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DEAD-box helicase 17 (DDX17) is a typical member of the DEAD-box family with transcriptional cofactor activity. Although DDX17 is abundantly expressed in the myocardium, its role in heart is not fully understood. We generated cardiomyocyte-specific Ddx17-knockout mice (Ddx17-cKO), cardiomyocyte-specific Ddx17 transgenic mice (Ddx17-Tg), and various models of cardiomyocyte injury and heart failure (HF). DDX17 is downregulated in the myocardium of mouse models of heart failure and cardiomyocyte injury. Cardiomyocyte-specific knockout of Ddx17 promotes autophagic flux blockage and cardiomyocyte apoptosis, leading to progressive cardiac dysfunction, maladaptive remodeling and progression to heart failure. Restoration of DDX17 expression in cardiomyocytes protects cardiac function under pathological conditions. Further studies showed that DDX17 can bind to the transcriptional repressor B-cell lymphoma 6 (BCL6) and inhibit the expression of dynamin-related protein 1 (DRP1). When DDX17 expression is reduced, transcriptional repression of BCL6 is attenuated, leading to increased DRP1 expression and mitochondrial fission, which in turn leads to impaired mitochondrial homeostasis and heart failure. We also investigated the correlation of DDX17 expression with cardiac function and DRP1 expression in myocardial biopsy samples from patients with heart failure. These findings suggest that DDX17 protects cardiac function by promoting mitochondrial homeostasis through the BCL6-DRP1 pathway in heart failure.
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RNA Helicases DEAD-box , Insuficiência Cardíaca , Miócitos Cardíacos , Animais , Humanos , Camundongos , Apoptose/genética , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Dinaminas/genética , Dinaminas/metabolismo , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/patologia , Insuficiência Cardíaca/metabolismo , Homeostase/genética , Camundongos Knockout , Camundongos Transgênicos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Dinâmica Mitocondrial/genética , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Proteínas Proto-Oncogênicas c-bcl-6/genética , Proteínas Proto-Oncogênicas c-bcl-6/metabolismoRESUMO
[This corrects the article DOI: 10.3389/fonc.2022.943032.].
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OBJECTIVES: To investigate the function and regulatory mechanisms of delphinidin in the treatment of hepatocellular carcinoma. METHODS: HepG2 and HuH-7 cells were treated with different concentrations of delphinidin. Cell viability was analysed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The cell autophagy and autophagic flux were analysed by LC3b-green fluorescent protein (GFP)-Adv and LC3b-GFP-monomeric red fluorescent protein-Adv transfected HepG2 and HuH-7 cells, respectively. Cell apoptosis was analysed by Hoechst33342 staining, terminal deoxynucleotidyl transferase dUTP nick end labeling staining and DNA laddering. Cell autophagy, apoptosis and survival related protein expressions were detected by Western blotting. KEY FINDINGS: After treatment with different concentrations of delphinidin, the cell survival rate was significantly decreased. Delphinidin could block the autophagic flux, resulting in a significant increase in autophagosomes, and led to an increase in cell apoptosis. The combined application of delphinidin and cisplatin could promote the antitumour effect and reduce the dose of cisplatin in tumour cells. Further mechanism studies reveal that delphinidin could inhibit the multidrug resistance gene 1 (MDR1) and the tumour-promoting transcription cofactor DEAD-box helicase 17 (DDX17) expression in tumour cells. Overexpression of DDX17 could reverse delphinidin's antitumor function in tumour cells. CONCLUSIONS: Delphinidin has a strong anti-tumour effect by inducing tumour cell autophagic flux blockage and apoptosis by inhibiting of both MDR1 and DDX17 expression.
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Cisplatino , Neoplasias Hepáticas , Humanos , Cisplatino/farmacologia , Genes MDR , Apoptose , Autofagia , Linhagem Celular Tumoral , RNA Helicases DEAD-box/farmacologiaRESUMO
Aging is a gradual and irreversible pathophysiological process. It presents with declines in tissue and cell functions and significant increases in the risks of various aging-related diseases, including neurodegenerative diseases, cardiovascular diseases, metabolic diseases, musculoskeletal diseases, and immune system diseases. Although the development of modern medicine has promoted human health and greatly extended life expectancy, with the aging of society, a variety of chronic diseases have gradually become the most important causes of disability and death in elderly individuals. Current research on aging focuses on elucidating how various endogenous and exogenous stresses (such as genomic instability, telomere dysfunction, epigenetic alterations, loss of proteostasis, compromise of autophagy, mitochondrial dysfunction, cellular senescence, stem cell exhaustion, altered intercellular communication, deregulated nutrient sensing) participate in the regulation of aging. Furthermore, thorough research on the pathogenesis of aging to identify interventions that promote health and longevity (such as caloric restriction, microbiota transplantation, and nutritional intervention) and clinical treatment methods for aging-related diseases (depletion of senescent cells, stem cell therapy, antioxidative and anti-inflammatory treatments, and hormone replacement therapy) could decrease the incidence and development of aging-related diseases and in turn promote healthy aging and longevity.
