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Targeted protein degradation (TPD) using the ubiquitin proteasome system (UPS) is a rapidly growing drug discovery modality to eliminate pathogenic proteins. Strategies for TPD have focused on heterobifunctional degraders that often suffer from poor drug-like properties, and molecular glues that rely on serendipitous discovery. Monovalent "direct" degraders represent an alternative approach, in which small molecules bind to a target protein and induce degradation of that protein through the recruitment of an E3 ligase complex. Using an ultra-high throughput cell-based screening platform, degraders of the bromodomain extraterminal protein BRD4 were identified and optimized to yield a lead compound, PLX-3618. In this paper, we demonstrate that PLX-3618 elicited UPS-mediated selective degradation of BRD4, resulting in potent antitumor activity in vitro and in vivo. Characterization of the degradation mechanism identified DCAF11 as the E3 ligase required for PLX-3618-mediated degradation of BRD4. Protein-protein interaction studies verified a BRD4:PLX-3618:DCAF11 ternary complex, and mutational studies provided further insights into the DCAF11-mediated degradation mechanism. Collectively, these results demonstrate the discovery and characterization of a novel small molecule that selectively degrades BRD4 through the recruitment of the E3 substrate receptor, DCAF11, and promotes potent antitumor activity in vivo.
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Large intestinal intussusception is rare in adults. Among potential pathologic lead points for intussusception are lipomas, benign tumors very infrequently found in the large bowel. A 30-year-old woman presented to the emergency department with a chief complaint of generalized abdominal pain for two weeks. A computed tomography scan of her abdomen and pelvis showed an ileocolic intussusception with a lead point of 6.7 cm. The lead point appeared to be predominantly fat. A colonoscopy revealed a large, obstructing lesion in the transverse colon. The patient underwent exploratory laparotomy with a right hemicolectomy, and a pathologic diagnosis of a lipoma was made. The patient recovered from surgery without complications and returned to her normal diet three weeks after discharge. This case highlights an unusual and rare presentation of an ileocolic intussusception caused by a cecal lipoma acting as a lead point.
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Targeted protein degradation (TPD) using the ubiquitin proteasome system (UPS) is a rapidly growing drug discovery modality to eliminate pathogenic proteins. Strategies for TPD have focused on heterobifunctional degraders that often suffer from poor drug-like properties, and molecular glues that rely on serendipitous discovery. Monovalent "direct" degraders represent an alternative approach, in which small molecules bind to a target protein and induce degradation of that protein through the recruitment of an E3 ligase complex. Using an ultra-high throughput cell-based screening platform, degraders of the bromodomain extra-terminal (BET) protein BRD4 were identified and optimized to yield a lead compound, PLX-3618. In this paper, we demonstrate that PLX-3618 elicited UPS-mediated selective degradation of BRD4, resulting in potent anti-tumor activity in vitro and in vivo. Characterization of the degradation mechanism identified DCAF11 as the E3 ligase required for PLX-3618-mediated degradation of BRD4. Protein-protein interaction studies verified a BRD4:PLX-3618:DCAF11 ternary complex, and mutational studies provided further insights into the DCAF11-mediated degradation mechanism. Collectively, these results demonstrate the discovery and characterization of a novel small molecule that selectively degrades BRD4 through the recruitment of the E3 substrate receptor, DCAF11, and promotes potent anti-tumor activity in vivo.
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PURPOSE: The aim of this systematic review was to synthesise the research identifying possible influences on CBT outcomes in chronic pain. Variations in the effectiveness of psychological therapies, such as CBT, in chronic pain have led to research investigating predictors of improved treatment outcomes. MATERIALS AND METHODS: We identified randomised controlled and cohort studies of CBT for chronic pain, published between 1974 to 2nd August 2023, which identified predictors of CBT outcomes. RESULTS: Nineteen studies were included in the review. Baseline sociodemographic, physical and emotional factors that influence the outcomes of CBT for chronic pain were identified. The most commonly reported predictors of CBT outcome, with medium to large effect sizes, were anxiety, depression and negative cognitions about pain and coping. Sociodemographic predictors of outcomes demonstrated small effects and lacked replicability. CONCLUSIONS: There was variability across study designs, CBT delivery and outcomes measures. Further research is needed in chronic pain to identify the predictive factors which influence treatment outcomes, and consistency across study designs and outcome variables is needed to reduce heterogeneity.
