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T-cell acute lymphoblastic leukemia (T-ALL) is a highly aggressive hematologic malignancy originating from T progenitor cells. It accounts for 15% of childhood and 25% of adult ALL cases. GNE-987 is a novel chimeric molecule developed using proteolysis-targeting chimeras (PROTAC) technology for targeted therapy. It consists of a potent inhibitor of the bromodomain and extraterminal (BET) protein, as well as the E3 ubiquitin ligase Von Hippel-Lindau (VHL), which enables the effective induction of proteasomal degradation of BRD4. Although GNE-987 has shown persistent inhibition of cell proliferation and apoptosis, its specific antitumor activity in T-ALL remains unclear. In this study, we aimed to investigate the molecular mechanisms underlying the antitumor effect of GNE-987 in T-ALL. To achieve this, we employed technologies including RNA sequencing (RNA-seq), chromatin immunoprecipitation sequencing (ChIP-seq) and CUT&Tag. The degradation of BET proteins, specifically BRD4, by GNE-987 has a profound impact on T-ALL cell. In in vivo experiments, sh-BRD4 lentivirus reduced T-ALL cell proliferation and invasion, extending the survival time of mice. The RNA-seq and CUT&Tag analyses provided further insights into the mechanism of action of GNE-987 in T-ALL. These analyses revealed that GNE-987 possesses the ability to suppress the expression of various genes associated with super-enhancers (SEs), including lymphoblastic leukemia 1 (LCK). By targeting these SE-associated genes, GNE-987 effectively inhibits the progression of T-ALL. Importantly, SE-related oncogenes like LCK were identified as critical targets of GNE-987. Based on these findings, GNE-987 holds promise as a potential novel candidate drug for the treatment of T-ALL.
Assuntos
Apoptose , Proliferação de Células , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Fatores de Transcrição , Ensaios Antitumorais Modelo de Xenoenxerto , Humanos , Animais , Camundongos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Proliferação de Células/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Elementos Facilitadores Genéticos , Proteínas que Contêm BromodomínioRESUMO
BACKGROUND: Acute myeloid leukemia (AML) is a malignancy of the hematopoietic system, and childhood AML accounts for about 20% of pediatric leukemia. ANP32B, an important nuclear protein associated with proliferation, has been found to regulate hematopoiesis and CML leukemogenesis by inhibiting p53 activity. However, recent study suggests that ANP32B exerts a suppressive effect on B-cell acute lymphoblastic leukemia (ALL) in mice by activating PU.1. Nevertheless, the precise underlying mechanism of ANP32B in AML remains elusive. RESULTS: Super enhancer related gene ANP32B was significantly upregulated in AML patients. The expression of ANP32B exhibited a negative correlation with overall survival. Knocking down ANP32B suppressed the proliferation of AML cell lines MV4-11 and Kasumi-1, along with downregulation of C-MYC expression. Additionally, it led to a significant decrease in H3K27ac levels in AML cell lines. In vivo experiments further demonstrated that ANP32B knockdown effectively inhibited tumor growth. CONCLUSIONS: ANP32B plays a significant role in promoting tumor proliferation in AML. The downregulation of ANP32B induces cell cycle arrest and promotes apoptosis in AML cell lines. Mechanistic analysis suggests that ANP32B may epigenetically regulate the expression of MYC through histone H3K27 acetylation. ANP32B could serve as a prognostic biomarker and potential therapeutic target for AML patients.
