RESUMEN
Prostaglandin E2 (PGE2) is known to be effective in regenerating tissues, and bimatoprost, an analog of PGF2α, has been approved by the FDA as an eyelash growth promoter and has been proven effective in human hair follicles. Thus, to enhance PGE2 levels while improving hair loss, we found dihydroisoquinolinone piperidinylcarboxy pyrazolopyridine (DPP), an inhibitor of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), using DeepZema®, an AI-based drug development program. Here, we investigated whether DPP improved hair loss in human follicle dermal papilla cells (HFDPCs) damaged by dihydrotestosterone (DHT), which causes hair loss. We found that DPP enhanced wound healing and the expression level of alkaline phosphatase in DHT-damaged HFDPCs. We observed that DPP significantly down-regulated the generation of reactive oxygen species caused by DHT. DPP recovered the mitochondrial membrane potential in DHT-damaged HFDPCs. We demonstrated that DPP significantly increased the phosphorylation levels of the AKT/ERK and activated Wnt signaling pathways in DHT-damaged HFDPCs. We also revealed that DPP significantly enhanced the size of the three-dimensional spheroid in DHT-damaged HFDPCs and increased hair growth in ex vivo human hair follicle organ culture. These data suggest that DPP exhibits beneficial effects on DHT-damaged HFDPCs and can be utilized as a promising agent for improving hair loss.
Asunto(s)
Folículo Piloso , Hidroxiprostaglandina Deshidrogenasas , Humanos , Folículo Piloso/efectos de los fármacos , Folículo Piloso/metabolismo , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Dihidrotestosterona/farmacología , Dihidrotestosterona/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Dermis/metabolismo , Dermis/citología , Dermis/efectos de los fármacos , Células Cultivadas , Vía de Señalización Wnt/efectos de los fármacos , Alopecia/tratamiento farmacológico , Alopecia/metabolismo , Cicatrización de Heridas/efectos de los fármacos , Cabello/efectos de los fármacos , Cabello/crecimiento & desarrollo , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Inhibidores Enzimáticos/farmacologíaRESUMEN
The enzyme 15-hydroxyprostaglandin dehydrogenase (15-PGDH), which acts as a negative regulator of prostaglandin E2 (PGE2) levels and activity, represents a promising pharmacological target for promoting liver regeneration. In this study, we collected data on 15-PGDH homologous family proteins, their inhibitors, and traditional Chinese medicine (TCM) compounds. Leveraging machine learning and molecular docking techniques, we constructed a prediction model for virtual screening of 15-PGDH inhibitors from TCM compound library and successfully screened genistein as a potential 15-PGDH inhibitor. Through further validation, it was discovered that genistein considerably enhances liver regeneration by inhibiting 15-PGDH, resulting in a significant increase in the PGE2 level. Genistein's effectiveness suggests its potential as a novel therapeutic agent for liver diseases, highlighting this study's contribution to expanding the clinical applications of TCM.
Asunto(s)
Inhibidores Enzimáticos , Hidroxiprostaglandina Deshidrogenasas , Regeneración Hepática , Medicina Tradicional China , Simulación del Acoplamiento Molecular , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Animales , Regeneración Hepática/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Humanos , Dinoprostona/metabolismo , Simulación por Computador , Genisteína/farmacología , Genisteína/química , Masculino , Evaluación Preclínica de Medicamentos , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/química , Ratones , Aprendizaje AutomáticoRESUMEN
15-prostaglandin dehydrogenase (15-PGDH) is a negative regulator of tissue stem cells that acts via enzymatic activity of oxidizing and degrading PGE2, and related eicosanoids, that support stem cells during tissue repair. Indeed, inhibiting 15-PGDH markedly accelerates tissue repair in multiple organs. Here we have used cryo-electron microscopy to solve the solution structure of native 15-PGDH and of 15-PGDH individually complexed with two distinct chemical inhibitors. These structures identify key 15-PGDH residues that mediate binding to both classes of inhibitors. Moreover, we identify a dynamic 15-PGDH lid domain that closes around the inhibitors, and that is likely fundamental to the physiologic 15-PGDH enzymatic mechanism. We furthermore identify two key residues, F185 and Y217, that act as hinges to regulate lid closing, and which both inhibitors exploit to capture the lid in the closed conformation, thus explaining their sub-nanomolar binding affinities. These findings provide the basis for further development of 15-PGDH targeted drugs as therapeutics for regenerative medicine.