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Promoção da Saúde , Doenças Neurodegenerativas , Humanos , Idoso , Envelhecimento/metabolismo , Senescência Celular/genética , Instabilidade Genômica , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/terapiaRESUMO
DEAD-box (DDX)5 and DDX17, which belong to the DEAD-box RNA helicase family, are nuclear and cytoplasmic shuttle proteins. These proteins are expressed in most tissues and cells and participate in the regulation of normal physiological functions; their abnormal expression is closely related to tumorigenesis and tumor progression. DDX5/DDX17 participate in almost all processes of RNA metabolism, such as the alternative splicing of mRNA, biogenesis of microRNAs (miRNAs) and ribosomes, degradation of mRNA, interaction with long noncoding RNAs (lncRNAs) and coregulation of transcriptional activity. Moreover, different posttranslational modifications, such as phosphorylation, acetylation, ubiquitination, and sumoylation, endow DDX5/DDX17 with different functions in tumorigenesis and tumor progression. Indeed, DDX5 and DDX17 also interact with multiple key tumor-promoting molecules and participate in tumorigenesis and tumor progression signaling pathways. When DDX5/DDX17 expression or their posttranslational modification is dysregulated, the normal cellular signaling network collapses, leading to many pathological states, including tumorigenesis and tumor development. This review mainly discusses the molecular structure features and biological functions of DDX5/DDX17 and their effects on tumorigenesis and tumor progression, as well as their potential clinical application for tumor treatment.
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Hepatic lipid accumulation is an initiation factor in fatty liver disease, and promoting a reduction in hepatic lipid accumulation is an important treatment strategy. DEAD box RNA helicase 17 (DDX17) is a member of the DEAD-box family and a molecular chaperone. Previous studies have demonstrated that DDX17 is a transcriptional coregulator of tumorigenesis, inflammation, and macrophage cholesterol efflux. The liver is the main site for lipid metabolism, and metabolic (dysfunction)-associated fatty liver disease (MAFLD) is one of the most common chronic liver diseases. However, the impact of DDX17 on hepatic lipid accumulation has not been verified. In this study, we found, for the first time, that oleic acid/palmitic acid (OA/PA)-induced lipid accumulation was largely abrogated by DDX17 overexpression in both HepG2 (a human hepatocellular carcinoma line) and Hep1-6 (a murine hepatocellular carcinoma line) cells, and this effect was due to a marked reduction in cellular triglyceride (TG) content. Moreover, the overexpression of DDX17 was accompanied by a significant decrease in the expression of genes involved in de novo fatty acid synthesis (FAS, ACC, and SCD-1) in both HepG2 and Hep1-6 cells. In conclusion, DDX17 protected against OA/PA-induced lipid accumulation in hepatocytes through de novo lipogenesis inhibition.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Hepatopatia Gordurosa não Alcoólica , Animais , Carcinoma Hepatocelular/metabolismo , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Células Hep G2 , Hepatócitos/metabolismo , Humanos , Metabolismo dos Lipídeos , Lipogênese , Fígado/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Hepatopatia Gordurosa não Alcoólica/metabolismo , Ácido Oleico/metabolismo , Ácido Oleico/farmacologia , Ácido Palmítico/metabolismo , Ácido Palmítico/farmacologiaRESUMO
OBJECTIVE: To investigate the protective effects and regulatory mechanism of miR-488-3p on doxorubicin-induced cardiotoxicity. METHODS: The C57BL/6 mice and primary cardiomyocytes were used to construct doxorubicin-induced cardiomyocyte injury models in vivo and in vitro. The levels of miR-488-3p and its downstream target genes were analyzed by quantitative real-time PCR. Mouse cardiac function, cell survival, cellular injury-related proteins, and the apoptosis level of cardiomyocytes were analyzed by echocardiography, MTT analysis, Western blotting, and DNA laddering separately. RESULTS: Cardiomyocyte injury caused by a variety of stimuli can lead to the reduction of miR-488-3p level, especially when stimulated with doxorubicin. Doxorubicin led to significant decrease in cardiac function, cell autophagic flux blockage, and apoptosis in vivo and in vitro. The expression of miR-488-3p's target gene, CyclinG1, increased remarkably in the doxorubicin-treated neonatal mouse cardiomyocytes. Overexpression of miR-488-3p inhibited CyclinG1 expression, increased cardiomyocyte viability, and attenuated doxorubicin-induced cardiomyocyte autophagic flux blockage and apoptosis. CONCLUSIONS: miR-488-3p is one of the important protective miRNAs in doxorubicin-induced cardiotoxicity by inhibiting the expression of CyclinG1, which provides insight into the possible clinical application of miR-488-3p/CyclinG1 as therapeutic targets in doxorubicin-induced cardiovascular diseases.