This review synthesised research identifying factors predicting outcomes of Cognitive Behavioural Therapy for chronic pain.The most commonly reported predictors of Cognitive Behavioural Therapy outcome, with medium to large effect sizes, were anxiety, depression, and negative cognitions about pain and coping. sociodemographic predictors of outcomes demonstrated small effects and lacked replicability.There is a move towards more individualised treatments in chronic pain.Our results suggest that decisions regarding Cognitive Behavioural Therapy for chronic pain should carefully consider baseline levels of anxiety, depression, and negative cognitions about pain.
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BACKGROUND & AIMS: Src homology and collagen (Shc) proteins are major adapters to extracellular signals, however, the regulatory role of Shc isoforms in sterile inflammatory responses in alcoholic hepatitis (AH) has not been fully investigated. We hypothesized that in an isoform-specific manner Shc modulates pre-apoptotic signals, calreticulin (CRT) membrane exposure, and recruitment of inflammatory cells. METHODS: Liver biopsy samples from patients with AH vs healthy subjects were studied for Shc expression using DNA microarray data and immunohistochemistry. Shc knockdown (hypomorph) and age-matched wild-type mice were pair-fed according to the chronic-plus-binge alcohol diet. To analyze hepatocyte-specific effects, adeno-associated virus 8-thyroxine binding globulin-Cre (hepatocyte-specific Shc knockout)-mediated deletion was performed in flox/flox Shc mice. Lipid peroxidation, proinflammatory signals, redox radicals, reduced nicotinamide adenine dinucleotide/oxidized nicotinamide adenine dinucleotide ratio, as well as cleaved caspase 8, B-cell-receptor-associated protein 31 (BAP31), Bcl-2-associated X protein (Bax), and Bcl-2 homologous antagonist killer (Bak), were assessed in vivo. CRT translocation was studied in ethanol-exposed p46ShcáºSH2-transfected hepatocytes by membrane biotinylation in conjunction with phosphorylated-eukaryotic initiation factor 2 alpha, BAP31, caspase 8, and Bax/Bak. The effects of idebenone, a novel Shc inhibitor, was studied in alcohol/pair-fed mice. RESULTS: Shc was significantly induced in patients with AH (P < .01). Alanine aminotransferase, reduced nicotinamide adenine dinucleotide/oxidized nicotinamide adenine dinucleotide ratios, production of redox radicals, and lipid peroxidation improved (P < .05), and interleukin 1ß, monocyte chemoattractant protein 1, and C-X-C chemokine ligand 10 were reduced in Shc knockdown and hepatocyte-specific Shc knockout mice. In vivo, Shc-dependent induction, and, in hepatocytes, a p46Shc-dependent increase in pre-apoptotic proteins Bax/Bak, caspase 8, BAP31 cleavage, and membrane translocation of CRT/endoplasmic reticulum-resident protein 57 were seen. Idebenone protected against alcohol-mediated liver injury. CONCLUSIONS: Alcohol induces p46Shc-dependent activation of pre-apoptotic pathways and translocation of CRT to the membrane, where it acts as a damage-associated molecular pattern, instigating immunogenicity. Shc inhibition could be a novel treatment strategy in AH.
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Hepatite Alcoólica , Camundongos , Animais , Proteína X Associada a bcl-2 , Caspase 8 , Calreticulina , NAD , Camundongos Knockout , Etanol , Inflamação , ColágenoRESUMO
Oncoprotein expression is controlled at the level of mRNA translation and is regulated by the eukaryotic translation initiation factor 4F (eIF4F) complex. eIF4A, a component of eIF4F, catalyzes the unwinding of secondary structure in the 5'-untranslated region (5'-UTR) of mRNA to facilitate ribosome scanning and translation initiation. Zotatifin (eFT226) is a selective eIF4A inhibitor that increases the affinity between eIF4A and specific polypurine sequence motifs and has been reported to inhibit translation of driver oncogenes in models of lymphoma. Here we report the identification of zotatifin binding motifs in the 5'-UTRs of HER2 and FGFR1/2 Receptor Tyrosine Kinases (RTKs). Dysregulation of HER2 or FGFR1/2 in human cancers leads to activation of the PI3K/AKT and RAS/ERK signaling pathways, thus enhancing eIF4A activity and promoting the translation of select oncogenes that are required for tumor cell growth and survival. In solid tumor models driven by alterations in HER2 or FGFR1/2, downregulation of oncoprotein expression by zotatifin induces sustained pathway-dependent anti-tumor activity resulting in potent inhibition of cell proliferation, induction of apoptosis, and significant in vivo tumor growth inhibition or regression. Sensitivity of RTK-driven tumor models to zotatifin correlated with high basal levels of mTOR activity and elevated translational capacity highlighting the unique circuitry generated by the RTK-driven signaling pathway. This dependency identifies the potential for rational combination strategies aimed at vertical inhibition of the PI3K/AKT/eIF4F pathway. Combination of zotatifin with PI3K or AKT inhibitors was beneficial across RTK-driven cancer models by blocking RTK-driven resistance mechanisms demonstrating the clinical potential of these combination strategies.