Asunto(s)
Eicosanoides , Hidroxiprostaglandina Deshidrogenasas , Microscopía por Crioelectrón , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidoresRESUMEN
15-Prostaglandin dehydrogenase (15-PGDH) regulates the concentration of prostaglandin E2 in vivo. Inhibitors of 15-PGDH elevate PGE2 levels and promote tissue repair and regeneration. Here, we describe a novel class of quinoxaline amides that show potent inhibition of 15-PGDH, good oral bioavailability, and protective activity in mouse models of ulcerative colitis and recovery from bone marrow transplantation.
Asunto(s)
Hidroxiprostaglandina Deshidrogenasas , Quinoxalinas , Animales , Ratones , Colitis Ulcerosa/tratamiento farmacológico , Dinoprostona , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Quinoxalinas/farmacologíaRESUMEN
15-hydroxyprostaglandin dehydrogenase (15-PGDH; encoded by HPGD) is ubiquitously expressed in mammalian tissues and catalyzes the degradation of prostaglandins (PGs; mainly PGE2, PGD2, and PGF2α) in a process mediated by solute carrier organic anion transport protein family member 2A1 (SLCO2A1; also known as PGT, OATP2A1, PHOAR2, or SLC21A2). As a key enzyme, 15-PGDH catalyzes the rapid oxidation of 15-hydroxy-PGs into 15-keto-PGs with lower biological activity. Increasing evidence suggests that 15-PGDH plays a key role in many physiological and pathological processes in mammals and is considered a potential pharmacological target for preventing organ damage, promoting bone marrow graft recovery, and enhancing tissue regeneration. Additionally, results of whole-exome analyses suggest that recessive inheritance of an HPGD mutation is associated with idiopathic hypertrophic osteoarthropathy. Interestingly, as a tumor suppressor, 15-PGDH inhibits proliferation and induces the differentiation of cancer cells (including those associated with colorectal, lung, and breast cancers). Furthermore, a recent study identified 15-PGDH as a marker of aging tissue and a potential novel therapeutic target for resisting the complex pathology of aging-associated diseases. Here, we review and summarise recent information on the molecular functions of 15-PGDH and discuss its pathophysiological implications.
Asunto(s)
Envejecimiento/fisiología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Prostaglandinas/metabolismo , Animales , Biomarcadores/metabolismo , Hidroxiprostaglandina Deshidrogenasas/genéticaRESUMEN
We discovered SW033291 in a high throughput chemical screen aimed at identifying 15-prostaglandin dehydrogenase (15-PGDH) modulators. The compound exhibited inhibitory activity in in vitro biochemical and cell-based assays of 15-PGDH activity. We subsequently demonstrated that this compound, and several analogs thereof, are effective in in vivo mouse models of bone marrow transplant, colitis, and liver regeneration, where increased levels of PGE2 positively potentiate tissue regeneration. To better understand the binding of SW033291, we carried out docking studies for both the substrate, PGE2, and an inhibitor, SW033291, to 15-PGDH. Our models suggest similarities in the ways that PGE2 and SW033291 interact with key residues in the 15-PGDH-NAD+ complex. We carried out molecular dynamics simulations (MD) of SW033291 bound to this complex, in order to understand the dynamics of the binding interactions for this compound. The butyl side chain (including the sulfoxide) of SW033291 participates in crucial binding interactions that are similar to those observed for the C15-OH and the C16-C20 alkyl chain of PGE2. In addition, interactions with residues Ser138, Tyr151, and Gln148 play key roles in orienting and stabilizing SW033291 in the binding site and lead to enantioselectivity for the R-enantiomer. Finally, we compare the binding mode of (R)-S(O)-SW033291 with the binding interactions of published 15-PGDH inhibitors.
Asunto(s)
Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Piridinas/química , Piridinas/farmacología , Tiofenos/química , Tiofenos/farmacología , Sitios de Unión , Humanos , Simulación de Dinámica MolecularRESUMEN
The splenic microenvironment regulates hematopoietic stem and progenitor cell (HSPC) function, particularly during demand-adapted hematopoiesis; however, practical strategies to enhance splenic support of transplanted HSPCs have proved elusive. We have previously demonstrated that inhibiting 15-hydroxyprostaglandin dehydrogenase (15-PGDH), using the small molecule (+)SW033291 (PGDHi), increases BM prostaglandin E2 (PGE2) levels, expands HSPC numbers, and accelerates hematologic reconstitution after BM transplantation (BMT) in mice. Here we demonstrate that the splenic microenvironment, specifically 15-PGDH high-expressing macrophages, megakaryocytes (MKs), and mast cells (MCs), regulates steady-state hematopoiesis and potentiates recovery after BMT. Notably, PGDHi-induced neutrophil, platelet, and HSPC recovery were highly attenuated in splenectomized mice. PGDHi induced nonpathologic splenic extramedullary hematopoiesis at steady state, and pretransplant PGDHi enhanced the homing of transplanted cells to the spleen. 15-PGDH enzymatic activity localized specifically to macrophages, MK lineage cells, and MCs, identifying these cell types as likely coordinating the impact of PGDHi on splenic HSPCs. These findings suggest that 15-PGDH expression marks HSC niche cell types that regulate hematopoietic regeneration. Therefore, PGDHi provides a well-tolerated strategy to therapeutically target multiple HSC niches, promote hematopoietic regeneration, and improve clinical outcomes of BMT.