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Antibióticos Antineoplásicos/efeitos adversos , Cardiotoxicidade/etiologia , Ciclina G1/antagonistas & inibidores , Doxorrubicina/efeitos adversos , MicroRNAs/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Animais , Humanos , Masculino , Camundongos , RatosRESUMO
Lysophosphatidylcholine (LPC), the active metabolite of palmitate, triggers hepatocyte death by activating endoplasmic reticulum stress and JNK signalling-mediated lipoapoptosis. However, LPC-induced cytotoxicity in hepatocytes is not well understood. Here, we found for the first time that LPC-induced cell rounding occurred prior to apoptosis. LPC-induced rounding of cells reduced both cell-extracellular matrix (ECM) adhesion and cell-cell junctions, which promoted detachment-induced apoptosis (defined as anoikis) in hepatocytes. Further study revealed that LPC altered cellular morphology and cell adhesion by inhibiting integrin and cadherin signalling-mediated microfilament polymerization. We also found that ECM supplementation and microfilament cytoskeletal stabilization inhibited LPC-induced hepatocyte death by attenuating anoikis. Our data indicate a novel cytotoxic process and signalling pathway induced by LPC.
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Anoikis/efeitos dos fármacos , Caderinas/genética , Adesão Celular/efeitos dos fármacos , Integrinas/genética , Junções Intercelulares/efeitos dos fármacos , Lisofosfatidilcolinas/farmacologia , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestrutura , Anoikis/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Caderinas/metabolismo , Caspase 8/genética , Caspase 8/metabolismo , Linhagem Celular Tumoral , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/genética , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Matriz Extracelular/ultraestrutura , Regulação da Expressão Gênica , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/ultraestrutura , Humanos , Integrinas/metabolismo , Junções Intercelulares/metabolismo , Junções Intercelulares/ultraestrutura , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais , Vinculina/genética , Vinculina/metabolismoRESUMO
OBJECTIVE: In this study, we aim to explore the role of bone marrow macrophage-derived exosomes in hepatic insulin resistance, investigate the substance in exosomes that regulates hepatic insulin signalling pathways, reveal the specific molecular mechanisms involved in hepatic insulin resistance and further explore the role of exosomes in type 2 diabetes. MATERIALS AND METHODS: High-fat diet (HFD)-fed mice were used as obesity-induced hepatic insulin resistance model, exosomes were isolated from BMMs which were extracted from HFD-fed mice by ultracentrifugation. Exosomes were analysed the spectral changes of microRNA expression using a microRNA array. The activation of the insulin signalling pathway and the level of glycogenesis were examined in hepatocytes after transfected with miR-143-5p mimics. Luciferase assay and western blot were used to assess the target of miR-143-5p. RESULTS: BMMs from HFD-fed mice were polarized towards M1, and miR-143-5p was significantly upregulated in exosomes of BMMs from HFD-fed mice. Overexpression of miR-143-5p in Hep1-6 cells led to decreased phosphorylation of AKT and GSK and glycogen synthesis. Dual-luciferase reporter assay and western blot demonstrated that mitogen-activated protein kinase phosphatase-5 (Mkp5, also known as Dusp10) was the target gene of miR-143-5p. Moreover, the overexpression of MKP5 could rescue the insulin resistance induced by transfection miR-143-5p mimics in Hep1-6. CONCLUSION: Bone marrow macrophage-derived exosomal miR-143-5p induces insulin resistance in hepatocytes through repressing MKP5.