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Alzheimer's disease(AD) is an age-associated neurodegenerative disease that results in deterioration of memory and cognitive function. As a currently untreatable disorder, AD has emerged as one of the defining biomedical challenges of our time. Thus, new approaches that can examine the cellular and molecular mechanisms underlying age-related AD pathology are sorely needed. One of the hallmarks of Alzheimer's disease is the hyperphosphorylation of the tau protein. Caenorhabditis elegans have been previously used to study the genetic pathways impacted by tau proteotoxic stress; however, currently, available C. elegans tau models express the human protein solely in neurons, which are unresponsive to global RNA interference (RNAi). This limits powerful RNAi screening methods from being utilized effectively in these disease models. Our goal was to develop a C. elegans tau model that has pronounced tau-induced disease phenotypes in cells that can be modified by feeding RNAi methods. Towards this end, we generated a novel C. elegans transgenic line with codon-optimized human 0N4R V337M tau expressed in the body wall muscle under the myo-3 promoter. Immunoblotting experiments revealed that the expressed tau is phosphorylated on epitopes canonically associated with human AD pathology. The tau line has significantly reduced health metrics, including egg laying, growth rate, paralysis, thrashing frequency, crawling speed, and lifespan. These defects are suppressed by RNAi directed against the tau mRNA. Taken together, our results suggest that this alternative tau genetic model could be a useful tool for uncovering the mechanisms that influence the hyperphosphorylation and toxicity of human tau via RNAi screening and other approaches.
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Proteínas de Caenorhabditis elegans , Doenças Neurodegenerativas , Tauopatias , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Modelos Animais de Doenças , Humanos , Tauopatias/genéticaRESUMO
Flavaglines such as silvestrol (1) and rocaglamide (2) constitute an interesting class of natural products with promising anticancer activities. Their mode of action is based on inhibition of eukaryotic initiation factor 4A (eIF4A) dependent translation through formation of a stable ternary complex with eIF4A and mRNA, thus blocking ribosome scanning. Herein we describe initial SAR studies in a novel series of 1-aminomethyl substituted flavagline-inspired eIF4A inhibitors. We discovered that a variety of N-substitutions at the 1-aminomethyl group are tolerated, making this position pertinent for property and ADME profile tuning. The findings presented herein are relevant to future drug design efforts towards novel eIF4A inhibitors with drug-like properties.
