RESUMEN

BACKGROUND: Epidermal remodeling and hypertrophy are hallmarks of skin fibrotic disorders, and keratinocyte to mesenchymal (EMT)-like transformations drive epidermis alteration in skin fibrosis such as keloids and hypertrophic scars (HTS). While phosphodiesterase 4 (PDE4) inhibitors have shown effectiveness in various fibrotic disorders, their role in skin fibrosis is not fully understood. This study aimed to explore the specific role of PDE4B in epidermal remodeling and hypertrophy seen in skin fibrosis. METHODS: In vitro experiments examined the effects of inhibiting PDE4A-D (with Roflumilast) or PDE4B (with siRNA) on TGFß1-induced EMT differentiation and dedifferentiation in human 3D epidermis. In vivo studies investigated the impact of PDE4 inhibition on HOCl-induced skin fibrosis and epidermal hypertrophy in mice, employing both preventive and therapeutic approaches. RESULTS: The study found increased levels of PDE4B (mRNA, protein) in keloids > HTS compared to healthy epidermis, as well as in TGFß-stimulated 3D epidermis. Keloids and HTS epidermis exhibited elevated levels of collagen Iα1, fibronectin, αSMA, N-cadherin, and NOX4 mRNA, along with decreased levels of E-cadherin and ZO-1, confirming an EMT process. Inhibition of both PDE4A-D and PDE4B prevented TGFß1-induced Smad3 and ERK1/2 phosphorylation and mesenchymal differentiation in vitro. PDE4A-D inhibition also promoted mesenchymal dedifferentiation and reduced TGFß1-induced ROS and keratinocyte senescence by rescuing PPM1A, a Smad3 phosphatase. In vivo, PDE4 inhibition mitigated HOCl-induced epidermal hypertrophy in mice in both preventive and therapeutic settings. CONCLUSIONS: Overall, the study supports the potential of PDE4 inhibitors, particularly PDE4B, in treating skin fibrosis, including keloids and HTS, shedding light on their functional role in this condition.

Asunto(s)

Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Fibrosis , Queloide , Queratinocitos , Inhibidores de Fosfodiesterasa 4 , Humanos , Queloide/metabolismo , Queloide/patología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Queratinocitos/metabolismo , Queratinocitos/efectos de los fármacos , Inhibidores de Fosfodiesterasa 4/farmacología , Animales , Ratones , Epidermis/metabolismo , Epidermis/patología , Factor de Crecimiento Transformador beta1/metabolismo , Transición Epitelial-Mesenquimal/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Masculino

RESUMEN

Sulfonylureas (SUs) are a class of antidiabetic drugs widely used in the management of diabetes mellitus type 2. They promote insulin secretion by inhibiting the ATP-sensitive potassium channel in pancreatic ß-cells. Recently, the exchange protein directly activated by cAMP (Epac) was identified as a new class of target proteins of SUs that might contribute to their antidiabetic effect, through the activation of the Ras-like guanosine triphosphatase Rap1, which has been controversially discussed. We used human embryonic kidney (HEK) 293 cells expressing genetic constructs of various Förster resonance energy transfer (FRET)-based biosensors containing different versions of Epac1 and Epac2 isoforms, alone or fused to different phosphodiesterases (PDEs), to monitor SU-induced conformational changes in Epac or direct PDE inhibition in real time. We show that SUs can both induce conformational changes in the Epac2 protein but not in Epac1, and directly inhibit the PDE3 and PDE4 families, thereby increasing cAMP levels in the direct vicinity of these PDEs. Furthermore, we demonstrate that the binding site of SUs in Epac2 is distinct from that of cAMP and is located between the amino acids E443 and E460. Using biochemical assays, we could also show that tolbutamide can inhibit PDE activity through an allosteric mechanism. Therefore, the cAMP-elevating capacity due to allosteric PDE inhibition in addition to direct Epac activation may contribute to the therapeutic effects of SU drugs.

Asunto(s)

AMP Cíclico , Factores de Intercambio de Guanina Nucleótido , Compuestos de Sulfonilurea , Humanos , Compuestos de Sulfonilurea/farmacología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Células HEK293 , AMP Cíclico/metabolismo , Transferencia Resonante de Energía de Fluorescencia , Inhibidores de Fosfodiesterasa/farmacología , Inhibidores de Fosfodiesterasa/química , Hipoglucemiantes/farmacología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Sitios de Unión , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 3/metabolismo

RESUMEN

Phosphodiesterase 4 (PDE4) enzymes catalyze cyclic adenosine monophosphate (cAMP) hydrolysis and are involved in a variety of physiological processes, including brain function, monocyte and macrophage activation, and neutrophil infiltration. Among different PDE4 isoforms, Phosphodiesterases 4D (PDE4Ds) play a fundamental role in cognitive, learning and memory consolidation processes and cancer development. Selective PDE4D inhibitors (PDE4Dis) could represent an innovative and valid therapeutic strategy for the treatment of various neurodegenerative diseases, such as Alzheimer's, Parkinson's, Huntington's, and Lou Gehrig's diseases, but also for stroke, traumatic brain and spinal cord injury, mild cognitive impairment, and all demyelinating diseases such as multiple sclerosis. In addition, small molecules able to block PDE4D isoforms have been recently studied for the treatment of specific cancer types, particularly hepatocellular carcinoma and breast cancer. This review overviews the PDE4DIsso far identified and provides useful information, from a medicinal chemistry point of view, for the development of a novel series of compounds with improved pharmacological properties.