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Células da Medula Óssea/metabolismo , Fosfatases de Especificidade Dupla/biossíntese , Regulação Enzimológica da Expressão Gênica , Hepatócitos/metabolismo , Resistência à Insulina , Macrófagos/metabolismo , MicroRNAs/metabolismo , Animais , Dieta Hiperlipídica , Exossomos , CamundongosRESUMO
Lipotoxicity-induced apoptosis, also referred to as lipoapoptosis, is one of the important initial factors promoting the progression from hepatosteatosis to nonalcoholic steatohepatitis (NASH). Saturated free fatty acids (SFAs), which are increased significantly in NASH, are directly hepatotoxic which induce hepatocyte lipoapoptosis. Previously, we reported that protein phosphatase 4 (PP4) was a novel regulator of hepatic insulin resistance and lipid metabolism, but its role in hepatic lipoapoptosis remains unexplored. In this study, we found out that PP4 was upregulated in the livers of western diet-fed-induced NASH mice and SFA-treated murine primary hepatocytes and HepG2 cells. In addition, we found for the first time that suppression of PP4 decreased SFA-induced JNK activation and expression of key modulators of hepatocyte lipoapoptosis including p53-upregulated modulator of apoptosis (PUMA) and Bcl-2-interacting mediator (Bim) and reduced hepatocyte lipoapoptosis level as well both in vitro and in vivo. Further study revealed that PP4 induced JNK activation and lipoapoptosis-related protein expression by regulating the RAC1/MLK3 pathway instead of the PERK/CHOP pathway. The effects of palmitate-treated and PP4-induced lipoapoptosis pathway activation were largely abolished by RAC1 inhibition. Moreover, we identified that PP4 interacted with RAC1 and regulated GTPase activity of RAC1. In conclusion, these results demonstrated that PP4 was a novel regulator of hepatocyte lipoapoptosis and mediated hepatocyte lipoapoptosis by regulating the RAC1/MLK3/JNK signaling pathway. Our finding provided new insights into the mechanisms of this process.
Assuntos
Hepatócitos/metabolismo , Calicreínas/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Sistema de Sinalização das MAP Quinases , Neuropeptídeos/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Antígeno Prostático Específico/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular Tumoral , Hepatócitos/citologia , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MAP Quinase Quinase Quinase 11 Ativada por MitógenoRESUMO
Adiponectin is a small peptide secreted and a key component of the endocrine system and immune system. Although globular adiponectin protects myocardial ischemia/reperfusion-induced cardiomyocyte injury, the protective mechanisms remain largely unresolved. Using a neonatal rat ventricular myocyte hypoxia/reoxygenation model, we investigated the role of its potential mechanisms of necroptosis in globular adiponectin-mediated protection in hypoxia/reoxygenation-induced cardiomyocyte injury as compared to apoptosis. We found that globular adiponectin treatment attenuated cardiomyocyte injury as indicated by increased cell viability and reduced lactate dehydrogenase release following hypoxia/reoxygenation. Immunofluorescence staining and Western blotting demonstrated that both necroptosis and apoptosis were triggered by hypoxia/reoxygenation and diminished by globular adiponectin. Necrostatin-1 (RIP1-specific inhibitor) and Z-VAD-FMK (pan-caspase inhibitor) only mimicked the inhibition of necroptosis and apoptosis, respectively, by globular adiponectin in hypoxia/reoxygenation-treated cardiomyocytes. Globular adiponectin attenuated reactive oxygen species production, oxidative damage, and p38MAPK and NF-κB signaling, all important for necroptosis and apoptosis. Collectively, our study suggests that globular adiponectin inhibits hypoxia/reoxygenation-induced necroptosis and apoptosis in cardiomyocytes probably by reducing oxidative stress and interrupting p38MAPK signaling.