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Antineoplásicos/farmacologia , Benzofuranos/farmacologia , Produtos Biológicos/farmacologia , Fator de Iniciação 4A em Eucariotos/antagonistas & inibidores , Triterpenos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Benzofuranos/síntese química , Benzofuranos/química , Produtos Biológicos/síntese química , Produtos Biológicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Fator de Iniciação 4A em Eucariotos/metabolismo , Humanos , Estrutura Molecular , Relação Estrutura-Atividade , Triterpenos/síntese química , Triterpenos/químicaRESUMO
The PI3K/AKT/mTOR pathway is often activated in lymphoma through alterations in PI3K, PTEN, and B-cell receptor signaling, leading to dysregulation of eIF4A (through its regulators, eIF4B, eIF4G, and PDCD4) and the eIF4F complex. Activation of eIF4F has a direct role in tumorigenesis due to increased synthesis of oncogenes that are dependent on enhanced eIF4A RNA helicase activity for translation. eFT226, which inhibits translation of specific mRNAs by promoting eIF4A1 binding to 5'-untranslated regions (UTR) containing polypurine and/or G-quadruplex recognition motifs, shows potent antiproliferative activity and significant in vivo efficacy against a panel of diffuse large B-cell lymphoma (DLBCL), and Burkitt lymphoma models with ≤1 mg/kg/week intravenous administration. Evaluation of predictive markers of sensitivity or resistance has shown that activation of eIF4A, mediated by mTOR signaling, correlated with eFT226 sensitivity in in vivo xenograft models. Mutation of PTEN is associated with reduced apoptosis in vitro and diminished efficacy in vivo in response to eFT226. In models evaluated with PTEN loss, AKT was stimulated without a corresponding increase in mTOR activation. AKT activation leads to the degradation of PDCD4, which can alter eIF4F complex formation. The association of eFT226 activity with PTEN/PI3K/mTOR pathway regulation of mRNA translation provides a means to identify patient subsets during clinical development.
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Fator de Iniciação 4A em Eucariotos/antagonistas & inibidores , Linfoma de Células B/genética , Linfoma de Células B/patologia , Oncogenes , Biossíntese de Proteínas/genética , RNA Mensageiro/genética , Animais , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Fator de Iniciação 4A em Eucariotos/metabolismo , Feminino , Humanos , Camundongos Endogâmicos NOD , Camundongos SCID , PTEN Fosfo-Hidrolase/metabolismo , RNA Mensageiro/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
INTRODUCTION: Despite its popularity, there have been no reports outlining adult craniofacial trauma in the setting of golf injuries. Our main objective was to identify and describe trends in head and neck injuries incurred while participating in golf including injury type, anatomic location, and patient disposition. METHODS: A retrospective cohort analysis of the National Electronic Injury Surveillance System (NEISS) was conducted from 2009-2018. Population characteristics of golf-related injuries were recorded, including, age, sex, and race distributions. Distribution of injury anatomic location, injury type, distribution of fracture location, patient disposition was further analyzed. A one way ANOVA was utilized to obtain the mean ages for all injuries and compare them for any statistical difference. To identify statistical significance, a Fisher exact test with a Monte Carlo simulation was performed. RESULTS: A total of 509 golf-related injuries (national estimate 24,425 cases) were recorded over the study period. Lacerations were the most common injury overall (54.2%), while contusions and abrasions were the next most common injuries (27.3%). The most common fracture subtype observed was midface (40.43%), followed by nasal bone (27.66%), mandible (12.77%), skull (12.77%), and cervical spine (6.38%). The highest proportion of patients admitted for further treatment were individuals >70 years of age. Fractures had the highest rate of admission (29.8%). CONCLUSIONS: A better understanding of golf injuries can allow for rapid detection and appropriate treatment when encountered. This knowledge can also help to develop safety precautions by potentially reforming rules and regulations as well as protective equipment.
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Contusões , Traumatismos Craniocerebrais , Golfe , Adulto , Traumatismos Craniocerebrais/epidemiologia , Traumatismos Craniocerebrais/etiologia , Bases de Dados Factuais , Humanos , Estudos RetrospectivosRESUMO
Rocaglates, rocaglamides, and related flavagline natural products exert their remarkable anticancer activity through inhibition of eukaryotic initiation factor 4A (eIF4A) but generally display suboptimal drug-like properties. In our efforts to identify potent drug-like eIF4A inhibitors, we developed synthetic strategies for diastereoselectively functionalizing the C1 position of aza-rocaglamide scaffolds (cf. 14 and 18), which proceed via retention or inversion of configuration at C1 depending on the C2 substituent (cf. 15 and 21) and ultimately enabled the discovery of novel and potent eIF4A inhibitors such as 25.