Asunto(s)

Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Inhibidores de Fosfodiesterasa 4 , Humanos , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/uso terapéutico , Inhibidores de Fosfodiesterasa 4/química , Animales , Enfermedades Neurodegenerativas/tratamiento farmacológico , Enfermedades Neurodegenerativas/metabolismo , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo

RESUMEN

Cyclic nucleotide phosphodiesterases (PDEs) consist of a family of enzymes expressed in several types of cells, including inflammatory cells, that play a pivotal role in inflammation. Several studies have demonstrated that the inhibition of PDE4 results in a reduced inflammatory response via PKA and CREB signaling. Hence, PDE4 suppression improves the inflammatory feedback typical of several diseases, such as inflammatory bowel disease (IBD). In our previous studies, we have demonstrated that miR-369-3p regulates inflammatory responses, modulating different aspects of the inflammatory process. The aim of this study was to demonstrate an additional anti-inflammatory effect of miR-369-3p targeting PDE4B, one of the widely expressed isoforms in immune cells. We found that miR-369-3p was able to reduce the expression of PDE4B, elevating the intracellular levels of cAMP. This accumulation increased the expression of PKA and pCREB, mitigating the release of pro-inflammatory cytokines and promoting the release of anti-inflammatory cytokines. To prove that PDE4B is a good therapeutic target in IBD, we also demonstrate that the expression of PDE4B was increased in UC patients compared to healthy controls, affecting the immune infiltrate. PDE4B is considered an important player in inflammatory progression; hence, our results show the ability of miR-369-3p to ameliorate inflammation by targeting PDE4B, supporting its future application as a new therapeutic approach in IBD.

Asunto(s)

Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Enfermedades Inflamatorias del Intestino , MicroARNs , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , AMP Cíclico/metabolismo , Inflamación/genética , Inflamación/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Citocinas/metabolismo , Masculino , Transducción de Señal , Femenino

RESUMEN

Loss of retinal ganglion cells (RGCs) is central to the pathogenesis of optic neuropathies such as glaucoma. Increased RGC cAMP signaling is neuroprotective. We have shown that displacement of the cAMP-specific phosphodiesterase PDE4D3 from an RGC perinuclear compartment by expression of the modified PDE4D3 N-terminal peptide 4D3(E) increases perinuclear cAMP and protein kinase A activity in cultured neurons and in vivo RGC survival after optic nerve crush (ONC) injury. To explore mechanisms by which PDE4D3 displacement promotes neuroprotection, in this study mice intravitreally injected with an adeno-associated virus to express an mCherry-tagged 4D3(E) peptide were subjected to ONC injury and analyzed by single cell RNA-sequencing (scRNA-seq). 4D3(E)-mCherry expression was associated with an attenuation of injury-induced changes in gene expression, thereby supporting the hypothesis that enhanced perinuclear PKA signaling promotes neuroprotective RGC gene expression.

Asunto(s)

Ratones Endogámicos C57BL , Traumatismos del Nervio Óptico , Células Ganglionares de la Retina , Células Ganglionares de la Retina/metabolismo , Células Ganglionares de la Retina/patología , Células Ganglionares de la Retina/efectos de los fármacos , Animales , Ratones , Traumatismos del Nervio Óptico/metabolismo , Traumatismos del Nervio Óptico/tratamiento farmacológico , Traumatismos del Nervio Óptico/genética , Regulación de la Expresión Génica , Modelos Animales de Enfermedad , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Compresión Nerviosa , Supervivencia Celular , Inyecciones Intravítreas , Transducción de Señal , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Masculino , Células Cultivadas

RESUMEN

There is a pressing medical need for improved treatments in skin fibrosis including keloids and hypertrophic scars (HTS). This study aimed to characterize the role of phosphodiesterase 4 (PDE4), specifically PDE4B in fibrotic skin remodeling in vitro and in vivo. In vitro, effects of PDE4A-D (Roflumilast) or PDE4B (siRNA) inhibition on TGFß1-induced myofibroblast differentiation and dedifferentiation were studied in normal (NHDF) and keloid (KF) human dermal fibroblasts. In vivo, the role of PDE4 on HOCl-induced skin fibrosis in mice was addressed in preventive and therapeutic protocols. PDE4B (mRNA, protein) was increased in Keloid > HTS compared to healthy skin and in TGFß-stimulated NHDF and KF. In Keloid > HTS, collagen Iα1, αSMA, TGFß1 and NOX4 mRNA were all elevated compared to healthy skin confirming skin fibrosis. In vitro, inhibition of PDE4A-D and PDE4B similarly prevented TGFß1-induced Smad3 and ERK1/2 phosphorylation and myofibroblast differentiation, elevated NOX4 protein and proliferation in NHDF. PDE4A-D inhibition enabled myofibroblast dedifferentiation and curbed TGFß1-induced reactive oxygen species and fibroblast senescence. In KF PDE4A-D inhibition restrained TGFß1-induced Smad3 and ERK1/2 phosphorylation, myofibroblast differentiation and senescence. Mechanistically, PDE4A-D inhibition rescued from TGFß1-induced loss in PPM1A, a Smad3 phosphatase. In vivo, PDE4 inhibition mitigated HOCl-induced skin fibrosis in mice in preventive and therapeutic protocols. The current study provides novel evidence evolving rationale for PDE4 inhibitors in skin fibrosis (including keloids and HTS) and delivered evidence for a functional role of PDE4B in this fibrotic condition.