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Adiponectina/metabolismo , Traumatismo por Reperfusão Miocárdica/imunologia , Miócitos Cardíacos/patologia , Animais , Animais Recém-Nascidos , Apoptose/imunologia , Hipóxia Celular/imunologia , Sobrevivência Celular , Células Cultivadas , Meios de Cultura/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Traumatismo por Reperfusão Miocárdica/patologia , Miócitos Cardíacos/imunologia , Necroptose/imunologia , Estresse Oxidativo/imunologia , Gravidez , Cultura Primária de Células , Ratos , Espécies Reativas de Oxigênio/metabolismoRESUMO
Cholesterol efflux from macrophages is the initial step of reverse cholesterol transport, an important process for high-density lipoprotein-mediated atheroprotection. G protein-coupled receptor (GPR) 120, which functions as long-chain fatty acid receptor, is well known for its anti-inflammatory and insulin-sensitizing function in macrophages. However, the role of GPR120 on macrophage foam cell formation, the hallmark of atherosclerotic plaques, has not been verified. In this study, we found for the first time that stimulation of GPR120 by its agonist GW9508 elevated the expression of ATP-binding cassette transporters (ABC) A1 and ABCG1 in THP-1 macrophage-derived foam cells and Raw264.7 macrophages, and promoted ABCA1- and ABCG1-mediated cholesterol efflux and reduced cellular cholesteryl ester (CE) content as well. In addition, GPR120 activation was accompanied with the stimulation of AMPK pathway in macrophages; however, the effect of GPR120 on macrophage cholesterol efflux was largely abolished by AMPK inhibition. Moreover, the AMPK activity and the expression of ABCA1 and ABCG1 were markedly abrogated by knockdown of GPR120, or application of phospholipase C (PLC) inhibitor, calcium chelator, or CaMKK inhibitor. Because only free cholesterol can be effluxed from macrophages, we found that activation of AMPK could lead to increase both neutral CEs hydrolysis by upregulation of neutral cholesterol ester hydrolase expression and acid CEs hydrolysis by activation of ULK1. In conclusion, these results demonstrated that GPR120 facilitated ABCA1- and ABCG1-mediated cholesterol efflux through activation of PLC/Ca2+ /CaMKK/AMPK signaling pathway, which induced CE hydrolysis and elevated the expression of ABCA1 and ABCG1 in macrophages.
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Proteínas Quinases Ativadas por AMP/metabolismo , Cálcio/metabolismo , Ésteres do Colesterol/metabolismo , Colesterol/metabolismo , Macrófagos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Fosfolipases Tipo C/metabolismo , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Células Espumosas , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Metilaminas/farmacologia , Camundongos , Propionatos/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Transdução de SinaisRESUMO
The usage of doxorubicin is hampered by its life-threatening cardiotoxicity in clinical practice. Dexrazoxane is the only cardioprotective medicine approved by the FDA for preventing doxorubicin-induced cardiac toxicity. Nevertheless, the mechanism of dexrazoxane is incompletely understood. The aim of our study is to investigate the possible molecular mechanism of dexrazoxane against doxorubicin-induced cardiotoxicity. We established a doxorubicin-induced mouse and cardiomyocyte injury model. Male C57BL/6J mice were randomly distributed into a control group (Con), a doxorubicin treatment group (DOX), a doxorubicin plus dexrazoxane treatment group (DOX+DEX), and a dexrazoxane treatment group (DEX). Echocardiography and histology analyses were performed to evaluate heart function and structure. DNA laddering, qRT-PCR, and Western blot were performed on DOX-treated cardiomyocytes with/without DEX treatment in vitro. Cardiomyocytes were then transfected with miR-17-5p mimics or inhibitors in order to analyze its downstream target. Our results demonstrated that dexrazoxane has a potent effect on preventing cardiac injury induced by doxorubicin in vivo and in vitro by reducing cardiomyocyte apoptosis. MicroRNA plays an important role in cardiovascular diseases. Our data revealed that dexrazoxane could upregulate the expression of miR-17-5p, which plays a cytoprotective role in response to hypoxia by regulating cell apoptosis. Furthermore, the miRNA and protein analysis revealed that miR-17-5p significantly attenuated phosphatase and tensin homolog (PTEN) expression in cardiomyocytes exposed to doxorubicin. Taken together, dexrazoxane might exert a cardioprotective effect against doxorubicin-induced cardiomyocyte apoptosis by regulating the expression of miR-17-5p/PTEN cascade.