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BACKGROUND: Osteosarcoma (OS) is one of the most common primary malignant tumors of bone in children, which develops from osteoblasts and typically occurs during the rapid growth phase of the bone. Recently, Super-Enhancers(SEs)have been reported to play a crucial role in osteosarcoma growth and metastasis. Therefore, there is an urgent need to identify specific targeted inhibitors of SEs to assist clinical therapy. This study aimed to elucidate the role of BRD4 inhibitor GNE-987 targeting SEs in OS and preliminarily explore its mechanism. METHODS: We evaluated changes in osteosarcoma cells following treatment with a BRD4 inhibitor GNE-987. We assessed the anti-tumor effect of GNE-987 in vitro and in vivo by Western blot, CCK8, flow cytometry detection, clone formation, xenograft tumor size measurements, and Ki67 immunohistochemical staining, and combined ChIP-seq with RNA-seq techniques to find its anti-tumor mechanism. RESULTS: In this study, we found that extremely low concentrations of GNE-987(2-10 nM) significantly reduced the proliferation and survival of OS cells by degrading BRD4. In addition, we found that GNE-987 markedly induced cell cycle arrest and apoptosis in OS cells. Further study indicated that VHL was critical for GNE-987 to exert its antitumor effect in OS cells. Consistent with in vitro results, GNE-987 administration significantly reduced tumor size in xenograft models with minimal toxicity, and partially degraded the BRD4 protein. KRT80 was identified through analysis of the RNA-seq and ChIP-seq data. U2OS HiC analysis suggested a higher frequency of chromatin interactions near the KRT80 binding site. The enrichment of H3K27ac modification at KRT80 was significantly reduced after GNE-987 treatment. KRT80 was identified as playing an important role in OS occurrence and development. CONCLUSIONS: This research revealed that GNE-987 selectively degraded BRD4 and disrupted the transcriptional regulation of oncogenes in OS. GNE-987 has the potential to affect KRT80 against OS.
Assuntos
Apoptose , Neoplasias Ósseas , Proteínas de Ciclo Celular , Proliferação de Células , Osteossarcoma , Fatores de Transcrição , Animais , Humanos , Camundongos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Proteínas que Contêm Bromodomínio , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/antagonistas & inibidores , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Elementos Facilitadores Genéticos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Camundongos Nus , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Osteossarcoma/genética , Osteossarcoma/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Glioblastoma (GBM) is a relatively prevalent primary tumor of the central nervous system in children, characterized by its high malignancy and mortality rates, along with the intricate challenges of achieving complete surgical resection. Recently, an increasing number of studies have focused on the crucial role of super-enhancers (SEs) in the occurrence and development of GBM. This study embarks on the task of evaluating the effectiveness of MZ1, an inhibitor of BRD4 meticulously designed to specifically target SEs, within the intricate framework of GBM. METHODS: The clinical data of GBM patients was sourced from the Chinese Glioma Genome Atlas (CGGA) and the Gene Expression Profiling Interactive Analysis 2 (GEPIA2), and the gene expression data of tumor cell lines was derived from the Cancer Cell Line Encyclopedia (CCLE). The impact of MZ1 on GBM was assessed through CCK-8, colony formation assays, EdU incorporation analysis, flow cytometry, and xenograft mouse models. The underlying mechanism was investigated through RNA-seq and ChIP-seq analyses. RESULTS: In this investigation, we made a noteworthy observation that MZ1 exhibited a substantial reduction in the proliferation of GBM cells by effectively degrading BRD4. Additionally, MZ1 displayed a notable capability in inducing significant cell cycle arrest and apoptosis in GBM cells. These findings were in line with our in vitro outcomes. Notably, MZ1 administration resulted in a remarkable decrease in tumor size within the xenograft model with diminished toxicity. Furthermore, on a mechanistic level, the administration of MZ1 resulted in a significant suppression of pivotal genes closely associated with cell cycle regulation and epithelial-mesenchymal transition (EMT). Interestingly, our analysis of RNA-seq and ChIP-seq data unveiled the discovery of a novel prospective oncogene, SDC1, which assumed a pivotal role in the tumorigenesis and progression of GBM. CONCLUSION: In summary, our findings revealed that MZ1 effectively disrupted the aberrant transcriptional regulation of oncogenes in GBM by degradation of BRD4. This positions MZ1 as a promising candidate in the realm of therapeutic options for GBM treatment.