Asunto(s)

Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Fibroblastos , Fibrosis , Queloide , Inhibidores de Fosfodiesterasa 4 , Piel , Factor de Crecimiento Transformador beta1 , Queloide/patología , Queloide/metabolismo , Humanos , Animales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fibroblastos/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/patología , Ratones , Inhibidores de Fosfodiesterasa 4/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , Piel/patología , Piel/metabolismo , Piel/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Masculino , Células Cultivadas , NADPH Oxidasa 4/metabolismo , NADPH Oxidasa 4/antagonistas & inhibidores , NADPH Oxidasa 4/genética , Ácido Hipocloroso/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Proteína smad3/metabolismo , Proliferación Celular/efectos de los fármacos , Femenino

RESUMEN

Inflammatory bowel disease (IBD) is a chronic and progressive condition with a significant global burden. Currently, available treatments primarily provide symptomatic relief and retard disease progression, yet they do not offer a cure and are frequently associated with adverse effects. Therefore, the discovery of new targets and therapeutic drugs for IBD is crucial. Phosphodiesterase 4 (PDE4) inhibitors have emerged as promising candidates in the search for effective IBD treatments, although dose-dependent side effects hamper their clinical utility. In this study, building upon heterocyclic biaryl derivatives (TPA16), we designed and synthesized a series of N2-substituted indazole-based PDE4D inhibitors, emphasizing improving safety profiles. An enzyme activity screening discovered an optimized compound, LZ-14 (Z21115), which exhibited high PDE4D7 (IC50 = 10.5 nM) inhibitory activity and good selectivity. More interestingly, LZ-14 has demonstrated promising effects in treating IBD in mouse models by improving the inflammatory response and colon injury. Furthermore, LZ-14 displayed low emetogenic potential in ketamine/xylazine anesthesia mice alternative models.

Asunto(s)

Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Descubrimiento de Drogas , Indazoles , Enfermedades Inflamatorias del Intestino , Inhibidores de Fosfodiesterasa 4 , Animales , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Relación Estructura-Actividad , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/química , Inhibidores de Fosfodiesterasa 4/síntesis química , Inhibidores de Fosfodiesterasa 4/uso terapéutico , Ratones , Indazoles/farmacología , Indazoles/química , Indazoles/síntesis química , Humanos , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Estructura Molecular , Relación Dosis-Respuesta a Droga , Masculino , Ratones Endogámicos C57BL

RESUMEN

Immune-mediated inflammatory diseases (IMIDs) comprise a broad spectrum of conditions characterized by systemic inflammation affecting various organs and tissues, for which there is no known cure. The isoform-specific inhibition of phosphodiesterase-4B (PDE4B) over PDE4D constitutes an effective therapeutic strategy for the treatment of IMIDs that minimizes the adverse effects associated with non-selective PDE4 inhibitors. Thus, we report a new class of isoquinolone derivatives as next-generation PDE4 inhibitors for effective management of rheumatoid arthritis (RA) and psoriasis. Among the series, 8 compounds i.e. 1e, 1l, 1m, 1n, 1o, 2m, 2o and 3o showed promising PDE4B inhibition (>80 %) in vitro with IC50 ∼ 1.4-6.2 µM. The compound 1l was identified as an initial hit and was pursued for further studies. According to structure-activity relationship (SAR), an allyl group at C-4 position improved PDE4B inhibition. The correlation between in vitro activity data and binding affinities obtained via molecular docking suggested that the high-affinity binding to PDE4B is a prerequisite for the effective inhibition of PDE4B. Notably, the hit 1l showed selectivity towards PDE4B over PDE4D in vitro. Furthermore, 1l treatment (30 mg/kg) in the adjuvant-induced arthritis (AIA) rat model induced by complete Freund's adjuvant (CFA) demonstrated anti-arthritic potential via ameliorating paw swelling and body weight, narrowing joint space, reducing excessive immune cells infiltration and pannus formation in addition to reducing mRNA expression of pro-inflammatory cytokines such as TNF-α and IL-6 in synovial tissues of experimental rats. Additionally, 1l reduced the hyper-proliferative state and colony forming potential of IMQ-induced psoriatic keratinocytes. The treatment of these cells with 1l markedly reduced the protein levels of Ki67 and mRNA levels of pro-inflammatory cytokines e.g. IL-17A and TNF-α suggesting its potent anti-psoriatic potential. Furthermore, 1l did not show any significant adverse effects when evaluated in a systematic toxicity (e.g. teratogenicity, hepatotoxicity and cardiotoxicity) studies in zebrafish at the tested concentrations (1-100 µM) and the NOAEL (no-observed-adverse-effect level) was found to be 100 µM. Thus, with promising anti-inflammatory effects both in vitro and in vivo along with PDE4B selectivity with an acceptable safety margin, 1l emerged as a new and promising inhibitor for further studies.