Assuntos
Apoptose/efeitos dos fármacos , Dexrazoxano/farmacologia , Doxorrubicina/efeitos adversos , MicroRNAs/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Animais , Cardiotoxicidade/tratamento farmacológico , Cardiotoxicidade/patologia , Sobrevivência Celular/efeitos dos fármacos , Dexrazoxano/metabolismo , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , PTEN Fosfo-Hidrolase/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
Mitochondria are currently known as novel targets for treating cancer, especially for tumors displaying multidrug resistance (MDR). This present study aimed to develop a mitochondria-targeted delivery system by using triphenylphosphonium cation (TPP+)-conjugated Brij 98 as the functional stabilizer to modify paclitaxel (PTX) nanocrystals (NCs) against drug-resistant cancer cells. Evaluations were performed on 2D monolayer and 3D multicellular spheroids (MCs) of MCF-7 cells and MCF-7/ADR cells. In comparison with free PTX and the non-targeted PTX NCs, the targeted PTX NCs showed the strongest cytotoxicity against both 2D MCF-7 and MCF-7/ADR cells, which was correlated with decreased mitochondrial membrane potential. The targeted PTX NCs exhibited deeper penetration on MCF-7 MCs and more significant growth inhibition on both MCF-7 and MCF-7/ADR MCs. The proposed strategy indicated that the TPP+-modified NCs represent a potentially viable approach for targeted chemotherapeutic molecules to mitochondria. This strategy might provide promising therapeutic outcomes to overcome MDR.
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Hyperglycaemia is a characteristic of type 2 diabetes. In hepatocytes, impaired insulin sensitivity leads to increased gluconeogenesis and decreased glycogenesis. MicroRNA (miR)3383p is associated with tumour necrosis factor (TNF)αinduced suppression of hepatic glycogenesis via regulation of protein phosphatase 4 regulatory subunit 1 (PP4R1). However, the effect of miR3383p on gluconeogenesis in hepatocytes remains unknown. In a previous study, it was demonstrated that miR3383p is downregulated in the livers of mice and in mouse HEPA16 hepatocytes following treatment with TNFα. In the present study, the effect of miR3383p on TNFαinduced gluconeogenesis in hepatocytes was investigated. The levels of phosphorylatedFOXO1/FOXO1, phosphoenolpyruvate carboxykinase (PEPCK), peroxisome proliferatoractivated receptor γ coactivator (PGC1α) and glucose6phosphatase (G6Pase) were measured by western blotting. The mRNA levels of PEPCK, PGC1α and G6Pase were determined by quantitative polymerase chain reaction. Pyruvate tolerance testing was used to determine the gluconeogenesis of mouse livers. The results demonstrated that treatment with TNFα resulted in increased levels of gluconeogenesis in the livers of mice and decreased miR3383p expression levels in HEPA16 cells. Overexpression of miR3383p reversed TNFαinduced glucose production via enhancement of phosphorylated forkhead box O1 levels and downregulation of the expression levels of genes associated with gluconeogenesis, including peroxisome proliferatoractivated receptor γ coactivator1α, phosphoenolpyruvate carboxykinase and glucose6phosphatase. However, inhibition of miR3383p expression was revealed to enhance gluconeogenesis in the livers of mice and in HEPA16 cells. Furthermore, downregulation of PP4R1 was revealed to attenuate the effect on glucose production following treatment with miR3383p inhibitors. In conclusion, the results of the present study revealed that miR3383p may be involved in TNFαmediated gluconeogenesis via targeting of PP4R1 in hepatocytes.
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Gluconeogênese , Hepatócitos/metabolismo , MicroRNAs/metabolismo , Fosfoproteínas Fosfatases/genética , Animais , Linhagem Celular , Regulação para Baixo/efeitos dos fármacos , Gluconeogênese/efeitos dos fármacos , Glucose/metabolismo , Hepatócitos/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Fosfoproteínas Fosfatases/metabolismo , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Ultraconserved (uc) RNAs, a class of long non-coding RNAs (lncRNAs), are conserved across humans, mice, and rats, but the physiological significance and pathological role of ucRNAs is largely unknown. Here we show that uc.372 is upregulated in the livers of db/db mice, HFD-fed mice, and NAFLD patients. Gain-of-function and loss-of-function studies indicate that uc.372 drives hepatic lipid accumulation in mice by promoting lipogenesis. We further demonstrate that uc.372 binds to pri-miR-195/pri-miR-4668 and suppresses maturation of miR-195/miR-4668 to regulate expression of genes related to lipid synthesis and uptake, including ACC, FAS, SCD1, and CD36. Finally, we identify that uc.372 is located downstream of the insulinoma-associated 2 (INSM2) gene that is transcriptionally activated by upstream transcription factor 1 (USF1). Our findings reveal a novel mechanism by which uc.372 drives hepatic steatosis through inhibition of miR-195/miR-4668 maturation to relieve miR-195/miR-4668-mediated suppression of functional target gene expression.