Asunto(s)
Células de la Médula Ósea/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Hematopoyesis Extramedular/efectos de los fármacos , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Regeneración , Bazo/efectos de los fármacos , Animales , Células de la Médula Ósea/citología , Femenino , Perfilación de la Expresión Génica , Ratones , Ratones Endogámicos C57BL , Bazo/enzimología , Bazo/metabolismoRESUMEN
The two primary mechanisms by which iodinated contrast media (CM) causes contrast-induced acute kidney injury (CIAKI) are the hemodynamic effect causing intrarenal vasoconstriction and the tubular toxic effect causing acute tubular necrosis. Inhibition of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), which degrades prostaglandin E2 (PGE2), promotes tissue repair and regeneration in many organs. PGE2 causes intrarenal arterial vasodilation. In this study, we investigated whether a 15-PGDH inhibitor can act as a candidate for blocking these two major mechanisms of CIAKI. We established a CIAKI mouse model by injecting a 10 gram of iodine per body weight (gI/kg) dose of iodixanol into each mouse tail vein. A 15-PGDH inhibitor (SW033291), PGE1, or PGE2 were administered to compare the renal functional parameters, histologic injury, vasoconstriction, and renal blood flow changes. In addition, human renal proximal tubular epithelial cells were cultured in a CM-treated medium. SW033291, PGE1, or PGE2 were added to compare any changes in cell viability and apoptosis rate. CIAKI mice that received SW033291 had lower serum levels of creatinine, neutrophil gelatinase-associated lipocalin, and kidney injury molecule 1 (p < 0.001); lower histologic injury score and TUNEL positive rates (p < 0.001); and higher medullary arteriolar area (p < 0.05) and renal blood flow (p < 0.001) than CM + vehicle group. In cell culture experiments, Adding SW033291 increased the viability rate (p < 0.05) and decreased the apoptosis rate of the tubular epithelial cells (p < 0.001). This 15-PGDH inhibitor blocks the two primary mechanisms of CIAKI, intrarenal vasoconstriction and tubular cell toxicity, and thus has the potential to be a novel prophylaxis for CIAKI. Abbreviations: 15-PGDH: 15-hydroxyprostaglandin dehydrogenase; AMP: adenosine monophosphate; CIAKI: contrast-induced acute kidney injury; CM: contrast media; EP: prostaglandin E2 receptor; hRPTECs: human-derived renal proximal tubule epithelial cells; KIM-1: kidney injury molecule-1; MTT: 3-(4,5-Dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide; NGAL: neutrophil gelatinase-associated lipocalin; PBS: phosphate-buffered saline; PGE1: prostaglandin E1; PGE2: prostaglandin E2; RBF: renal blood flow; TUNEL: terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling; α-SMA: α-Smooth muscle actin.
Asunto(s)
Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/prevención & control , Medios de Contraste/efectos adversos , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Piridinas/farmacología , Tiofenos/farmacología , Animales , Creatinina/sangre , Femenino , Humanos , Riñón/fisiopatología , Lipocalina 2/sangre , Ratones , Ratones Endogámicos C57BL , Prostaglandinas E/farmacología , Ácidos Triyodobenzoicos/efectos adversosRESUMEN
Solidagenone (SOL) is a labdane-type diterpenoid found in Solidago chilensis, a plant traditionally used to treat skin diseases, kidney pain and ovarian inflammation. In this study, the topical anti-inflammatory activity of SOL was evaluated using in vivo and in silico assays. Croton oil-, arachidonic acid (AA)- and phenol-induced ear oedema mouse models were applied in the in vivo studies. Myeloperoxidase (MPO) and N-acetyl-ß-D-glucosaminidase (NAG) activities and tumour necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and nitric oxide (NO) levels were determined, as well as histopathological analyses were conducted. Interaction profiles between SOL and cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), glucocorticoid receptor, estradiol-17-ß-dehydrogenase and prostaglandin-E(2)-9-reductase were established using molecular docking. SOL significantly inhibited croton oil-, AA- and phenol-induced ear oedema (P < .001) at doses of 0.1, 0.5 and 1.0 mg/ear. The MPO and NAG activities and TNF-α, IL-6 and NO levels were decreased (P < .001). The histopathological data revealed that inflammatory parameters (oedema thickness, leucocyte infiltration and vasodilatation) were reduced by treatment with SOL at doses of 0.1, 0.5 and 1.0 mg/ear. The docking study showed that SOL interacts with COX-1 and prostaglandin-E(2)-9-reductase through hydrogen bonding, inhibiting these enzymes. These results indicate that SOL may be a promising compound for the treatment of cutaneous inflammatory disorders and has potential as a topical anti-inflammatory agent.