Assuntos
Neoplasias Encefálicas , Proteínas que Contêm Bromodomínio , Glioblastoma , Animais , Criança , Humanos , Camundongos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Proteínas que Contêm Bromodomínio/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Glioblastoma/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Estudos Prospectivos , Sindecana-1/antagonistas & inibidores , Fatores de Transcrição/genéticaRESUMO
Acute myeloid leukemia (AML) is a rare and heterogeneous disease. Over the past few decades, patient prognosis has improved with continuous improvements in treatment, but outcomes for some patients with primary drug resistance or relapse after treatment remain poor. Additional therapies to improve outcomes for these patients are urgently needed. FYB1 expression differs substantially between AML tissues and normal tissues. High FYB1 expression is correlated with poorer overall survival (OS), indicating that FYB1 may regulate AML progression. Therefore, understanding the effect of FYB1 on AML could improve the success rate of therapeutic approaches and prognosis for patients with AML. In this study, through analysis of large databases and both in vivo and in vitro experiments, we assessed the expression and role of FYB1 in AML and the relationship of FYB with patient prognosis. Downstream targets of the FYB1 gene were analyzed by RNA-seq. Database mining and in vitro experiments were used to further clarify the effect of the downstream target gelsolin-like actin-capping protein (CAPG) on AML cells and its relationship with patient prognosis. FYB1 expression was significantly higher in AML tissue and corresponded with a poor prognosis. FYB1 knockdown inhibited AML cell proliferation, promoted cell apoptosis, reduced cell adhesion capability and significantly reduced the tumor formation rate in mice. In addition, FYB1 knockdown induced a notable decrease in CAPG expression. The suppression of CAPG significantly inhibited cell proliferation and increased cell apoptosis. The conclusions of this study underscore the pivotal role of the FYB1/CAPG axis in promoting AML. We propose that the FYB1/CAPG axis could serve as a new thread in the development of therapeutic strategies for AML.
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BACKGROUND: Overweight/obesity is considered an independent risk factor for nephrolithiasis, but little is known about its effect on nephrolithiasis according to metabolic health status. OBJECTIVES: We aimed to investigate the association between various metabolic overweight phenotypes and the occurrence of nephrolithiasis. It also explores whether changes in these phenotypes over time influence the risk of nephrolithiasis. MATERIALS AND METHODS: A total of 10,315 participants free of nephrolithiasis who underwent an annual health checkup from 2017 to 2022 were included in our prospective cohort study. They were categorized into four groups according to the presence of overweight and metabolic abnormalities (MA). The primary endpoint was the occurrence of renal stones. Multivariable Cox analysis was conducted to elucidate the relationship between metabolic overweight phenotypes and incident nephrolithiasis. RESULTS: During a median follow-up duration of 4.02 years, nephrolithiasis occurred in 1,468 (14.23%) participants. In the full cohort, we observed that the 5-year cumulative incidences of nephrolithiasis were highest in the metabolically healthy overweight (MHO) and metabolically abnormal overweight (MAO) groups. The hazard ratios (HRs) for nephrolithiasis, relative to metabolically healthy normal weight (MHNW), ranged from 1.19 (95% CI:1.03-1.37; MHO) to 1.32 (95% CI:1.15-1.51; MAO). Furthermore, individuals with persistent MHO throughout follow-up were at a 1.42-fold increased risk of nephrolithiasis (P < 0.001), and 32.17% of individuals experienced changes in phenotype during follow-up. Among MAO subjects, those who transitioned to MHO and MHNW had a 26% and 45% lower risk of incident nephrolithiasis, respectively, compared to those who persisted in the MAO phenotype. CONCLUSION: Individuals in the MHO and MAO groups exhibit an elevated risk of incident nephrolithiasis in this prospective cohort study. A significant proportion of nephrolithiasis cases may be potentially preventable through the appropriate management of metabolic risk factors for MAO subjects.