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Lipídeos/biossíntese , Fígado/metabolismo , MicroRNAs/genética , RNA Longo não Codificante/genética , Animais , Dieta Hiperlipídica , Ácido Graxo Sintases/genética , Ácido Graxo Sintases/metabolismo , Regulação da Expressão Gênica , Células Hep G2 , Humanos , Metabolismo dos Lipídeos/genética , Lipogênese/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/metabolismo , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismoRESUMO
As a member of miR-17-92 miRNA clusters, miR19a has been considered to regulate hepatic glycogenesis by mediating the PI3K/AKT signaling pathway. However, whether miR19a serves an important role in gluconeogenesis in hepatocytes remains unknown. In the present study, the impact of miR19a on gluconeogenesis in HEP16 cells and its mechanisms of action were investigated. It was observed that overexpression of miR19a led to decreased glucose production, accompanied by increased activation of the AKT/FOXO1 signaling pathway and downregulated expression of gluconeogenesisassociated genes, including peroxisome proliferatoractivated receptor γ coactivator 1α, phosphoenolpyruvate carboxykinase and glucose 6phosphatase in the HEP16 cells transfected with the miR19a mimic. In contrast, suppression of miR19a impaired the activation of the AKT/FOXO1 signaling pathway and increased the expression of gluconeogenesisassociated genes, accompanied by an elevated glucose production. Additionally, phosphatase and tensin homolog (PTEN) was identified as a target of miR19a and participated in the miR19amediated gluconeogenesis in hepatocytes. These findings provide mechanistic insight into the effects of miR19a on the regulation of the AKT/FOXO1 signaling pathway and the expression of gluconeogenesisassociated genes. MiR19a may mediate gluconeogenesis in hepatocytes by downregulating PTEN expression.
Assuntos
Regulação da Expressão Gênica , Gluconeogênese/genética , Glucose/metabolismo , Hepatócitos/metabolismo , MicroRNAs/genética , PTEN Fosfo-Hidrolase/genética , Animais , Antagomirs/genética , Antagomirs/metabolismo , Linhagem Celular , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Glucose-6-Fosfatase/genética , Glucose-6-Fosfatase/metabolismo , Hepatócitos/citologia , Fígado/citologia , Fígado/metabolismo , Camundongos , MicroRNAs/agonistas , MicroRNAs/antagonistas & inibidores , MicroRNAs/metabolismo , Oligorribonucleotídeos/genética , Oligorribonucleotídeos/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fosfoenolpiruvato Carboxiquinase (ATP)/genética , Fosfoenolpiruvato Carboxiquinase (ATP)/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
OBJECTIVE: Insulin resistance is a critical factor contributing to the pathogenesis of type 2 diabetes and other metabolic diseases. Recent studies have indicated that miR-338-3p plays an important role in cancer. Here, we investigated whether miR-338-3p mediates tumour necrosis factor-α (TNF-α)-induced hepatic insulin resistance. METHODS: The activation of the insulin signalling pathway and the level of glycogenesis were examined in the livers of the db/db and high fat diet (HFD)-fed mice and in HEP1-6 cells transfected with miR-338-3p mimic or inhibitor. Computational prediction of microRNA target, luciferase assay and Western blot were used to assess the miR-338-3p target. Chromatin immunoprecipitation (ChIP) assay was used to determine the transcriptional regulator of miR-338-3p. RESULTS: miR-338-3p was down-regulated in the livers of the db/db, HFD-fed and TNF-α-treated C57BL/6J mice, as well as in mouse HEP1-6 hepatocytes treated with TNF-α. Importantly the down-regulation of miR-338-3p induced insulin resistance, as indicated by impaired glucose tolerance and insulin tolerance. Further research showed that the down-regulated miR-338-3p resulted in the impaired AKT/ glycogen synthase kinase 3 beta (GSl·Gß) signalling pathway and glycogen synthesis. In contrast, hepatic over-expression of miR-338-3p rescued the TNF-α-induced insulin resistance. Moreover, protein phosphatase 4 regulator subunit 1 (PP4R1) was identified as a direct target of miR-338-3p that mediated hepatic insulin signalling by regulating protein phosphatase 4 (PP4). Finally we identified hepatic nuclear factor 4 alpha (HNF-4α) as the transcriptional regulator of miRNA-338-3p. CONCLUSIONS: Our studies provide novel insight into the critical role and molecular mechanism by which miR-338-3p is involved in TNF-α-induced hepatic insulin resistance. miR-338-3p might mediate TNF-α-induced hepatic insulin resistance by targeting PP4R1 to regulate PP4 expression.