Asunto(s)

Artritis Reumatoide , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Inhibidores de Fosfodiesterasa 4 , Psoriasis , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/química , Inhibidores de Fosfodiesterasa 4/síntesis química , Inhibidores de Fosfodiesterasa 4/uso terapéutico , Psoriasis/tratamiento farmacológico , Artritis Reumatoide/tratamiento farmacológico , Animales , Relación Estructura-Actividad , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Humanos , Ratas , Estructura Molecular , Relación Dosis-Respuesta a Droga , Simulación del Acoplamiento Molecular , Pez Cebra , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/inducido químicamente , Artritis Experimental/patología , Masculino

RESUMEN

Background and Objectives: Colorectal cancer (CRC) is the most frequently diagnosed malignant disease of the gastrointestinal system, and new diagnostic and prognostic markers are needed to elucidate the complete tumor profile. Materials and Methods: We used CRC tumor tissues (Dukes' A-D) and adjacent noncancerous tissues of 43 patients. Immunohistochemistry was used to examine the expression of phosphodiesterase 4B (PDE4B), phosphodiesterase 4D (PDE4D), and secreted frizzled related protein 5 (SFRP5) markers. We also analyzed the expression levels of PDE4B, PDE4D, and SFRP5 in CRC tissues compared to control tissues using RNA-sequencing data from the UCSC Xena browser. Results: In CRC stages, the distribution of PDE4B-positive cells varied, with differing percentages between epithelium and lamina propria. Statistically significant differences were found in the number of PDE4B-positive epithelial cells between healthy controls and all CRC stages, as well as between different CRC stages. Similarly, significant differences were observed in the number of PDE4B-positive cells in the lamina propria between healthy controls and all CRC stages, as well as between different CRC stages. CRC stage Dukes' C exhibited a significantly higher number of PDE4B-positive cells in the lamina propria compared to CRC stage Dukes' B. Significant differences were noted in the number of PDE4D-positive epithelial cells between healthy controls and CRC stages Dukes' A, B, and D, as well as between CRC stage Dukes' C and stages A, B, and D. CRC stage Dukes' A had significantly more PDE4D-positive cells in the lamina propria compared to stage D. Significant differences were also observed in the number of SFRP5-positive cells in the lamina propria between healthy controls and all CRC stages, as well as between CRC stages Dukes' A and D. While the expression of PDE4D varied across CRC stages, the expression of SFRP5 remained consistently strong in both epithelium and lamina propria, with significant differences noted mainly in the lamina propria. The expression levels of PDE4B, PDE4D, and SFRP5 reveal significant differences in the expression of these genes between CRC patients and healthy controls, with notable implications for patient prognosis. Namely, our results demonstrate that PDE4B, PDE4D, and SFRP5 are significantly under-expressed in CRC tissues compared to control tissues. The Kaplan-Meier survival analysis and the log-rank (Mantel-Cox) test revealed distinct prognostic implications where patients with lower expression levels of SFRP5 exhibited significantly longer overall survival. The data align with our immunohistochemical results and might suggest a potential tumor-suppressive role for these genes in CRC. Conclusions: Considering significantly lower gene expression, aligned with our immunohistochemical data in tumor tissue in comparison to the control tissue, as well as the significantly poorer survival rate in the case of its higher expression, we can hypothesize that SFRP5 is the most promising biomarker for CRC out of the observed proteins. These findings suggest alterations in PDE4B, PDE4D, and SFRP5 expression during CRC progression, as well as between different stages of CRC, with potential implications for understanding the molecular mechanisms involved in CRC development and progression.

Asunto(s)

Biomarcadores de Tumor , Neoplasias Colorrectales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Humanos , Neoplasias Colorrectales/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Masculino , Femenino , Persona de Mediana Edad , Biomarcadores de Tumor/análisis , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Anciano , Proteínas Adaptadoras Transductoras de Señales/análisis , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Inmunohistoquímica/métodos , Adulto , Proteínas de la Membrana/análisis , Proteínas de la Membrana/genética

RESUMEN

BACKGROUND: ß-adrenergic receptor (ß-AR) overactivation is a major pathological cue associated with cardiac injury and diseases. AMPK (AMP-activated protein kinase), a conserved energy sensor, regulates energy metabolism and is cardioprotective. However, whether AMPK exerts cardioprotective effects via regulating the signaling pathway downstream of ß-AR remains unclear. METHODS: Using immunoprecipitation, mass spectrometry, site-specific mutation, in vitro kinase assay, and in vivo animal studies, we determined whether AMPK phosphorylates ß-arrestin-1 at serine (Ser) 330. Wild-type mice and mice with site-specific mutagenesis (S330A knock-in [KI]/S330D KI) were subcutaneously injected with the ß-AR agonist isoproterenol (5 mg/kg) to evaluate the causality between ß-adrenergic insult and ß-arrestin-1 Ser330 phosphorylation. Cardiac transcriptomics was used to identify changes in gene expression from ß-arrestin-1-S330A/S330D mutation and ß-adrenergic insult. RESULTS: Metformin could decrease cAMP/PKA (protein kinase A) signaling induced by isoproterenol. AMPK bound to ß-arrestin-1 and phosphorylated Ser330 with the highest phosphorylated mass spectrometry score. AMPK activation promoted ß-arrestin-1 Ser330 phosphorylation in vitro and in vivo. Neonatal mouse cardiomyocytes overexpressing ß-arrestin-1-S330D (active form) inhibited the ß-AR/cAMP/PKA axis by increasing PDE (phosphodiesterase) 4 expression and activity. Cardiac transcriptomics revealed that the differentially expressed genes between isoproterenol-treated S330A KI and S330D KI mice were mainly involved in immune processes and inflammatory response. ß-arrestin-1 Ser330 phosphorylation inhibited isoproterenol-induced reactive oxygen species production and NLRP3 (NOD-like receptor protein 3) inflammasome activation in neonatal mouse cardiomyocytes. In S330D KI mice, the ß-AR-activated cAMP/PKA pathways were attenuated, leading to repressed inflammasome activation, reduced expression of proinflammatory cytokines, and mitigated macrophage infiltration. Compared with S330A KI mice, S330D KI mice showed diminished cardiac fibrosis and improved cardiac function upon isoproterenol exposure. However, the cardiac protection exerted by AMPK was abolished in S330A KI mice. CONCLUSIONS: AMPK phosphorylation of ß-arrestin-1 Ser330 potentiated PDE4 expression and activity, thereby inhibiting ß-AR/cAMP/PKA activation. Subsequently, ß-arrestin-1 Ser330 phosphorylation blocks ß-AR-induced cardiac inflammasome activation and remodeling.