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Nefrolitíase , Sobrepeso , Fenótipo , Humanos , Masculino , Feminino , Nefrolitíase/epidemiologia , Pessoa de Meia-Idade , Sobrepeso/epidemiologia , Adulto , Estudos Prospectivos , Fatores de Risco , Incidência , Estudos de CoortesRESUMO
AIMS: It is acknowledged that aberrant liver immunity contributes to the development of type 2 diabetes mellitus (T2DM). Mucosal-associated invariant T (MAIT) cells, an innate-like T-cell subset, are enriched in the human liver. Nevertheless, the characterisation and potential role of hepatic MAIT cells in T2DM remain unclear. MATERIALS AND METHODS: Fourteen newly diagnosed T2DM subjects and 15 controls received liver biopsy. The frequency and cytokine production of MAIT cells were analysed by flow cytometry. The expression of genes involved in glucose metabolism was determined in HepG2 cells co-cultured with hepatic MAIT cells. RESULTS: Compared with controls, hepatic MAIT cell frequency was significantly increased in T2DM patients (24.66% vs. 14.61%, p = 0.001). There were more MAIT cells producing interferon-γ (IFN-γ, 60.49% vs. 33.33%, p = 0.021) and tumour necrosis factor-α (TNF-α, 46.84% vs. 5.91%, p = 0.021) in T2DM than in controls, whereas their production of interleukin 17 (IL-17) was comparable (15.25% vs. 4.55%, p = 0.054). Notably, an IFN-γ+ TNF-α+ IL-17+/- producing MAIT cell subset was focussed, which showed an elevated proportion in T2DM (42.66% vs. 5.85%, p = 0.021) and positively correlated with plasma glucose levels. A co-culture experiment further indicated that hepatic MAIT cells from T2DM upregulated the gene expression of pyruvate carboxylase, a key molecule involved in gluconeogenesis, in HepG2 cells, and this response was blocked with neutralising antibodies against IFN-γ and TNF-α. CONCLUSIONS: Our data implicate an increased Th1-like MAIT cell subset in the liver of newly diagnosed T2DM subjects, which induces hyperglycaemia by promoting hepatic gluconeogenesis. It provides novel insights into the immune regulation of metabolic homoeostasis. CLINICAL TRIAL REGISTRATION NUMBER: NCT03296605 (registered at www. CLINICALTRIALS: gov on 12 October 2018).
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Diabetes Mellitus Tipo 2 , Células T Invariantes Associadas à Mucosa , Humanos , Células T Invariantes Associadas à Mucosa/fisiologia , Interleucina-17 , Fator de Necrose Tumoral alfa , Gluconeogênese , FígadoRESUMO
T cell activation initiates protective adaptive immunity, but counterbalancing mechanisms are critical to prevent overshooting responses and to maintain immune homeostasis. The CARD11-BCL10-MALT1 (CBM) complex bridges T cell receptor engagement to NF-κB signaling and MALT1 protease activation. Here, we show that ABIN-1 is modulating the suppressive function of A20 in T cells. Using quantitative mass spectrometry, we identified ABIN-1 as an interactor of the CBM signalosome in activated T cells. A20 and ABIN-1 counteract inducible activation of human primary CD4 and Jurkat T cells. While A20 overexpression is able to silence CBM complex-triggered NF-κB and MALT1 protease activation independent of ABIN-1, the negative regulatory function of ABIN-1 depends on A20. The suppressive function of A20 in T cells relies on ubiquitin binding through the C-terminal zinc finger (ZnF)4/7 motifs, but does not involve the deubiquitinating activity of the OTU domain. Our mechanistic studies reveal that the A20/ABIN-1 module is recruited to the CBM complex via A20 ZnF4/7 and that proteasomal degradation of A20 and ABIN-1 releases the CBM complex from the negative impact of both regulators. Ubiquitin binding to A20 ZnF4/7 promotes destructive K48-polyubiquitination to itself and to ABIN-1. Further, after prolonged T cell stimulation, ABIN-1 antagonizes MALT1-catalyzed cleavage of re-synthesized A20 and thereby diminishes sustained CBM complex signaling. Taken together, interdependent post-translational mechanisms are tightly controlling expression and activity of the A20/ABIN-1 silencing module and the cooperative action of both negative regulators is critical to balance CBM complex signaling and T cell activation.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Linfócitos T/metabolismo , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/fisiologia , Proteína 10 de Linfoma CCL de Células B/metabolismo , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Células Cultivadas , Guanilato Ciclase/metabolismo , Células HEK293 , Humanos , Células Jurkat , Ativação Linfocitária/genética , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/metabolismo , Complexos Multiproteicos/metabolismo , NF-kappa B/metabolismo , Ligação Proteica , Interferência de RNA/imunologia , Transdução de Sinais/fisiologia , Linfócitos T/imunologiaRESUMO