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Benzofuranos/química , Fator de Iniciação 4A em Eucariotos/antagonistas & inibidores , Sítios de Ligação , Produtos Biológicos , Fator de Iniciação 4A em Eucariotos/metabolismo , Humanos , Estrutura MolecularRESUMO
Type 2 diabetes is clinically associated with progressive necroinflammation and fibrosis in nonalcoholic steatohepatitis (NASH). Advanced glycation end-products (AGEs) accumulate during prolonged hyperglycemia, but the mechanistic pathways that lead to accelerated liver fibrosis have not been well defined. In this study, we show that the AGEs clearance receptor AGER1 was downregulated in patients with NASH and diabetes and in our NASH models, whereas the proinflammatory receptor RAGE was induced. These findings were associated with necroinflammatory, fibrogenic, and pro-oxidant activity via the NADPH oxidase 4. Inhibition of AGEs or RAGE deletion in hepatocytes in vivo reversed these effects. We demonstrate that dysregulation of NRF2 by neddylation of cullin 3 was linked to AGER1 downregulation and that induction of NRF2 using an adeno-associated virus-mediated approach in hepatocytes in vivo reversed AGER1 downregulation, lowered the level of AGEs, and improved proinflammatory and fibrogenic responses in mice on a high AGEs diet. In patients with NASH and diabetes or insulin resistance, low AGER1 levels were associated with hepatocyte ballooning degeneration and ductular reaction. Collectively, prolonged exposure to AGEs in the liver promotes an AGER1/RAGE imbalance and consequent redox, inflammatory, and fibrogenic activity in NASH.
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Diabetes Mellitus Tipo 2/metabolismo , Regulação para Baixo , Cirrose Hepática/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Receptor para Produtos Finais de Glicação Avançada/biossíntese , Animais , Ácido Ascórbico , Colecalciferol , Desidroepiandrosterona/análogos & derivados , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Hepatócitos/metabolismo , Hepatócitos/patologia , Cirrose Hepática/genética , Cirrose Hepática/patologia , Camundongos , Camundongos Knockout , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Ácidos Nicotínicos , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/patologia , Extratos Vegetais , Receptor para Produtos Finais de Glicação Avançada/genéticaRESUMO
Dysregulation of protein translation is a key driver for the pathogenesis of many cancers. Eukaryotic initiation factor 4A (eIF4A), an ATP-dependent DEAD-box RNA helicase, is a critical component of the eIF4F complex, which regulates cap-dependent protein synthesis. The flavagline class of natural products (i.e., rocaglamide A) has been shown to inhibit protein synthesis by stabilizing a translation-incompetent complex for select messenger RNAs (mRNAs) with eIF4A. Despite showing promising anticancer phenotypes, the development of flavagline derivatives as therapeutic agents has been hampered because of poor drug-like properties as well as synthetic complexity. A focused effort was undertaken utilizing a ligand-based design strategy to identify a chemotype with optimized physicochemical properties. Also, detailed mechanistic studies were undertaken to further elucidate mRNA sequence selectivity, key regulated target genes, and the associated antitumor phenotype. This work led to the design of eFT226 (Zotatifin), a compound with excellent physicochemical properties and significant antitumor activity that supports clinical development.
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Benzofuranos/química , Desenho de Fármacos , Fator de Iniciação 4A em Eucariotos/antagonistas & inibidores , Animais , Benzofuranos/farmacocinética , Benzofuranos/uso terapêutico , Sítios de Ligação , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Cristalografia por Raios X , Fator de Iniciação 4A em Eucariotos/genética , Fator de Iniciação 4A em Eucariotos/metabolismo , Feminino , Meia-Vida , Humanos , Ligantes , Camundongos , Camundongos Nus , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Estrutura Terciária de Proteína , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Ratos , Relação Estrutura-AtividadeRESUMO
BACKGROUND AND AIMS: Older patients with obesity/type II diabetes mellitus frequently present with advanced NASH. Whether this is due to specific molecular pathways that accelerate fibrosis during aging is unknown. Activation of the Src homology 2 domain-containing collagen-related (Shc) proteins and redox stress have been recognized in aging; however, their link to NASH has not been explored. APPROACH AND RESULTS: Shc expression increased in livers of older patients with NASH, as assessed by real time quantitative PCR (RT-qPCR) or western blots. Fibrosis, Shc expression, markers of senescence, and nicotinamide adenine dinucleotide phosphate, reduced form oxidases (NOXs) were studied in young/old mice on fast food diet (FFD). To inhibit Shc in old mice, lentiviral (LV)-short hairpin Shc versus control-LV were used during FFD. For hepatocyte-specific effects, floxed (fl/fl) Shc mice on FFD were injected with adeno-associated virus 8-thyroxine-binding globulin-Cre-recombinase versus control. Fibrosis was accelerated in older mice on FFD, and Shc inhibition by LV in older mice or hepatocyte-specific deletion resulted in significantly improved inflammation, reduction in senescence markers in older mice, lipid peroxidation, and fibrosis. To study NOX2 activation, the interaction of p47phox (NOX2 regulatory subunit) and p52Shc was evaluated by proximity ligation and coimmunoprecipitations. Palmitate-induced p52Shc binding to p47phox , activating the NOX2 complex, more so at an older age. Kinetics of binding were assessed in Src homology 2 domain (SH2) or phosphotyrosine-binding (PTB) domain deletion mutants by biolayer interferometry, revealing the role of SH2 and the PTB domains. Lastly, an in silico model of p52Shc/p47phox interaction using RosettaDock was generated. CONCLUSIONS: Accelerated fibrosis in the aged is modulated by p52Shc/NOX2. We show a pathway for direct activation of the phagocytic NOX2 in hepatocytes by p52Shc binding and activating the p47phox subunit that results in redox stress and accelerated fibrosis in the aged.