Asunto(s)

Proteínas Quinasas Activadas por AMP , Isoproterenol , Miocitos Cardíacos , beta-Arrestina 1 , Animales , Fosforilación , beta-Arrestina 1/metabolismo , beta-Arrestina 1/genética , Ratones , Proteínas Quinasas Activadas por AMP/metabolismo , Isoproterenol/toxicidad , Isoproterenol/farmacología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Ratones Endogámicos C57BL , Masculino , Receptores Adrenérgicos beta/metabolismo , Serina/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Agonistas Adrenérgicos beta/farmacología , Agonistas Adrenérgicos beta/toxicidad , Células Cultivadas , Transducción de Señal , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Humanos

RESUMEN

Microglia activity can drive excessive synaptic loss during the prodromal phase of Alzheimer's disease (AD) and is associated with lowered cyclic adenosine monophosphate (cAMP) due to cAMP phosphodiesterase 4B (PDE4B). This study aimed to investigate whether long-term inhibition of PDE4B by A33 (3 mg/kg/day) can prevent synapse loss and its associated cognitive decline in APPswe/PS1dE9 mice. This model is characterized by a chimeric mouse/human APP with the Swedish mutation and human PSEN1 lacking exon 9 (dE9), both under the control of the mouse prion protein promoter. The effects on cognitive function of prolonged A33 treatment from 20 days to 4 months of age, was assessed at 7-8 months. PDE4B inhibition significantly improved both the working and spatial memory of APPswe/PSdE9 mice after treatment ended. At the cellular level, in vitro inhibition of PDE4B induced microglial filopodia formation, suggesting that regulation of PDE4B activity can counteract microglia activation. Further research is needed to investigate if this could prevent microglia from adopting their 'disease-associated microglia (DAM)' phenotype in vivo. These findings support the possibility that PDE4B is a potential target in combating AD pathology and that early intervention using A33 may be a promising treatment strategy for AD.

Asunto(s)

Enfermedad de Alzheimer , Cognición , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Modelos Animales de Enfermedad , Ratones Transgénicos , Microglía , Inhibidores de Fosfodiesterasa 4 , Animales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Ratones , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/patología , Cognición/efectos de los fármacos , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/uso terapéutico , Inhibidores de Fosfodiesterasa 4/administración & dosificación , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Presenilina-1/genética , Presenilina-1/metabolismo , Humanos , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Masculino

RESUMEN

Sepsis can quickly result in fatality for critically ill individuals, while liver damage can expedite the progression of sepsis, necessitating the exploration of new strategies for treating hepatic sepsis. PDE4 has been identified as a potential target for the treatment of liver damage. The scaffold hopping of lead compounds FCPR16 and Z19153 led to the discovery of a novel 7-methoxybenzofuran PDE4 inhibitor 4e, demonstrating better PDE4B (IC50 = 10.0 nM) and PDE4D (IC50 = 15.2 nM) inhibitor activity as a potential anti-hepatic sepsis drug in this study. Compared with FCPR16 and Z19153, 4e displayed improved oral bioavailability (F = 66 %) and longer half-life (t1/2 = 2.0 h) in SD rats, which means it can be more easily administered and has a longer-lasting effect. In the D-GalN/LPS-induced liver injury model, 4e exhibited excellent hepatoprotective activity against hepatic sepsis by decreasing ALT and AST levels and inflammatory infiltrating areas.

Asunto(s)

Benzofuranos , Galactosamina , Inhibidores de Fosfodiesterasa 4 , Sepsis , Animales , Humanos , Masculino , Ratas , Benzofuranos/farmacología , Benzofuranos/química , Benzofuranos/síntesis química , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Relación Dosis-Respuesta a Droga , Descubrimiento de Drogas , Galactosamina/farmacología , Lipopolisacáridos/farmacología , Lipopolisacáridos/antagonistas & inhibidores , Hígado/efectos de los fármacos , Hígado/patología , Simulación del Acoplamiento Molecular , Estructura Molecular , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/química , Inhibidores de Fosfodiesterasa 4/síntesis química , Sustancias Protectoras/farmacología , Sustancias Protectoras/química , Sustancias Protectoras/síntesis química , Ratas Sprague-Dawley , Sepsis/tratamiento farmacológico , Relación Estructura-Actividad