Sulfur and selenium occupy a distinguished position in biology owing to their redox activities, high nucleophilicity, and acyl transfer capabilities. Thiolated/selenolated amino acids, including cysteine, selenocysteine, and their derivatives, play critical roles in regulating the conformation and function of proteins and serve as an important motif for peptide design and bioconjugation. Unfortunately, a general and concise method to attain enantiopure ß-thiolated/selenolated amino acids remains an unsolved problem. Herein, we present a photoredox-catalyzed asymmetric method for the preparation of enantiopure ß-thiolated/selenolated amino acids using a simple chiral auxiliary, which controls the diastereoselectivity of the key alkylation step and acts as an orthogonal protecting group in the subsequent peptide synthesis. Our protocol can be used to prepare a wide range of ß-thiolated/selenolated amino acids on a gram scale, which would otherwise be difficult to obtain using conventional methods. The effect of our chemistry was further highlighted and validated through the preparation of a series of peptidyl thiol/selenol analogues, including cytochrome c oxidase subunit protein 7C and oxytocin.
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Aminoácidos/síntese química , Selênio/química , Compostos de Sulfidrila/química , Aminoácidos/química , Catálise , Conformação Molecular , Oxirredução , Processos FotoquímicosRESUMO
Here, we describe a convergent synthesis of the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) harboring both O- and N-glycosylation sites using a Se-auxiliary-mediated ligation protocol together with native chemical ligation methodology. The robust and rapid Se-mediated ligation enables assembly of the N-terminus of hGM-CSF in just 4 h, which is significantly faster than the traditional ligation/desulfurization method. Therefore, this new methodology could help to produce hGM-CSF glycoform libraries more efficiently for future elucidation of the importance of glycosylation.
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Fator Estimulador de Colônias de Granulócitos e Macrófagos , Fenômenos Biofísicos , Glicosilação , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , HumanosRESUMO
BACKGROUND/AIMS: Cardiac fibrosis is an important cardiac remodeling event that can ultimately lead to the development of severe arrhythmia and heart failure. MicroRNAs (miRNAs) are involved in the pathogenesis of many cardiovascular diseases. Here, we aimed to investigate the effects of caveolin-3 (Cav3) on the pathogenesis of cardiac fibrosis and the underlying molecular mechanisms. METHODS: Cav3 expression was decreased in cardiac fibrosis in vivo and in vitro model. To investigate the role of Cav3 in cardiac fibrosis, we transfected cardiac fibroblasts (CFs) with the siRNA of Cav3 and Cav3-overexpressing plasmid. The collagen content and proliferation of CFs were detected by qRT-PCR, western blot, MTT, and immunofluorescence. A luciferase reporter gene assay and gain/loss of function were used to detect the relationship between miR-22 and Cav3. RESULTS: Cav3 depletion in CFs induced an increase in collagen content, cell proliferation, and phenotypic conversion of fibroblasts to myofibroblasts. Conversely, Cav3 overexpression in CFs was shown to inhibit angiotensin II-mediated excessive collagen deposition through protein kinase C (PKC)ε inactivation. Cav3 was experimentally confirmed as a direct target of miR-22, containing two seed binding sites in its 3'-untranslated region, and miR-22 was demonstrated to be significantly upregulated in the ischemic border zone in mice after myocardial infarction and in neonatal rat CFs pretreated with angiotensin II. miR-22 overexpression increased CFs proliferation, and collagen and α-smooth muscle actin levels in CFs, while the knockdown of endogenous miR-22 decreased CFs numbers. CONCLUSIONS: Our findings demonstrate that miR-22 accelerates cardiac fibrosis through the miR-22-Cav3-PKCε pathway, which, therefore, may represent a new therapeutic target for treatment of excessive fibrosis-associated cardiac diseases.