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Envelhecimento/metabolismo , NADPH Oxidase 2/fisiologia , Hepatopatia Gordurosa não Alcoólica/etiologia , Animais , Hepatócitos/metabolismo , Humanos , Cirrose Hepática/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NADPH Oxidases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Adaptadoras da Sinalização Shc/antagonistas & inibidores , Proteínas Adaptadoras da Sinalização Shc/fisiologia , Domínios de Homologia de srcRESUMO
Recruitment of inflammatory cells is a major feature of alcoholic liver injury however; the signals and cellular sources regulating this are not well defined. C-C chemokine receptor type 2 (CCR2) is expressed by active hepatic stellate cells (HSC) and is a key monocyte recruitment signal. Activated HSC are also important sources of hydrogen peroxide resulting from the activation of NADPH oxidase 4 (NOX4). As the role of this NOX in early alcoholic liver injury has not been addressed, we studied NOX4-mediated regulation of CCR2/CCL2 mRNA stability. NOX4 mRNA was significantly induced in patients with alcoholic liver injury, and was co-localized with αSMA-expressing activated HSC. We generated HSC-specific NOX4 KO mice and these were pair-fed on alcohol diet. Lipid peroxidation have not changed significantly however, the expression of CCR2, CCL2, Ly6C, TNFα, and IL-6 was significantly reduced in NOX4HSCKO compared to fl/fl mice. NOX4 promoter was induced in HSC by acetaldehyde treatment, and NOX4 has significantly increased mRNA half-life of CCR2 and CCL2 in conjunction with Ser221 phosphorylation and cytoplasmic shuttling of HuR. In conclusion, NOX4 is induced in early alcoholic liver injury and regulates CCR2/CCL2 mRNA stability thereby promoting recruitment of inflammatory cells and production of proinflammatory cytokines.
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Quimiocina CCL2/genética , Hepatopatias Alcoólicas/genética , NADPH Oxidase 4/metabolismo , Estabilidade de RNA/genética , Receptores CCR2/genética , Acetaldeído/farmacologia , Consumo de Bebidas Alcoólicas/genética , Animais , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Células Cultivadas , Quimiocina CCL2/metabolismo , Proteína Semelhante a ELAV 1/metabolismo , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Hepatopatias Alcoólicas/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transporte Proteico/efeitos dos fármacos , Estabilidade de RNA/efeitos dos fármacos , Receptores CCR2/metabolismoRESUMO
Macrophage activation is an important feature of primary biliary cholangitis (PBC) pathogenesis and other cholestatic liver diseases. Galectin-3 (Gal3), a pleiotropic lectin, is produced by monocytic cells and macrophages. However, its role in PBC has not been addressed. We hypothesized that Gal3 is a key to induce NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome in macrophages and in turn to propagate proinflammatory IL-17 signaling. In liver tissues from patients with PBC and dnTGF-ßRII mice, a model of autoimmune cholangitis, the expression of Gal3, NLRP3, and the adaptor protein adaptor apoptosis-associated speck-like protein was induced, with the downstream activation of caspase-1 and IL-1ß. In wild-type hepatic macrophages, deoxycholic acid induced the association of Gal3 and NLRP3 with direct activation of the inflammasome, resulting in an increase in IL-1ß. Downstream retinoid-related orphan receptor C mRNA, IL-17A, and IL-17F were induced. In Gal3-/- macrophages, no inflammasome activation was detected. To confirm the key role of Gal3 in the pathogenesis of cholestatic liver injury, we generated dnTGF-ßRII/galectin-3-/- (dn/Gal3-/-) mice, which showed impaired inflammasome activation along with significantly improved inflammation and fibrosis. Taken together, our data point to a novel role of Gal3 as an initiator of inflammatory signaling in autoimmune cholangitis, mediating the activation of NLRP3 inflammasome and inducing IL-17 proinflammatory cascades. These studies provide a rationale to target Gal3 in autoimmune cholangitis and potentially other cholestatic diseases.-Tian, J., Yang, G., Chen, H.-Y., Hsu, D. K., Tomilov, A., Olson, K. A., Dehnad, A., Fish, S. R., Cortopassi, G., Zhao, B., Liu, F.-T., Gershwin, M. E., Török, N. J., Jiang, J. X. Galectin-3 regulates inflammasome activation in cholestatic liver injury.