RESUMEN

INTRODUCTION: Chronic liver disease (CLD) is a complex disease associated with profound dysfunction. Despite an incredible burden, the first and only pharmacotherapy for metabolic-associated steatohepatitis was only approved in March of this year, indicating a gap in the translation of preclinical studies. There is a body of preclinical work on the application of phosphodiesterase 4 inhibitors in CLD, none of these molecules have been successfully translated into clinical use. AREAS COVERED: To design therapies to combat CLD, it is essential to consider the dysregulation of other tissues that contribute to its development and progression. As such, proper therapies must combat this throughout the body rather than focusing only on the liver. To detail this, literature characterizing the pathogenesis of CLD was pulled from PubMed, with a particular focus placed on the role of PDE4 in inflammation and metabolism. Then, the focus is shifted to detailing the available information on existing PDE4 inhibitors. EXPERT OPINION: This review gives a brief overview of some of the pathologies of organ systems that are distinct from the liver but contribute to disease progression. The demonstrated efficacy of PDE4 inhibitors in other human inflammatory diseases should earn them further examination for the treatment of CLD.

Asunto(s)

Tejido Adiposo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Inhibidores de Fosfodiesterasa 4 , Humanos , Inhibidores de Fosfodiesterasa 4/farmacología , Animales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Tejido Adiposo/metabolismo , Tejido Adiposo/efectos de los fármacos , Hepatopatías/tratamiento farmacológico , Hepatopatías/fisiopatología , Inflamación/tratamiento farmacológico , Inflamación/fisiopatología , Progresión de la Enfermedad , Enfermedad Crónica , Desarrollo de Medicamentos

RESUMEN

Four new p-terphenyl derivatives, talaroterphenyls A-D (1-4), together with three biosynthetically related known ones (5-7), were obtained from the mangrove sediment-derived Talaromyces sp. SCSIO 41412. Compounds 1-3 are rare p-terphenyls, which are completely substituted on the central benzene ring by oxygen atoms; this is the first report of their isolation from natural sources. Their structures were elucidated through NMR spectroscopy, HRESIMS, and X-ray diffraction. Genome sequence analysis revealed that 1-7 were biosynthesized from tyrosine and phenylalanine, involving four key biosynthetic genes (ttpB-ttpE). These p-terphenyls (1-7) and 36 marine-derived terphenyl analogues (8-43) were screened for phosphodiesterase 4 (PDE4) inhibitory activities, and 1-5, 14, 17, 23, and 26 showed notable activities with IC50 values of 0.40-16 µM. The binding pattern of p-terphenyl inhibitors 1-3 with PDE4 were explored by molecular docking analysis. Talaroterphenyl A (1), with a low cytotoxicity, showed obvious anti-inflammatory activity in LPS-stimulated RAW264.7 cells. Furthermore, in the TGF-ß1-induced medical research council cell strain-5 (MRC-5) pulmonary fibrosis model, 1 could down-regulate the expression levels of FN1, COL1, and α-SMA significantly at concentrations of 5-20 µM. This study suggests that the oxidized p-terphenyl 1, as a marine-derived PDE4 inhibitor, could be used as a promising antifibrotic agent.

Asunto(s)

Inhibidores de Fosfodiesterasa 4 , Compuestos de Terfenilo , Inhibidores de Fosfodiesterasa 4/farmacología , Inhibidores de Fosfodiesterasa 4/química , Inhibidores de Fosfodiesterasa 4/aislamiento & purificación , Ratones , Animales , Compuestos de Terfenilo/farmacología , Compuestos de Terfenilo/química , Compuestos de Terfenilo/aislamiento & purificación , Estructura Molecular , Talaromyces/química , Células RAW 264.7 , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Biología Marina

RESUMEN

Cyclic adenosine monophosphate (cAMP) is a key second messenger that regulates signal transduction pathways pivotal for numerous biological functions. Intracellular cAMP levels are spatiotemporally regulated by their hydrolyzing enzymes called phosphodiesterases (PDEs). It has been shown that increased cAMP levels in the central nervous system (CNS) promote neuroplasticity, neurotransmission, neuronal survival, and myelination while suppressing neuroinflammation. Thus, elevating cAMP levels through PDE inhibition provides a therapeutic approach for multiple CNS disorders, including multiple sclerosis, stroke, spinal cord injury, amyotrophic lateral sclerosis, traumatic brain injury, and Alzheimer's disease. In particular, inhibition of the cAMP-specific PDE4 subfamily is widely studied because of its high expression in the CNS. So far, the clinical translation of full PDE4 inhibitors has been hampered because of dose-limiting side effects. Hence, focusing on signaling cascades downstream activated upon PDE4 inhibition presents a promising strategy, offering novel and pharmacologically safe targets for treating CNS disorders. Yet, the underlying downstream signaling pathways activated upon PDE(4) inhibition remain partially elusive. This review provides a comprehensive overview of the existing knowledge regarding downstream mediators of cAMP signaling induced by PDE4 inhibition or cAMP stimulators. Furthermore, we highlight existing gaps and future perspectives that may incentivize additional downstream research concerning PDE(4) inhibition, thereby providing novel therapeutic approaches for CNS disorders.