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Caveolina 3/metabolismo , MicroRNAs/metabolismo , Infarto do Miocárdio/patologia , Actinas/metabolismo , Angiotensina II/farmacologia , Animais , Sequência de Bases , Caveolina 3/antagonistas & inibidores , Caveolina 3/genética , Células Cultivadas , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Cadeia alfa 1 do Colágeno Tipo I , Colágeno Tipo III/genética , Colágeno Tipo III/metabolismo , Modelos Animais de Doenças , Regulação para Baixo , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Masculino , Camundongos , MicroRNAs/genética , Infarto do Miocárdio/metabolismo , Miocárdio/citologia , Miocárdio/metabolismo , Proteína Quinase C-épsilon/metabolismo , Interferência de RNA , Ratos , Ratos Sprague-DawleyRESUMO
Nonalcoholic fatty liver disease (NAFLD)1 is the most common chronic liver disease worldwide. Cichorium glandulosum Boiss. et Huet (CG) is a common traditional Uighur medicine, and it has been widely used as active therapy on various hepatic diseases. Recently, lipid-lowering effect has been revealed on CG. Polysaccharides are principal component of CG which could be the possible lipid-lowering compound in CG. In this study, extraction and purification of CG polysaccharides (CGP) was performed, and the lipid regulation effect of CGP was investigated on NAFLD zebrafish model. The results showed that CGP significantly decreased the levels of TC, TG, and decreased the mRNA expression of srebf-1, and fas, increased the expression of pparab. The findings suggest that the lipid-lowering effects of CGP mainly depend on facilitation of lipolysis (mainly beta-oxidation) or inhibition of lipogenesis. Furthermore, CGP could prevent and causes the regression of steatosis in NAFLD via its lipid metabolism regulation effect.
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Asteraceae , Hipolipemiantes/uso terapêutico , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Polissacarídeos/uso terapêutico , Animais , Modelos Animais de Doenças , Hipolipemiantes/farmacologia , Larva , Metabolismo dos Lipídeos/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , Fitoterapia , Polissacarídeos/farmacologia , Transcriptoma/efeitos dos fármacos , Peixe-ZebraRESUMO
Isoquercetin (IQ), a glucoside derivative of quercetin, has been reported to have beneficial effects in nonalcoholic fatty liver disease (NAFLD). In this study, we investigated the potential improvement of IQ in liver lipid accumulation, inflammation, oxidative condition, and activation in Kupffer cells (KCs) on a high-fat diet (HFD) induced NAFLD models. Male Sprague-Dawley (SD) rats were induced by HFD, lipopolysaccharides/free fatty acids (LPS/FFA) induced co-culture cells model between primary hepatocytes and Kupffer cells was used to test the effects and the underlying mechanism of IQ. Molecular docking was performed to predict the potential target of IQ. Significant effects of IQ were found on reduced lipid accumulation, inflammation, and oxidative stress. In addition, AMP-activated protein kinase (AMPK) pathway was activated by IQ, and is plays an important role in lipid regulation. Meanwhile, IQ reversed the increase of activated KCs which caused by lipid overload, and also suppression of Transforming growth factor beta (TGF-ß) signaling by TGF-ß Recptor-1 and SMAD2/3 signaling. Finally, TGF-ßR1 and TGF-ßR2 were both found may involve in the mechanism of IQ. IQ can improve hepatic lipid accumulation and decrease inflammation and oxidative stress by its activating AMPK pathway and suppressing TGF-ß signaling to alleviate NAFLD.