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Galectina 3/metabolismo , Inflamassomos/metabolismo , Fígado/metabolismo , Macrófagos/metabolismo , Transdução de Sinais/fisiologia , Animais , Caspase 1/metabolismo , Células Cultivadas , Galectina 3/genética , Humanos , Interleucina-17/metabolismo , Interleucina-23/metabolismo , Fígado/lesões , Ativação de Macrófagos/fisiologia , Camundongos Transgênicos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismoRESUMO
MicroRNAs (miRNAs), small RNA molecules that post-transcriptionally regulate mRNA expression, are crucial in diverse developmental and physiological programs and their misregulation can lead to disease. Chemically modified oligonucleotides have been developed to modulate miRNA activity for therapeutic intervention in disease settings, but their mechanism of action has not been fully elucidated. Here we show that the miRNA inhibitors (anti-miRs) physically associate with Argonaute proteins in the context of the cognate target miRNA in vitro and in vivo. The association is mediated by the seed region of the miRNA and is sensitive to the placement of chemical modifications. Furthermore, the targeted miRNAs are stable and continue to be associated with Argonaute. Our results suggest that anti-miRs specifically associate with Argonaute-bound miRNAs, preventing association with target mRNAs, which leads to subsequent stabilization and thus increased expression of the targeted mRNAs.
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Proteínas Argonautas/metabolismo , MicroRNAs/antagonistas & inibidores , MicroRNAs/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Animais , Feminino , Humanos , Masculino , Camundongos Endogâmicos C57BL , Ligação Proteica/efeitos dos fármacosRESUMO
Aminoglycoside antibiotics target an internal RNA loop within the bacterial ribosomal decoding site. Here, we describe the synthesis and SAR of novel 3,5-diamino-piperidine derivatives as aminoglycoside mimetics, and show they act as inhibitors of bacterial translation and growth.
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Antibacterianos/síntese química , Piperidinas/síntese química , Piperidinas/farmacologia , Aminoglicosídeos , Antibacterianos/farmacologia , Mimetismo Molecular , Biossíntese de Proteínas/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/crescimento & desenvolvimento , Relação Estrutura-AtividadeRESUMO
We report the structure-guided discovery, synthesis, and initial characterization of 3,5-diamino-piperidinyl triazines (DAPT), a novel translation inhibitor class that targets bacterial rRNA and exhibits broad-spectrum antibacterial activity. DAPT compounds were designed as structural mimetics of aminoglycoside antibiotics which bind to the bacterial ribosomal decoding site and thereby interfere with translational fidelity. We found that DAPT compounds bind to oligonucleotide models of decoding-site RNA, inhibit translation in vitro, and induce translation misincorporation in vivo, in agreement with a mechanism of action at the ribosomal decoding site. The novel DAPT antibacterials inhibit growth of gram-positive and gram-negative bacteria, including the respiratory pathogen Pseudomonas aeruginosa, and display low toxicity to human cell lines. In a mouse protection model, an advanced DAPT compound demonstrated efficacy against an Escherichia coli infection at a 50% protective dose of 2.4 mg/kg of body weight by single-dose intravenous administration.