Asunto(s)
Inhibidores de la Ciclooxigenasa/farmacología , Dermatitis/prevención & control , Edema/prevención & control , Furanos/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Proteínas de la Membrana/antagonistas & inhibidores , Naftalenos/farmacología , Extractos Vegetales/farmacología , Piel/efectos de los fármacos , Solidago , Acetilglucosaminidasa/metabolismo , Animales , Ciclooxigenasa 1/metabolismo , Inhibidores de la Ciclooxigenasa/aislamiento & purificación , Inhibidores de la Ciclooxigenasa/metabolismo , Dermatitis/metabolismo , Dermatitis/patología , Modelos Animales de Enfermedad , Edema/inducido químicamente , Edema/metabolismo , Edema/patología , Furanos/aislamiento & purificación , Furanos/metabolismo , Enlace de Hidrógeno , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Interleucina-6/metabolismo , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Simulación del Acoplamiento Molecular , Naftalenos/aislamiento & purificación , Naftalenos/metabolismo , Óxido Nítrico/metabolismo , Peroxidasa/metabolismo , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/metabolismo , Unión Proteica , Transducción de Señal , Piel/metabolismo , Piel/patología , Solidago/química , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Treatments are lacking for sarcopenia, a debilitating age-related skeletal muscle wasting syndrome. We identifed increased amounts of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), the prostaglandin E2 (PGE2)-degrading enzyme, as a hallmark of aged tissues, including skeletal muscle. The consequent reduction in PGE2 signaling contributed to muscle atrophy in aged mice and results from 15-PGDH-expressing myofibers and interstitial cells, such as macrophages, within muscle. Overexpression of 15-PGDH in young muscles induced atrophy. Inhibition of 15-PGDH, by targeted genetic depletion or a small-molecule inhibitor, increased aged muscle mass, strength, and exercise performance. These benefits arise from a physiological increase in PGE2 concentrations, which augmented mitochondrial function and autophagy and decreased transforming growth factor-ß signaling and activity of ubiquitin-proteasome pathways. Thus, PGE2 signaling ameliorates muscle atrophy and rejuvenates muscle function, and 15-PGDH may be a suitable therapeutic target for countering sarcopenia.
Asunto(s)
Envejecimiento/metabolismo , Dinoprostona/metabolismo , Hidroxiprostaglandina Deshidrogenasas/fisiología , Músculo Esquelético/patología , Rejuvenecimiento , Sarcopenia/enzimología , Animales , Muerte Celular Autofágica/genética , Muerte Celular Autofágica/fisiología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/genética , Macrófagos/enzimología , Ratones , Ratones Endogámicos C57BL , Mitocondrias Musculares/ultraestructura , Fuerza Muscular/genética , Fuerza Muscular/fisiología , Músculo Esquelético/enzimología , Miofibrillas/enzimología , Sarcopenia/genéticaRESUMEN
In the present study, we demonstrated the marked activity of SW033291, an inhibitor of 15-hydoxyprostaglandin dehydrogenase (15-PGDH), in preventing acute kidney injury (AKI) in a murine model of ischemia-reperfusion injury. AKI due to ischemic injury represents a significant clinical problem. PGE2 is vasodilatory in the kidney, but it is rapidly degraded in vivo due to catabolism by 15-PGDH. We investigated the potential of SW033291, a potent and specific 15-PGDH inhibitor, as prophylactic treatment for ischemic AKI. Prophylactic administration of SW033291 significantly increased renal tissue PGE2 levels and increased post-AKI renal blood flow and renal arteriole area. In parallel, prophylactic SW033291 decreased post-AKI renal morphology injury scores and tubular apoptosis and markedly reduced biomarkers of renal injury that included blood urea nitrogen, creatinine, neutrophil gelatinase-associated lipocalin, and kidney injury molecule-1. Prophylactic SW033291 also reduced post-AKI induction of proinflammatory cytokines, high-mobility group box 1, and malondialdehyde. Protective effects of SW033291 were mediated by PGE2 signaling, as they could be blocked by pharmacological inhibition of PGE2 synthesis. Consistent with activation of PGE2 signaling, SW033291 induced renal levels of both EP4 receptors and cAMP, along with other vasodilatory effectors, including AMP, adenosine, and the adenosine A2A receptor. The protective effects of SW0333291 could largely be achieved with a single prophylactic dose of the drug. Inhibition of 15-PGDH may thus represent a novel strategy for prophylaxis of ischemic AKI in multiple clinical settings, including renal transplantation and cardiovascular surgery.