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Proteínas Quinases Ativadas por AMP/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo , Quercetina/análogos & derivados , Fator de Crescimento Transformador beta/metabolismo , Animais , Biomarcadores/sangue , Técnicas de Cocultura , Citocinas/sangue , Dieta Hiperlipídica , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Inflamação/sangue , Células de Kupffer/metabolismo , Fígado/efeitos dos fármacos , Masculino , Simulação de Acoplamento Molecular , Hepatopatia Gordurosa não Alcoólica/sangue , Estresse Oxidativo/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Quercetina/farmacologia , Quercetina/uso terapêutico , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND/AIMS: Acute myocardial infarction (AMI) is a devastating cardiovascular disease with a high rate of morbidity and mortality, partly due to enhanced arrhythmogenicity. MicroRNAs (miRNAs) have been shown to participate in the regulation of cardiac ion channels and the associated arrhythmias. The purpose of this study was to test our hypothesis that miR-223-3p contributes to the electrical disorders in AMI via modulating KCND2, the gene encoding voltage-gated channel Kv4.2 that carries transient outward K+ current Ito. METHODS: AMI model was established in male Sprague-Dawley (SD) rats by left anterior descending artery (LAD) ligation. Evans blue and TTC staining was used to measure infarct area. Ito was recorded in isolated ventricular cardiomyocytes or cultured neonatal rat ventricular cells (NRVCs) by whole-cell patch-clamp techniques. Western blot analysis was employed to detect the protein level of Kv4.2 and real-time RT-PCR to determine the transcript level of miR-223-3p. Luciferase assay was used to examine the interaction between miR-223-3p and KCND2 in cultured NRVCs. RESULTS: Expression of miR-223-3p was remarkably upregulated in AMI relative to sham control rats. On the contrary, the protein level of Kv4.2 and Ito density were significantly decreased in AMI. Consistently, transfection of miR-223-3p mimic markedly reduced Kv4.2 protein level and Ito current in cultured NRVCs. Co-transfection of AMO-223-3p (an antisense inhibitor of miR-223-3p) reversed the repressive effect of miR-223-3p. Luciferase assay showed that miR-223-3p, but not the negative control, substantially suppressed the luciferase activity, confirming the direct binding of miR-223-3p to the seed site within the KCND2 sequence. Finally, direct intramuscular injection of AMO-223-3p into the ischemic myocardium to knockdown endogenous miR-223-3p decreased the propensity of ischemic arrhythmias. CONCLUSIONS: Upregulation of miR-223-3p in AMI repressed the expression of KCND2/Kv4.2 resulting in reduction of Ito density that can cause APD prolongation and promote arrhythmias in AMI, and therefore knockdown of endogenous miR-223-3p might be considered a new approach for antiarrhythmic therapy of ischemic arrhythmias.
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Regulação da Expressão Gênica , MicroRNAs/genética , Infarto do Miocárdio/genética , Canais de Potássio Shal/genética , Animais , Animais Recém-Nascidos , Western Blotting , Células Cultivadas , Ativação do Canal Iônico/genética , Ativação do Canal Iônico/fisiologia , Masculino , Potenciais da Membrana/genética , Potenciais da Membrana/fisiologia , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/fisiopatologia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/fisiologia , Técnicas de Patch-Clamp , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Canais de Potássio Shal/metabolismo , Canais de Potássio Shal/fisiologiaRESUMO
The pituitary is the central endocrine gland with effects on metabolic dysfunction-associated steatotic liver disease (MASLD). However, it is not clear whether the pituitary responds to free fatty acid (FFA) toxicity, thus dysregulating hepatic lipid metabolism. Here, we demonstrate that decreased prolactin (PRL) levels are involved in the association between FFA and MASLD based on a liver biospecimen-based cohort. Moreover, overloaded FFAs decrease serum PRL levels, thus promoting liver steatosis in mice with both dynamic diet intervention and stereotactic pituitary FFA injection. Mechanistic studies show that excessive FFA sensing in pituitary lactotrophs inhibits the synthesis and secretion of PRL in a cell-autonomous manner. Notably, inhibiting excessive lipid uptake using pituitary stereotaxic virus injection or a specific drug delivery system effectively ameliorates hepatic lipid accumulation by improving PRL levels. Targeted inhibition of pituitary FFA sensing may be a potential therapeutic target for liver steatosis.
