been shown to constrain reperfusion injury after acute myocardial ischaemia. However, it is unknown whether pharmacological inhibition of mPGES-1, a target with lower risk of thrombosis compared with selective inhibition of cyclooxygenase-2, affects chronic cardiac remodelling after MI. EXPERIMENTAL APPROACH: Mice were subjected to left anterior descending coronary artery ligation, followed by intraperitoneal treatment with the mPGES-1 inhibitor compound III (CIII) or 118, celecoxib (cyclooxygenase-2 inhibitor) or vehicle, once daily for 28 days. Urinary prostanoid metabolites were measured by liquid chromatography-tandem mass spectrometry. KEY RESULTS: Chronic administration of CIII improved cardiac function in mice after MI compared with vehicle or celecoxib. CIII did not affect thrombogenesis or blood pressure. In addition, CIII reduced infarct area, augmented scar thickness, decreased collagen I/III ratio, decreased the expression of fibrosis-related genes and increased capillary density in the ischaemic area. Shunting to urinary metabolites of PGI2 , not thromboxane B2 or PGD2 , after inhibition of mPGES-1 was positively correlated with cardiac function after MI. CIII administration significantly increased urinary PGI2 /PGE2 metabolite ratio compared to vehicle or celecoxib. The PGI2 /PGE2 metabolite ratio correlated positively with ejection fraction, fractional shortening and scar thickness. Treatment with 118 also improved cardiac function. CONCLUSION AND IMPLICATIONS: Inhibition of mPGES-1 prevented chronic adverse cardiac remodelling via an augmented PGI2 /PGE2 metabolite ratio and therefore represents a potential therapeutic strategy for development of HFrEF after MI.
Asunto(s)
Insuficiencia Cardíaca , Infarto del Miocardio , Animales , Ratones , Prostaglandina-E Sintasas/metabolismo , Celecoxib/farmacología , Cicatriz , Remodelación Ventricular , Volumen Sistólico , Infarto del Miocardio/genética , Inhibidores de la Ciclooxigenasa 2RESUMEN
Inhibition of microsomal prostaglandin E synthase-1 (mPGES-1) results in decreased production of proinflammatory PGE2 and can lead to shunting of PGH2 into the prostaglandin D2 (PGD2)/15-deoxy-Δ12,14-prostaglandin J2 (15dPGJ2) pathway. 15dPGJ2 forms Michael adducts with thiol-containing biomolecules such as GSH or cysteine residues on target proteins and is thought to promote resolution of inflammation. We aimed to elucidate the biosynthesis and metabolism of 15dPGJ2 via conjugation with GSH, to form 15dPGJ2-glutathione (15dPGJ2-GS) and 15dPGJ2-cysteine (15dPGJ2-Cys) conjugates and to characterize the effects of mPGES-1 inhibition on the PGD2/15dPGJ2 pathway in mouse and human immune cells. Our results demonstrate the formation of PGD2, 15dPGJ2, 15dPGJ2-GS, and 15dPGJ2-Cys in RAW264.7 cells after lipopolysaccharide stimulation. Moreover, 15dPGJ2-Cys was found in lipopolysaccharide-activated primary murine macrophages as well as in human mast cells following stimulation of the IgE-receptor. Our results also suggest that the microsomal glutathione S-transferase 3 is essential for the formation of 15dPGJ2 conjugates. In contrast to inhibition of cyclooxygenase, which leads to blockage of the PGD2/15dPGJ2 pathway, we found that inhibition of mPGES-1 preserves PGD2 and its metabolites. Collectively, this study highlights the formation of 15dPGJ2-GS and 15dPGJ2-Cys in mouse and human immune cells, the involvement of microsomal glutathione S-transferase 3 in their biosynthesis, and their unchanged formation following inhibition of mPGES-1. The results encourage further research on their roles as bioactive lipid mediators.