Asunto(s)
Lesión Renal Aguda/prevención & control , Adenosina/metabolismo , Dinoprostona/metabolismo , Inhibidores Enzimáticos/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Riñón/irrigación sanguínea , Riñón/efectos de los fármacos , Piridinas/farmacología , Receptor de Adenosina A2A/metabolismo , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Circulación Renal/efectos de los fármacos , Daño por Reperfusión/prevención & control , Tiofenos/farmacología , Vasodilatación/efectos de los fármacos , Lesión Renal Aguda/enzimología , Lesión Renal Aguda/patología , Lesión Renal Aguda/fisiopatología , Animales , Velocidad del Flujo Sanguíneo , Modelos Animales de Enfermedad , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Riñón/enzimología , Riñón/patología , Masculino , Ratones Endogámicos C57BL , Daño por Reperfusión/enzimología , Daño por Reperfusión/patología , Daño por Reperfusión/fisiopatología , Transducción de SeñalRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a progressive disease characterized by interstitial remodeling and pulmonary dysfunction. The etiology of IPF is not completely understood but involves pathologic inflammation and subsequent failure to resolve fibrosis in response to epithelial injury. Treatments for IPF are limited to anti-inflammatory and immunomodulatory agents, which are only partially effective. Prostaglandin E2 (PGE2) disrupts TGFß signaling and suppresses myofibroblast differentiation, however practical strategies to raise tissue PGE2 during IPF have been limited. We previously described the discovery of a small molecule, (+)SW033291, that binds with high affinity to the PGE2-degrading enzyme 15-hydroxyprostaglandin dehydrogenase (15-PGDH) and increases PGE2 levels. Here we evaluated pulmonary 15-PGDH expression and activity and tested whether pharmacologic 15-PGDH inhibition (PGDHi) is protective in a mouse model of bleomycin-induced pulmonary fibrosis (PF). Long-term PGDHi was well-tolerated, reduced the severity of pulmonary fibrotic lesions and extracellular matrix remodeling, and improved pulmonary function in bleomycin-treated mice. Moreover, PGDHi attenuated both acute inflammation and weight loss, and decreased mortality. Endothelial cells and macrophages are likely targets as these cell types highly expressed 15-PGDH. In conclusion, PGDHi ameliorates inflammatory pathology and fibrosis in murine PF, and may have clinical utility to treat human disease.