Asunto(s)

Enfermedades del Sistema Nervioso Central , Sistema Nervioso Central , AMP Cíclico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Inhibidores de Fosfodiesterasa 4 , Transducción de Señal , Humanos , AMP Cíclico/metabolismo , Inhibidores de Fosfodiesterasa 4/farmacología , Animales , Sistema Nervioso Central/efectos de los fármacos , Sistema Nervioso Central/metabolismo , Transducción de Señal/efectos de los fármacos , Enfermedades del Sistema Nervioso Central/tratamiento farmacológico , Enfermedades del Sistema Nervioso Central/metabolismo , Enfermedades del Sistema Nervioso Central/enzimología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo

RESUMEN

We have previously shown that phosphodiesterase 4 (PDE4) inhibition protects against neuronal injury in rats following middle cerebral artery occlusion/reperfusion (MCAO/R). However, the effects of PDE4 on brain edema and astrocyte swelling are unknown. In this study, we showed that inhibition of PDE4 by Roflumilast (Roflu) reduced brain edema and brain water content in rats subjected to MCAO/R. Roflu decreased the expression of aquaporin 4 (AQP4), while the levels of phosphorylated protein kinase B (Akt) and forkhead box O3a (FoxO3a) were increased. In addition, Roflu reduced cell volume and the expression of AQP4 in primary astrocytes undergoing oxygen and glucose deprivation/reoxygenation (OGD/R). Consistently, PDE4B knockdown showed similar effects as PDE4 inhibition; and PDE4B overexpression rescued the inhibitory role of PDE4B knockdown on AQP4 expression. We then found that the effects of Roflu on the expression of AQP4 and cell volume were blocked by the Akt inhibitor MK2206. Since neuroinflammation and astrocyte activation are the common events that are observed in stroke, we treated primary astrocytes with interleukin-1ß (IL-1ß). Astrocytes treated with IL-1ß showed decreased AQP4 and phosphorylated Akt and FoxO3a. Roflu significantly reduced AQP4 expression, which was accompanied by increased phosphorylation of Akt and FoxO3a. Furthermore, overexpression of FoxO3a partly reversed the effect of Roflu on AQP4 expression. Our findings suggest that PDE4 inhibition limits ischemia-induced brain edema and astrocyte swelling via the Akt/FoxO3a/AQP4 pathway. PDE4 is a promising target for the intervention of brain edema after cerebral ischemia.

Asunto(s)

Aminopiridinas , Acuaporina 4 , Astrocitos , Benzamidas , Edema Encefálico , Infarto de la Arteria Cerebral Media , Inhibidores de Fosfodiesterasa 4 , Ratas Sprague-Dawley , Daño por Reperfusión , Animales , Acuaporina 4/metabolismo , Acuaporina 4/genética , Astrocitos/metabolismo , Astrocitos/efectos de los fármacos , Daño por Reperfusión/metabolismo , Inhibidores de Fosfodiesterasa 4/farmacología , Masculino , Edema Encefálico/metabolismo , Edema Encefálico/etiología , Edema Encefálico/patología , Aminopiridinas/farmacología , Benzamidas/farmacología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Ciclopropanos/farmacología , Proteína Forkhead Box O3/metabolismo , Ratas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Células Cultivadas , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Modelos Animales de Enfermedad , Interleucina-1beta/metabolismo

RESUMEN

Objective: Microfold cells (M cells) are specific intestinal epithelial cells for monitoring and transcytosis of antigens, microorganisms, and pathogens in the intestine. However, the mechanism for M-cell development remained elusive. Materials and Methods: Real-time polymerase chain reaction, immunofluorescence, and western blotting were performed to analyze the effect of sorbitol-regulated M-cell differentiation in vivo and in vitro, and luciferase and chromatin Immunoprecipitation were used to reveal the mechanism through which sorbitol-modulated M-cell differentiation. Results: Herein, in comparison to the mannitol group (control group), we found that intestinal M-cell development was inhibited in response to sorbitol treatment as evidenced by impaired enteroids accompanying with decreased early differentiation marker Annexin 5, Marcksl1, Spib, sox8, and mature M-cell marker glycoprotein 2 expression, which was attributed to downregulation of receptor activator of nuclear factor kappa-В ligand (RANKL) expression in vivo and in vitro. Mechanically, in the M-cell model, sorbitol stimulation caused a significant upregulation of phosphodiesterase 4 (PDE4) phosphorylation, leading to decreased protein kinase A (PKA)/cAMP-response element binding protein (CREB) activation, which further resulted in CREB retention in cytosolic and attenuated CREB binds to RANKL promoter to inhibit RANKL expression. Interestingly, endogenous PKA interacted with CREB, and this interaction was destroyed by sorbitol stimulation. Most importantly, inhibition of PDE4 by dipyridamole could rescue the inhibitory effect of sorbitol on intestinal enteroids and M-cell differentiation and mature in vivo and in vitro. Conclusion: These findings suggested that sorbitol suppressed intestinal enteroids and M-cell differentiation and matured through PDE4-mediated RANKL expression; targeting to inhibit PDE4 was sufficient to induce M-cell development.