Asunto(s)
Cisteína , Prostaglandinas , Ratones , Humanos , Animales , Lipopolisacáridos/metabolismo , Mastocitos , Prostaglandina-E Sintasas/metabolismo , Macrófagos/metabolismo , Ciclooxigenasa 2/metabolismo , Glutatión/metabolismo , Glutatión Transferasa/metabolismo , Prostaglandina D2/farmacologíaRESUMEN
B cells play a significant role in established Rheumatoid Arthritis (RA). However, it is unclear to what extent differentiated B cells are present in joint tissue already at the onset of disease. Here, we studied synovial biopsies (n = 8) captured from untreated patients at time of diagnosis. 3414 index-sorted B cells underwent RNA sequencing and paired tissue pieces were subjected to spatial transcriptomics (n = 4). We performed extensive bioinformatics analyses to dissect the local B cell composition. Select plasma cell immunoglobulin sequences were expressed as monoclonal antibodies and tested by ELISA. Memory and plasma cells were found irrespective of autoantibody status of the patients. Double negative memory B cells were prominent, but did not display a distinct transcriptional profile. The tissue architecture implicate both local B cell maturation via T cell help and plasma cell survival niches with a strong CXCL12-CXCR4 axis. The immunoglobulin sequence analyses revealed clonality between the memory B and plasma cell pools further supporting local maturation. One of the plasma cell-derived antibodies displayed citrulline autoreactivity, demonstrating local autoreactive plasma cell differentiation in joint biopsies captured from untreated early RA. Hence, plasma cell niches are not a consequence of chronic inflammation, but are already present at the time of diagnosis.
Asunto(s)
Artritis Reumatoide , Membrana Sinovial , Autoanticuerpos , Diferenciación Celular , Humanos , Membrana Sinovial/patología , TranscriptomaRESUMEN
Chinese medicine has a long tradition of use against rheumatoid arthritis (RA). The formulations are based on combinations of typically 5-10 plants, which are usually boiled and administered as a decoction or tea. There are few clinical trials performed so the clinical evidence is sparse. One fundamental of traditional medicine is to prevent disease. RA is an autoimmune, inflammatory and chronic disease that primarily affects the joints of 0.5%-1% of the population. In two out of three of the cases, the patients are characterised by the presence of autoantibodies such as the rheumatoid factor and the more disease-specific autoantibody against citrullinated proteins, so-called 'ACPA' (anticitrullinated protein/peptide antibodies). ACPA positivity is also strongly associated with specific variations in the HLA-DRB1 gene, the shared epitope alleles. Together with smoking, these factors account for the major risks of developing RA. In this review, we will summarise the background using certain plant-based formulations based on Chinese traditional medicine for the treatment and prevention of RA and the strategy we have taken to explore the mechanisms of action. We also summarise the major pathophysiological pathways related to RA and how these could be analysed. Finally, we summarise our ideas on how a clinical trial using Chinese herbal medicine to prevent RA could be conducted.
Asunto(s)
Artritis Reumatoide , Medicamentos Herbarios Chinos , Alelos , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/prevención & control , Autoanticuerpos , Ensayos Clínicos como Asunto , Medicamentos Herbarios Chinos/uso terapéutico , Epítopos/genética , Predisposición Genética a la Enfermedad , Cadenas HLA-DRB1/genética , Humanos , Medicina Tradicional China , Péptidos , Factor Reumatoide/genética , TéRESUMEN
The inflamed rheumatic joint is a highly heterogeneous and complex tissue with dynamic recruitment and expansion of multiple cell types that interact in multifaceted ways within a localized area. Rheumatoid arthritis synovium has primarily been studied either by immunostaining or by molecular profiling after tissue homogenization. Here, we use Spatial Transcriptomics, where tissue-resident RNA is spatially labeled in situ with barcodes in a transcriptome-wide fashion, to study local tissue interactions at the site of chronic synovial inflammation. We report comprehensive spatial RNA-Seq data coupled to cell type-specific localization patterns at and around organized structures of infiltrating leukocyte cells in the synovium. Combining morphological features and high-throughput spatially resolved transcriptomics may be able to provide higher statistical power and more insights into monitoring disease severity and treatment-specific responses in seropositive and seronegative rheumatoid arthritis.