Asunto(s)
Antiinflamatorios/farmacología , Dinoprostona/metabolismo , Inhibidores Enzimáticos/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Piridinas/farmacología , Tiofenos/farmacología , Animales , Bleomicina/administración & dosificación , Peso Corporal/efectos de los fármacos , Dinoprostona/agonistas , Modelos Animales de Enfermedad , Células Endoteliales/efectos de los fármacos , Células Endoteliales/enzimología , Células Endoteliales/patología , Matriz Extracelular/efectos de los fármacos , Matriz Extracelular/enzimología , Femenino , Expresión Génica , Humanos , Hidroxiprostaglandina Deshidrogenasas/genética , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Fibrosis Pulmonar Idiopática/inducido químicamente , Fibrosis Pulmonar Idiopática/enzimología , Fibrosis Pulmonar Idiopática/mortalidad , Inflamación , Pulmón/efectos de los fármacos , Pulmón/enzimología , Pulmón/patología , Macrófagos/efectos de los fármacos , Macrófagos/enzimología , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Terapia Molecular Dirigida/métodos , Pruebas de Función Respiratoria , Análisis de SupervivenciaRESUMEN
Diabetes-induced hyperglycemia plays a key pathogenic role in degenerative retinal diseases. In diabetic hyperglycemia, aldose reductase (AR) is elevated and linked to the pathogenesis of diabetic retinopathy (DR) and cataract. Retinal microglia (RMG), the resident immune cells in the retina, are thought to contribute to the proinflammatory phenotype in the diabetic eye. However, we have a limited understanding of the potential role of AR expressed in RMG as a mediator of inflammation in the diabetic retina. Glycated proteins accumulate in diabetes, including Amadori-glycated albumin (AGA) which has been shown to induce a proinflammatory phenotype in various tissues. In this study, we investigated the ability of AGA to stimulate inflammatory changes to RMG and macrophages, and whether AR plays a role in this process. In macrophages, treatment with an AR inhibitor (Sorbinil) or genetic knockdown of AR lowered AGA-induced TNF-α secretion (56% and 40%, respectively) as well as cell migration. In a mouse RMG model, AR inhibition attenuated AGA-induced TNF-α secretion and cell migration (67% and 40%, respectively). To further mimic the diabetic milieu in retina, we cultured RMG under conditions of hypoxia and observed the induction of TNF-α and VEGF protein expression. Downregulation of AR in either a pharmacological or genetic manner prevented hypoxia-induced TNF-α and VEGF expression. In our animal study, increased numbers of RMG observed in streptozotocin (STZ)-induced diabetic retina was substantially lower when diabetes was induced in AR knockout mice. Thus, in vitro and in vivo studies demonstrated that AR is involved in diabetes-induced RMG activation, providing a rationale for targeting AR as a therapeutic strategy for DR.
Asunto(s)
Diabetes Mellitus Experimental/patología , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Animales , Hipoxia de la Célula , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Diabetes Mellitus Experimental/inducido químicamente , Regulación hacia Abajo/efectos de los fármacos , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/genética , Imidazolidinas/farmacología , Macrófagos/citología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Microglía/citología , Microglía/metabolismo , Células RAW 264.7 , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Retina/citología , Factor de Necrosis Tumoral alfa/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Whether sensory nerve can sense bone density or metabolic activity to control bone homeostasis is unknown. Here we found prostaglandin E2 (PGE2) secreted by osteoblastic cells activates PGE2 receptor 4 (EP4) in sensory nerves to regulate bone formation by inhibiting sympathetic activity through the central nervous system. PGE2 secreted by osteoblasts increases when bone density decreases as demonstrated in osteoporotic animal models. Ablation of sensory nerves erodes the skeletal integrity. Specifically, knockout of the EP4 gene in the sensory nerves or cyclooxygenase-2 (COX2) in the osteoblastic cells significantly reduces bone volume in adult mice. Sympathetic tone is increased in sensory denervation models, and propranolol, a ß2-adrenergic antagonist, rescues bone loss. Furthermore, injection of SW033291, a small molecule to increase PGE2 level locally, significantly boostes bone formation, whereas the effect is obstructed in EP4 knockout mice. Thus, we show that PGE2 mediates sensory nerve to control bone homeostasis and promote regeneration.
Asunto(s)
Huesos/metabolismo , Dinoprostona/metabolismo , Osteoporosis/patología , Subtipo EP4 de Receptores de Prostaglandina E/metabolismo , Células Receptoras Sensoriales/metabolismo , Fibras Adrenérgicas/efectos de los fármacos , Fibras Adrenérgicas/metabolismo , Antagonistas Adrenérgicos beta/farmacología , Animales , Densidad Ósea/efectos de los fármacos , Regeneración Ósea/efectos de los fármacos , Huesos/citología , Huesos/inervación , Huesos/patología , Células Cultivadas , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Modelos Animales de Enfermedad , Retroalimentación Fisiológica , Femenino , Humanos , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteoporosis/etiología , Propranolol/farmacología , Piridinas/farmacología , Subtipo EP4 de Receptores de Prostaglandina E/genética , Células Receptoras Sensoriales/efectos de los fármacos , Tiofenos/farmacologíaRESUMEN
Hematopoietic stem cell transplantation following myeloablative chemotherapy is a curative treatment for many hematopoietic malignancies. However, profound granulocytopenia during the interval between transplantation and marrow recovery exposes recipients to risks of fatal infection, a significant source of transplant-associated morbidity and mortality. We have previously described the discovery of a small molecule, SW033291, that potently inhibits the prostaglandin degrading enzyme 15-PGDH, increases bone marrow prostaglandin E2, and accelerates hematopoietic recovery following murine transplant. Here we describe the efficacy of (+)-SW209415, a second-generation 15-PGDH inhibitor, in an expanded range of models relevant to human transplantation. (+)-SW209415 is 10,000-fold more soluble, providing the potential for intravenous delivery, while maintaining potency in inhibiting 15-PGDH, increasing in vivo prostaglandin E2, and accelerating hematopoietic regeneration following transplantation. In additional models, (+)-SW209415: (i) demonstrated synergy with granulocyte colony-stimulating factor, the current standard of care; (ii) maintained efficacy as transplant cell dose was escalated; (iii) maintained efficacy when transplant donors and recipients were aged; and (iv) potentiated homing in xenotransplants using human hematopoietic stem cells. (+)-SW209415 showed no adverse effects, no potentiation of in vivo growth of human myeloma and leukemia xenografts, and, on chronic high-dose administration, no toxicity as assessed by weight, blood counts and serum chemistry. These studies provide independent chemical confirmation of the activity of 15-PGDH inhibitors in potentiating hematopoietic recovery, extend the range of models in which inhibiting 15-PGDH demonstrates activity, allay concerns regarding potential for adverse effects from increasing prostaglandin E2, and thereby, advance 15-PGDH as a therapeutic target for potentiating hematopoietic stem cell transplantation.