Assuntos
Regulação da Expressão Gênica , Resistência à Insulina , Fígado/efeitos dos fármacos , MicroRNAs/metabolismo , Fosfoproteínas Fosfatases/genética , Fator de Necrose Tumoral alfa/farmacologia , Animais , Sequência de Bases , Linhagem Celular , Dieta Hiperlipídica , Glicogênio Sintase Quinase 3 beta/metabolismo , Fator 4 Nuclear de Hepatócito/antagonistas & inibidores , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Insulina/farmacologia , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Fosfoproteínas Fosfatases/química , Fosfoproteínas Fosfatases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , Transdução de SinaisRESUMO
Several studies have suggested an important role of miR-291b-3p in the development of embryonic stem cells. In previous study, we found that the expression of miR-291b-3p was significantly upregulated in the liver of db/db mice. However, the role of miR-291b-3p in glucose metabolism and its underlying mechanisms remain unknown. In the present study, we demonstrated that miR-291b-3p was abundantly expressed in the liver. Of note, hepatic miR-291b-3p expression was upregulated in HFD-fed mice and induced by fasting in C57BL/6 J normal mice. Importantly, hepatic inhibition miR-291b-3p expression ameliorated hyperglycemia and insulin resistance in HFD-fed mice, whereas hepatic overexpression of miR-291b-3p led to hyperglycemia and insulin resistance in C57BL/6 J normal mice. Further study revealed that miR-291b-3p suppressed insulin-stimulated AKT/GSK signaling and increased the expression of gluconeogenic genes in hepatocytes. Moreover, we identified that p65, a subunit of nuclear factor-κB (NF-κB), is a target of miR-291b-3p by bioinformatics analysis and luciferase reporter assay. Silencing of p65 significantly augmented the expression of PTEN and impaired AKT activation. In conclusion, we found novel evidence suggesting that hepatic miR-291b-3p mediated glycogen synthesis and gluconeogenesis through targeting p65 to regulate PTEN expression. Our findings indicate the therapeutic potential of miR-291b-3p inhibitor in hyperglycemia and insulin resistance.
Assuntos
Glucose/metabolismo , Fígado/metabolismo , MicroRNAs/genética , Fator de Transcrição RelA/metabolismo , Animais , Linhagem Celular , Gluconeogênese , Glicogênio Hepático/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Fator de Transcrição RelA/genética , Regulação para CimaRESUMO
Doxorubicin (adriamycin), an anthracycline antibiotic, is commonly used to treat many types of solid and hematological malignancies. Unfortunately, clinical usage of doxorubicin is limited due to the associated acute and chronic cardiotoxicity. Previous studies demonstrated that Astragalus polysaccharide (APS), the extracts of Astragalus membranaceus, had strong anti-tumor activities and anti-inflammatory effects. However, whether APS could mitigate chemotherapy-induced cardiotoxicity is unclear thus far. We used a doxorubicin-induced neonatal rat cardiomyocyte injury model and a mouse heart failure model to explore the function of APS. GFP-LC3 adenovirus-mediated autophagic vesicle assays, GFP and RFP tandemly tagged LC3 (tfLC3) assays and Western blot analyses were performed to analyze the cell function and cell signaling changes following APS treatment in cardiomyocytes. First, doxorubicin treatment led to C57BL/6J mouse heart failure and increased cardiomyocyte apoptosis, with a disturbed cell autophagic flux. Second, APS restored autophagy in doxorubicin-treated primary neonatal rat ventricular myocytes and in the doxorubicin-induced heart failure mouse model. Third, APS attenuated doxorubicin-induced heart injury by regulating the AMPK/mTOR pathway. The mTOR inhibitor rapamycin significantly abrogated the protective effect of APS. These results suggest that doxorubicin could induce heart failure by disturbing cardiomyocyte autophagic flux, which may cause excessive cell apoptosis. APS could restore normal autophagic flux, ameliorating doxorubicin-induced cardiotoxicity by regulating the AMPK/mTOR pathway.