Asunto(s)
Artritis Reumatoide , Transcriptoma , Artritis Reumatoide/genética , Artritis Reumatoide/metabolismo , Humanos , Membrana Sinovial/metabolismoRESUMEN
BACKGROUND: The microvasculature is a target organ for the early manifestations of cardiovascular disease. Therefore, a better understanding of the prostaglandin system and characterising the effects of mPGES-1 inhibition and concomitant reduction of PGE2 in vascular beds are of interest. EXPERIMENTAL APPROACH: The effects of mPGES-1 inhibition on constriction and relaxation of resistance arteries (diameter: 100-400 µm) from patients with end stage kidney disease (ESKD) and controls (Non-ESKD) were studied using wire-myography in combination with immunological and mass-spectrometry based analyses. KEY RESULTS: Inhibition of mPGES-1 in arteries from ESKD patients and Non-ESKD controls significantly reduced adrenergic vasoconstriction, which was unaffected by the COX-2 inhibitors NS-398 and Etoricoxib, or by the COX-1/COX-2 inhibitor Indomethacin tested in Non-ESKD controls. However, a significant increase of acetylcholine-induced dilatation was observed for mPGES-1 inhibition. In IL-1ß treated arteries, inhibition of mPGES-1 significantly reduced PGE2 levels while PGI2 levels remained unchanged. In contrast, COX-2 inhibition blocked the formation of both prostaglandins. Blockade of PGI2 signalling with an IP receptor antagonist did not restore the reduced adrenergic constriction, neither did blocking PGE2 -EP4 or signalling through PPARγ. A biphasic effect was observed for PGE2 , inducing dilatation at nanomolar and constriction at micromolar concentrations. Immunohistochemistry demonstrated expression of mPGES-1, COX-1, PGIS, weak expression for COX-2, as well as receptor expression for PGE2 (EP1-4), thromboxane (TP) and PGI2 (IP) in ESKD and Non-ESKD. CONCLUSION: Our study demonstrates vasodilating effects following mPGES-1 inhibition in human microvasculature and suggests that several pathways besides shunting to PGI2 are involved.
Asunto(s)
Arterias , Fallo Renal Crónico , Prostaglandina-E Sintasas , Adrenérgicos , Arterias/metabolismo , Arterias/fisiología , Ciclooxigenasa 1/metabolismo , Ciclooxigenasa 2/metabolismo , Inhibidores de la Ciclooxigenasa 2/farmacología , Etoricoxib , Humanos , Fallo Renal Crónico/complicaciones , Microvasos/metabolismo , Microvasos/fisiología , Nitrobencenos , Prostaglandina-E Sintasas/antagonistas & inhibidores , Prostaglandinas , SulfonamidasRESUMEN
Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by synovial inflammation and joint destruction. Cell-derived small extracellular vesicles (sEV) mediate cell-to-cell communication in the synovial microenvironment by carrying microRNAs (miRs), a class of small non-coding RNAs. Herein, we report that sEV from synovial fluid promote osteoclast differentiation which is attributed to high levels of extracellular miR-574-5p. Moreover, we demonstrate for the first time that enhanced osteoclast maturation is mediated by Toll-like receptor (TLR) 7/8 signaling which is activated by miR-574-5p binding. This is a novel mechanism by which sEV and miRs contribute to RA pathogenesis and indicate that pharmacological inhibition of extracellular miR-574-5p might offer new therapeutic strategies to protect osteoclast-mediated bone destruction in RA.
Asunto(s)
Artritis Reumatoide , MicroARNs/metabolismo , Osteoclastos/metabolismo , Receptor Toll-Like 7/metabolismo , Receptor Toll-Like 8/metabolismo , Artritis Reumatoide/inmunología , Artritis Reumatoide/metabolismo , Artritis Reumatoide/patología , Diferenciación Celular/fisiología , Vesículas Extracelulares/metabolismo , Células HEK293 , Células HeLa , Humanos , Osteoclastos/patología , Osteogénesis/fisiología , Líquido Sinovial/metabolismoRESUMEN
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.