Asunto(s)
Antineoplásicos/farmacología , Inhibidores Enzimáticos/farmacología , Supervivencia de Injerto/efectos de los fármacos , Factor Estimulante de Colonias de Granulocitos/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Adulto , Factores de Edad , Animales , Trasplante de Médula Ósea , Femenino , Trasplante de Células Madre Hematopoyéticas , Xenoinjertos , Humanos , Masculino , RatonesRESUMEN
Nicotinamide adenine dinucleotide (NAD) levels decrease with aging as a result of aging-associated CD38 upregulation. Here, we established a cell model with decreased cellular NAD levels by overexpressing CD38 or treating cells with FK866, an inhibitor of nicotinamide phosphoribosyltransferase. We revealed that decreased NAD triggered reactive oxygen species (ROS)-mediated degradation of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), which drove cells to undergo epithelial-mesenchymal transition (EMT). Moreover, we showed that oxidation of the Cys44 residue to sulfonic acid in 15-PGDH led to its degradation via non-canonical ubiquitination-proteasome and autophagy pathways. Mutation of Cys44 to alanine abolished ROS-induced 15-PGDH degradation. We demonstrated that 15-PGDH silencing promoted EMT, whereas supplementation with NAD precursors increased NAD and 15-PGDH stability, and reversed the EMT process. Taken together, these results suggest that declining NAD levels contribute to age-dependent increases in cancer incidence, and repletion of NAD precursors is beneficial for increasing 15-PGDH expression.
Asunto(s)
Cisteína/química , Transición Epitelial-Mesenquimal , Hidroxiprostaglandina Deshidrogenasas/metabolismo , NAD/metabolismo , Especies Reactivas de Oxígeno/metabolismo , ADP-Ribosil Ciclasa 1/genética , ADP-Ribosil Ciclasa 1/metabolismo , Acrilamidas/farmacología , Autofagia , Línea Celular Tumoral , Movimiento Celular , Cisteína/metabolismo , Dinoprostona/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Hidroxiprostaglandina Deshidrogenasas/genética , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Mutagénesis Sitio-Dirigida , Niacina/farmacología , Oxidación-Reducción , Piperidinas/farmacología , Complejo de la Endopetidasa Proteasomal/metabolismo , Estabilidad Proteica , Ácidos Sulfónicos/químicaRESUMEN
The aldo-keto reductase 1C3 isoform (AKR1C3) plays a vital role in the biosynthesis of androgens, making this enzyme an attractive target for castration-resistant prostate cancer therapy. Although AKR1C3 is a promising drug target, no AKR1C3-targeted agent has to date been approved for clinical use. Flufenamic acid, a non-steroidal anti-inflammatory drug, is known to potently inhibit AKR1C3 in a non-selective manner as COX off-target effects are also observed. To diminish off-target effects, we have applied a scaffold hopping strategy replacing the benzoic acid moiety of flufenamic acid with an acidic hydroxyazolecarbonylic scaffold. In particular, differently N-substituted hydroxylated triazoles were designed to simultaneously interact with both subpockets 1 and 2 in the active site of AKR1C3, larger for AKR1C3 than other AKR1Cs isoforms. Through computational design and iterative rounds of synthesis and biological evaluation, novel compounds are reported, sharing high selectivity (up to 230-fold) for AKR1C3 over 1C2 isoform and minimal COX1 and COX2 off-target inhibition. A docking study of compound 8, the most interesting compound of the series, suggested that its methoxybenzyl substitution has the ability to fit inside subpocket 2, being involved in π-π staking interaction with Trp227 (partial overlapping) and in a T-shape π-π staking with Trp86. This compound was also shown to diminish testosterone production in the AKR1C3-expressing 22RV1 prostate cancer cell line while synergistic effect was observed when 8 was administered in combination with abiraterone or enzalutamide.