Asunto(s)

Proteína de Unión a Elemento de Respuesta al AMP Cíclico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Células M , Ligando RANK , Sorbitol , Animales , Masculino , Ratones , Diferenciación Celular/efectos de los fármacos , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Mucosa Intestinal/metabolismo , Células M/efectos de los fármacos , Ratones Endogámicos C57BL , Ligando RANK/metabolismo , Sorbitol/farmacología

RESUMEN

Inhibition of the cyclic-AMP degrading enzyme phosphodiesterase type 4 (PDE4) in the brains of animal models is protective in Alzheimer's disease (AD). We show for the first time that enzymes from the subfamily PDE4D not only colocalize with beta-amyloid (Aß) plaques in a mouse model of AD but that Aß directly associates with the catalytic machinery of the enzyme. Peptide mapping suggests that PDE4D is the preferential PDE4 subfamily for Aß as it possesses a unique binding site. Intriguingly, exogenous addition of Aß to cells overexpressing the PDE4D5 longform caused PDE4 activation and a decrease in cAMP. We suggest a novel mechanism where PDE4 longforms can be activated by Aß, resulting in the attenuation of cAMP signalling to promote loss of cognitive function in AD.

Asunto(s)

Enfermedad de Alzheimer , Péptidos beta-Amiloides , AMP Cíclico , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Neuronas , Animales , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Péptidos beta-Amiloides/metabolismo , AMP Cíclico/metabolismo , Ratones , Neuronas/metabolismo , Humanos , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/genética , Unión Proteica , Activación Enzimática , Ratones Transgénicos , Placa Amiloide/metabolismo , Placa Amiloide/patología

RESUMEN

Widespread metastasis is the primary reason for the high mortality associated with ovarian cancer (OC), and effective targeted therapy for tumor aggressiveness is still insufficient in clinical practice. Therefore, it is urgent to find new targets to improve prognosis of patients. PDE4A is a cyclic nucleotide phosphodiesterase that plays a crucial role in the occurrence and development in various malignancies. Our study firstly reported the function of PDE4A in OC. Expression of PDE4A was validated through bioinformatics analysis, RT-qPCR, Western blot, and immunohistochemistry. Additionally, its impact on cell growth and motility was assessed via in vitro and in vivo experiments. PDE4A was downregulated in OC tissues compared with normal tissues and low PDE4A expression was correlated with poor clinical outcomes in OC patients. The knockdown of PDE4A significantly promoted the proliferation, migration and invasion of OC cells while overexpression of PDE4A resulted in the opposite effect. Furthermore, smaller and fewer tumor metastatic foci were observed in mice bearing PDE4A-overexpressing OVCAR3 cells. Mechanistically, downregulation of PDE4A expression can induce epithelial-mesenchymal transition (EMT) and nuclear translocation of Snail, which suggests that PDE4A plays a pivotal role in suppressing OC progression. Notably, Rolipram, the PDE4 inhibitor, mirrored the effects observed with PDE4A deletion. In summary, the downregulation of PDE4A appears to facilitate OC progression by modulating the Snail/EMT pathway, underscoring the potential of PDE4A as a therapeutic target against ovarian cancer metastasis.

Asunto(s)

Movimiento Celular , Proliferación Celular , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Transición Epitelial-Mesenquimal , Regulación Neoplásica de la Expresión Génica , Neoplasias Ováricas , Factores de Transcripción de la Familia Snail , Humanos , Femenino , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Neoplasias Ováricas/patología , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Animales , Proliferación Celular/genética , Factores de Transcripción de la Familia Snail/metabolismo , Factores de Transcripción de la Familia Snail/genética , Ratones , Movimiento Celular/genética , Transición Epitelial-Mesenquimal/genética , Línea Celular Tumoral , Progresión de la Enfermedad , Ratones Desnudos , Ratones Endogámicos BALB C , Núcleo Celular/metabolismo , Pronóstico

RESUMEN

BACKGROUND: Canine atopic dermatitis (CAD) is a common genetically predisposed, inflammatory, and pruritic skin disorder that affects dogs globally. To date, there are no specific biomarkers available to diagnose CAD, and the current diagnosis is based on a combination of criteria including patient history, clinical signs, and exclusion of other relevant differential diagnoses. METHODS AND RESULTS: We examined the gene expression of phosphodiesterase 4D (PDE4D) in peripheral blood mononuclear cells (PBMCs), as well as miR-203 and miR-483 in plasma, in three groups: healthy dogs, CAD dogs, and other inflammatory pruritic skin diseases (OIPSD) such as pemphigus foliaceus, scabies, cutaneous lymphoma, and dermatophytosis. Our results showed that PDE4D gene expression in the CAD group is statistically higher compared to those in the healthy and OIPSD groups, suggesting PDE4D may be a specific marker for CAD. Nevertheless, no correlation was found between PDE4D gene expression levels and the lesion severity gauged by CAD severity index-4 (CADESI-4). We also showed that miR-203 is a generic marker for clinical dermatitis and differentiates both CAD and OIPSD inflammatory conditions from healthy controls. CONCLUSIONS: We show that PDE4D is a potential marker to differentiate CAD from non-atopic healthy and OIPSD while miR-203 may be a potential marker for general dermatologic inflammation. Future study of PDE4D and miR-203 on a larger scale is warranted.

Asunto(s)

Biomarcadores , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4 , Dermatitis Atópica , Enfermedades de los Perros , MicroARNs , Dermatitis Atópica/genética , Dermatitis Atópica/veterinaria , Dermatitis Atópica/sangre , Dermatitis Atópica/diagnóstico , Animales , Perros , MicroARNs/genética , MicroARNs/sangre , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Biomarcadores/sangre , Enfermedades de los Perros/genética , Enfermedades de los Perros/diagnóstico , Enfermedades de los Perros/sangre , Masculino , Leucocitos Mononucleares/metabolismo , Femenino