Asunto(s)
Antineoplásicos/farmacología , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/patología , Animales , Fibroblastos Asociados al Cáncer/metabolismo , Fibroblastos Asociados al Cáncer/patología , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ciclooxigenasa 2/metabolismo , Dinoprostona/metabolismo , Modelos Animales de Enfermedad , Sistemas de Liberación de Medicamentos/métodos , Evaluación Preclínica de Medicamentos/métodos , Femenino , Humanos , Ratones , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Neuroblastoma/metabolismo , Prostaglandina-E Sintasas/metabolismoRESUMEN
AIMS: Cardiovascular side effects caused by non-steroidal anti-inflammatory drugs (NSAIDs), which all inhibit cyclooxygenase (COX)-2, have prevented development of new drugs that target prostaglandins to treat inflammation and cancer. Microsomal prostaglandin E synthase-1 (mPGES-1) inhibitors have efficacy in the NSAID arena but their cardiovascular safety is not known. Our previous work identified asymmetric dimethylarginine (ADMA), an inhibitor of endothelial nitric oxide synthase, as a potential biomarker of cardiovascular toxicity associated with blockade of COX-2. Here, we have used pharmacological tools and genetically modified mice to delineate mPGES-1 and COX-2 in the regulation of ADMA. METHODS AND RESULTS: Inhibition of COX-2 but not mPGES-1 deletion resulted in increased plasma ADMA levels. mPGES-1 deletion but not COX-2 inhibition resulted in increased plasma prostacyclin levels. These differences were explained by distinct compartmentalization of COX-2 and mPGES-1 in the kidney. Data from prostanoid synthase/receptor knockout mice showed that the COX-2/ADMA axis is controlled by prostacyclin receptors (IP and PPARß/δ) and the inhibitory PGE2 receptor EP4, but not other PGE2 receptors. CONCLUSION: These data demonstrate that inhibition of mPGES-1 spares the renal COX-2/ADMA pathway and define mechanistically how COX-2 regulates ADMA.
Asunto(s)
Aorta/enzimología , Arginina/análogos & derivados , Ciclooxigenasa 2/metabolismo , Riñón/enzimología , Prostaglandina-E Sintasas/metabolismo , Animales , Aorta/efectos de los fármacos , Arginina/sangre , Inhibidores de la Ciclooxigenasa 2/farmacología , Sistema Enzimático del Citocromo P-450/genética , Sistema Enzimático del Citocromo P-450/metabolismo , Femenino , Oxidorreductasas Intramoleculares/genética , Oxidorreductasas Intramoleculares/metabolismo , Riñón/efectos de los fármacos , Masculino , Ratones Noqueados , PPAR-beta/genética , PPAR-beta/metabolismo , Prostaglandina-E Sintasas/antagonistas & inhibidores , Prostaglandina-E Sintasas/genética , Prostaglandinas I/sangre , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Epoprostenol/genética , Receptores de Epoprostenol/metabolismo , Receptores de Prostaglandina E/genética , Receptores de Prostaglandina E/metabolismoRESUMEN
Lately it has become possible to analyze transcriptomic profiles in tissue sections with retained cellular context. We aimed to explore synovial biopsies from rheumatoid arthritis (RA) and spondyloarthritis (SpA) patients, using Spatial Transcriptomics (ST) as a proof of principle approach for unbiased mRNA studies at the site of inflammation in these chronic inflammatory diseases. Synovial tissue biopsies from affected joints were studied with ST. The transcriptome data was subjected to differential gene expression analysis (DEA), pathway analysis, immune cell type identification using Xcell analysis and validation with immunohistochemistry (IHC). The ST technology allows selective analyses on areas of interest, thus we analyzed morphologically distinct areas of mononuclear cell infiltrates. The top differentially expressed genes revealed an adaptive immune response profile and T-B cell interactions in RA, while in SpA, the profiles implicate functions associated with tissue repair. With spatially resolved gene expression data, overlaid on high-resolution histological images, we digitally portrayed pre-selected cell types in silico. The RA displayed an overrepresentation of central memory T cells, while in SpA effector memory T cells were most prominent. Consequently, ST allows for deeper understanding of cellular mechanisms and diversity in tissues from chronic inflammatory diseases.