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3-Hidroxiesteroide Deshidrogenasas/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Triazoles/farmacología , 3-Hidroxiesteroide Deshidrogenasas/metabolismo , Miembro C3 de la Familia 1 de las Aldo-Ceto Reductasas , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Humanos , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Modelos Moleculares , Estructura Molecular , Relación Estructura-Actividad , Triazoles/síntesis química , Triazoles/química , Células Tumorales CultivadasRESUMEN
OBJECTIVE: To find an inhibitor of 15-hydroxyprostaglandin dehydrogenase (15-PGDH) that rapidly metabolises Prostaglandin E2 (PGE2) as a mediator of wound healing, we examined seven flavonoids for this role. RESULTS: 7,3',4'-Trimethoxyflavone (TMF) had the lowest IC50 value of 0.34 µM for 15-PGDH inhibition but >400 µM for cytotoxicity, indicating a high therapeutic index. TMF elevated PGE2 levels in a concentration-dependent manner in both A549 lung cancer and HaCaT cells. It also significantly increased mRNA expression of multidrug resistance-associated protein 4 (MRP4) and of prostaglandin transporter (PGT) slightly in HaCaT cells. In addition, TMF facilitated in vitro wound healing in a HaCaT scratch model, which was completely inhibited by adding both 15-PGDH and NAD+ as cofactor, confirming the involvement of PGE2 in its wound healing effect. CONCLUSION: TMF with a high therapeutic index can facilitate wound healing through PGE2 elevation by 15-PGDH inhibition.
Asunto(s)
Dinoprostona/metabolismo , Flavonas/farmacología , Hidroxiprostaglandina Deshidrogenasas/antagonistas & inhibidores , Cicatrización de Heridas/efectos de los fármacos , Células A549 , Línea Celular , Relación Dosis-Respuesta a Droga , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , NAD/metabolismo , Transportadores de Anión Orgánico/genéticaRESUMEN
17beta-hydroxysteroid dehydrogenase type 5 (17ß-HSD5) is an important enzyme associated with sex steroid metabolism in hormone-dependent cancer. However, reports on its expression and its prognostic value in breast cancer are inconsistent. Here, we demonstrate the impact of 17ß-HSD5 expression modulation on the proteome of estrogen receptor-positive (ER+) breast cancer cells. RNA interference technique (siRNA) was used to knock down 17ß-HSD5 gene expression in the ER+ breast cancer cell line MCF-7 and the proteome of the 17ß-HSD5-knockdown cells was compared to that of MCF-7 cells using two-dimensional (2-D) gel electrophoresis followed by mass spectrometry analysis. Ingenuity pathway analysis (IPA) was additionally used to assess functional enrichment analyses of the proteomic dataset, including protein network and canonical pathways. Our proteomic analysis revealed only four differentially expressed protein spots (fold change > 2, p<0.05) between the two cell lines. The four spots were up-regulated in 17ß-HSD5-knockdown MCF-7 cells, and comprised 21 proteins involved in two networks and in functions that include apoptosis inhibition, regulation of cell growth and differentiation, signal transduction and tumor metastasis. Among the proteins are nucleoside diphosphate kinase A (NME1), 78kDa glucose-regulated protein (GRP78) and phosphoglycerate kinase 1 (PGK1). We also showed that expression of 17ß-HSD5 and that of the apoptosis inhibitor GRP78 are strongly but negatively correlated. Consistent with their opposite regulation, GRP78 knockdown decreased MCF-7 cell viability whereas 17ß-HSD5 knockdown or inhibition increased cell viability and proliferation. Besides, IPA analysis revealed that ubiquitination pathway is significantly affected by 17ß-HSD5 knockdown. Furthermore, IPA predicted the proto-oncogene c-Myc as an upstream regulator linked to the tumor-secreted protein PGK1. The latter is over-expressed in invasive ductal breast carcinoma as compared with normal breast tissue and its expression increased following 17ß-HSD5 knockdown. Our present results indicate a 17ß-HSD5 role in down-regulating breast cancer development. We thus propose that 17ß-HSD5 may not be a potent target for breast cancer treatment but its low expression could represent a poor prognosis factor.