Asunto(s)
Artritis Reumatoide/inmunología , Espondiloartritis/inmunología , Membrana Sinovial/inmunología , Transcriptoma/inmunología , Adulto , Artritis Reumatoide/patología , Biopsia , Femenino , Perfilación de la Expresión Génica , Humanos , Inflamación/inmunología , Inflamación/patología , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Espondiloartritis/patología , Membrana Sinovial/patologíaRESUMEN
BACKGROUND AND PURPOSE: Microsomal PGE synthase-1 (mPGES-1), the inducible synthase that catalyses the terminal step in PGE2 biosynthesis, is of high interest as therapeutic target to treat inflammation. Inhibition of mPGES-1 is suggested to be safer than traditional NSAIDs, and recent data demonstrate anti-constrictive effects on vascular tone, indicating new therapeutic opportunities. However, there is a lack of potent mPGES-1 inhibitors lacking interspecies differences for conducting in vivo studies in relevant preclinical disease models. EXPERIMENTAL APPROACH: Potency was determined based on the reduction of PGE2 formation in recombinant enzyme assays, cellular assay, human whole blood assay, and air pouch mouse model. Anti-inflammatory properties were assessed by acute paw swelling in a paw oedema rat model. Effect on vascular tone was determined with human ex vivo wire myography. KEY RESULTS: We report five new mPGES-1 inhibitors (named 934, 117, 118, 322, and 323) that selectively inhibit recombinant human and rat mPGES-1 with IC50 values of 10-29 and 67-250 nM respectively. The compounds inhibited PGE2 production in a cellular assay (IC50 values 0.15-0.82 µM) and in a human whole blood assay (IC50 values 3.3-8.7 µM). Moreover, the compounds blocked PGE2 formation in an air pouch mouse model and reduced acute paw swelling in a paw oedema rat model. Human ex vivo wire myography analysis showed reduced adrenergic vasoconstriction after incubation with the compounds. CONCLUSION AND IMPLICATIONS: These mPGES-1 inhibitors can be used as refined tools in further investigations of the role of mPGES-1 in inflammation and microvascular disease.
Asunto(s)
Antiinflamatorios/farmacología , Arterias/efectos de los fármacos , Dinoprostona/biosíntesis , Edema/tratamiento farmacológico , Inhibidores Enzimáticos/farmacología , Tono Muscular/efectos de los fármacos , Prostaglandina-E Sintasas/antagonistas & inhibidores , Células A549 , Animales , Antiinflamatorios/química , Antiinflamatorios/farmacocinética , Arterias/enzimología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Edema/inmunología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacocinética , Escherichia coli/genética , Humanos , Técnicas In Vitro , Masculino , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Contracción Muscular/efectos de los fármacos , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/enzimología , Miografía , Prostaglandina-E Sintasas/sangre , Prostaglandina-E Sintasas/genéticaRESUMEN
Pharmacological inhibition of microsomal prostaglandin E synthase (mPGES)-1 for selective reduction in prostaglandin E2 (PGE2) biosynthesis is protective in experimental models of cancer and inflammation. Targeting mPGES-1 is envisioned as a safer alternative to traditional non-steroidal anti-inflammatory drugs (NSAIDs). Herein, we compared the effects of mPGES-1 inhibitor Compound III (CIII) with the cyclooxygenase (COX)-2 inhibitor NS-398 on protein and lipid profiles in interleukin (IL)-1ß-induced A549 lung cancer cells using mass spectrometry. Inhibition of mPGES-1 decreased PGE2 production and increased PGF2α and thromboxane B2 (TXB2) formation, while inhibition of COX-2 decreased the production of all three prostanoids. Our proteomics results revealed that CIII downregulated multiple canonical pathways including eIF2, eIF4/P70S6K, and mTOR signaling, compared to NS-398 that activated these pathways. Moreover, pathway analysis predicted that CIII increased cell death of cancer cells (Z = 3.8, p = 5.1E-41) while NS-398 decreased the same function (Z = -5.0, p = 6.5E-35). In our lipidomics analyses, we found alterations in nine phospholipids between the two inhibitors, with a stronger alteration in the lysophospholipid (LPC) profile with NS-398 compared to CIII. Inhibition of mPGES-1 increased the concentration of sphinganine and dihydroceramide (C16:0DhCer), while inhibition of COX-2 caused a general decrease in most ceramides, again suggesting different effects on cell death between the two inhibitors. We showed that CIII decreased proliferation and potentiated the cytotoxic effect of the cytostatic drugs cisplatin, etoposide, and vincristine when investigated in a live cell imaging system. Our results demonstrate differences in protein and lipid profiles after inhibition of mPGES-1 or COX-2 with important implications on the therapeutic potential of mPGES-1 inhibitors as adjuvant treatment in cancer. We encourage further investigations to illuminate the clinical benefit of mPGES-1 inhibitors in cancer.
RESUMEN
MicroRNAs (miRs) are important posttranscriptional regulators of gene expression. Besides their well-characterized inhibitory effects on mRNA stability and translation, miRs can also activate gene expression. In this study, we identified a novel noncanonical function of miR-574-5p. We found that miR-574-5p acts as an RNA decoy to CUG RNA-binding protein 1 (CUGBP1) and antagonizes its function. MiR-574-5p induces microsomal prostaglandin E synthase-1 (mPGES-1) expression by preventing CUGBP1 binding to its 3'UTR, leading to an enhanced alternative splicing and generation of an mPGES-1 3'UTR isoform, increased mPGES-1 protein expression, PGE2 formation, and tumor growth in vivo. miR-574-5p-induced tumor growth in mice could be completely inhibited with the mPGES-1 inhibitor CIII. Moreover, miR-574-5p is induced by IL-1ß and is strongly overexpressed in human nonsmall cell lung cancer where high mPGES-1 expression correlates with a low survival rate. The discovered function of miR-574-5p as a CUGBP1 decoy opens up new therapeutic opportunities. It might serve as a stratification marker to select lung tumor patients who respond to the pharmacological inhibition of PGE2 formation.-Saul, M. J., Baumann, I., Bruno, A., Emmerich, A. C., Wellstein, J., Ottinger, S. M., Contursi, A., Dovizio, M., Donnini, S., Tacconelli, S., Raouf, J., Idborg, H., Stein, S., Korotkova, M., Savai, R., Terzuoli, E., Sala, G., Seeger, W., Jakobsson, P.-J., Patrignani, P., Suess, B., Steinhilber, D. miR-574-5p as RNA decoy for CUGBP1 stimulates human lung tumor growth by mPGES-1 induction.
Asunto(s)
Proteínas CELF1/metabolismo , MicroARNs/metabolismo , Prostaglandina-E Sintasas/metabolismo , ARN/metabolismo , Células A549 , Animales , Proteínas CELF1/genética , Proliferación Celular , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HeLa , Humanos , Ratones , Ratones Desnudos , MicroARNs/genética , Imitación Molecular , Neoplasias Experimentales , Prostaglandina-E Sintasas/genética , Unión Proteica , Inhibidores de la Síntesis de la Proteína/farmacología , Puromicina/farmacología , ARN/genética , Interferencia de ARN , Isoformas de ARN , ARN MensajeroRESUMEN
BACKGROUND: Polymyositis (PM) and dermatomyositis (DM) are severe chronic autoimmune diseases, characterized by muscle fatigue and low muscle endurance. Conventional treatment includes high doses of glucocorticoids and immunosuppressive drugs; however, few patients recover full muscle function. One explanation of the persistent muscle weakness could be altered lipid metabolism in PM/DM muscle tissue as we previously reported. Using a targeted lipidomic approach we aimed to characterize serum lipid profiles in patients with PM/DM compared to healthy individuals (HI) in a cross-sectional study. Also, in the longitudinal study we compared serum lipid profiles in patients newly diagnosed with PM/DM before and after immunosuppressive treatment. METHODS: Lipidomic profiles were analyzed in serum samples from 13 patients with PM/DM, 12 HI and 8 patients newly diagnosed with PM/DM before and after conventional immunosuppressive treatment using liquid chromatography tandem mass spectrometry (LC-MS/MS) and a gas-chromatography flame ionization detector (GC-FID). Functional Index (FI), as a test of muscle performance and serum levels of creatine kinase (s-CK) as a proxy for disease activity were analyzed. RESULTS: The fatty acid (FA) composition of total serum lipids was altered in patients with PM/DM compared to HI; the levels of palmitic (16:0) acid were significantly higher while the levels of arachidonic (20:4, n-6) acid were significantly lower in patients with PM/DM. The profiles of serum phosphatidylcholine and triacylglycerol species were changed in patients with PM/DM compared to HI, suggesting disproportionate levels of saturated and polyunsaturated FAs that might have negative effects on muscle performance. After immunosuppressive treatment the total serum lipid levels of eicosadienoic (20:2, n-6) and eicosapentaenoic (20:5, n-3) acids were increased and serum phospholipid profiles were altered in patients with PM/DM. The correlation between FI or s-CK and levels of several lipid species indicate the important role of lipid changes in muscle performance and inflammation. CONCLUSIONS: Serum lipids profiles are significantly altered in patients with PM/DM compared to HI. Moreover, immunosuppressive treatment in patients newly diagnosed with PM/DM significantly affected serum lipid profiles. These findings provide new evidence of the dysregulated lipid metabolism in patients with PM/DM that could possibly contribute to low muscle performance.
Asunto(s)
Dermatomiositis/tratamiento farmacológico , Inmunosupresores/uso terapéutico , Metabolismo de los Lípidos/efectos de los fármacos , Lípidos/sangre , Metabolómica/métodos , Polimiositis/tratamiento farmacológico , Adulto , Estudios Transversales , Dermatomiositis/sangre , Dermatomiositis/metabolismo , Ácidos Grasos/sangre , Femenino , Humanos , Lípidos/química , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/fisiopatología , Evaluación de Resultado en la Atención de Salud/métodos , Evaluación de Resultado en la Atención de Salud/estadística & datos numéricos , Polimiositis/sangre , Polimiositis/metabolismoRESUMEN
Despite recent progress in diagnosis and treatment, survival for children with high-risk metastatic neuroblastoma is still poor. Prostaglandin E2 (PGE2)-driven inflammation promotes tumor growth, immune suppression, angiogenesis and resistance to established cancer therapies. In neuroblastoma, cancer-associated fibroblasts (CAFs) residing in the tumor microenvironment are the primary source of PGE2. However, clinical targeting of PGE2 with current non-steroidal anti-inflammatory drugs or cyclooxygenase inhibitors has been limited due to risk of adverse side effects. By specifically targeting microsomal prostaglandin E synthase-1 (mPGES-1) activity with a small molecule inhibitor we could block CAF-derived PGE2 production leading to reduced tumor growth, impaired angiogenesis, inhibited CAF migration and infiltration, reduced tumor cell proliferation and a favorable shift in the M1/M2 macrophage ratio. In this study, we provide proof-of-principle of the benefits of targeting mPGES-1 in neuroblastoma, applicable to a wide variety of tumors. This non-toxic single drug treatment targeting infiltrating stromal cells opens up for combination treatment options with established cancer therapies.