Assuntos
Ácidos Graxos não Esterificados , Fígado Gorduroso , Lactotrofos , Prolactina , Animais , Prolactina/metabolismo , Prolactina/sangue , Ácidos Graxos não Esterificados/sangue , Ácidos Graxos não Esterificados/metabolismo , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Camundongos , Lactotrofos/metabolismo , Lactotrofos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Humanos , Masculino , Metabolismo dos Lipídeos , Fígado/metabolismoRESUMO
Neuroblastoma (NB) is a common childhood tumor with a high incidence worldwide. The regulatory role of RNA N6-methyladenosine (m6A) in gene expression has attracted significant attention, and the impact of methyltransferase-like 14 (METTL14) on tumor progression has been extensively studied in various types of cancer. However, the specific influence of METTL14 on NB remains unexplored. Using data from the Target database, our study revealed significant upregulation of METTL14 expression in high-risk NB patients, with strong correlation with poor prognosis. Furthermore, we identified ETS1 and YY1 as upstream regulators that control the expression of METTL14. In vitro experiments involving the knockdown of METTL14 in NB cells demonstrated significant inhibition of cell proliferation, migration, and invasion. In addition, suppressing METTL14 inhibited NB tumorigenesis in nude mouse models. Through MeRIP-seq and RNA-seq analyses, we further discovered that YWHAH is a downstream target gene of METTL14. Mechanistically, we observed that methylated YWHAH transcripts, particularly those in the 5' UTR, were specifically recognized by the m6A "reader" protein YTHDF1, leading to the degradation of YWHAH mRNA. Moreover, the downregulation of YWHAH expression activated the PI3K/AKT signaling pathway, promoting NB cell activity. Overall, our study provides valuable insights into the oncogenic effects of METTL14 in NB cells, highlighting its role in inhibiting YWHAH expression through an m6A-YTHDF1-dependent mechanism. These findings also suggest the potential utility of a biomarker panel for prognostic prediction in NB patients.
RESUMO
BACKGROUND: Despite the use of targeted therapeutic approaches, T-cell acute lymphoblastic leukemia (T-ALL) is still associated with a high incidence of complications and a poor prognosis. Indisulam (also known as E7070), a newly identified molecular glue compound, has demonstrated increased therapeutic efficacy in several types of cancer through the rapid degradation of RBM39. This study aimed to evaluate the therapeutic potential of indisulam in T-ALL, elucidate its underlying mechanisms and explore the role of the RBM39 gene. METHODS: We verified the anticancer effects of indisulam in both in vivo and in vitro models. Additionally, the construction of RBM39-knockdown cell lines using shRNA confirmed that the malignant phenotype of T-ALL cells was dependent on RBM39. Through RNA sequencing, we identified indisulam-induced splicing anomalies, and proteomic analysis helped pinpoint protein changes caused by the drug. Comprehensive cross-analysis of these findings facilitated the identification of downstream effectors and subsequent validation of their functional roles. RESULTS: Indisulam has significant antineoplastic effects on T-ALL. It attenuates cell proliferation, promotes apoptosis and interferes with cell cycle progression in vitro while facilitating tumor remission in T-ALL in vivo models. This investigation provides evidence that the downregulation of RBM39 results in the restricted proliferation of T-ALL cells both in vitro and in vivo, suggesting that RBM39 is a potential target for T-ALL treatment. Indisulam's efficacy is attributed to its ability to induce RBM39 degradation, causing widespread aberrant splicing and abnormal translation of the critical downstream effector protein, THOC1, ultimately leading to protein depletion. Moreover, the presence of DCAF15 is regarded as critical for the effectiveness of indisulam, and its absence negates the ability of indisulam to induce the desired functional alterations. CONCLUSION: Our study revealed that indisulam, which targets RBM39 to induce tumor cell apoptosis, is an effective drug for treating T-ALL. Targeting RBM39 through indisulam leads to mis-splicing of pre-mRNAs, resulting in the loss of key effectors such as THOC1.
Assuntos
Leucemia-Linfoma Linfoblástico de Células T Precursoras , Proteínas de Ligação a RNA , Humanos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Camundongos , Animais , Linhagem Celular Tumoral , Apoptose/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proliferação de Células/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , Splicing de RNA , Sulfonamidas/farmacologia , FemininoRESUMO
Super enhancers (SEs) are genomic regions comprising multiple closely spaced enhancers, typically occupied by a high density of cell-type-specific master transcription factors (TFs) and frequently enriched in key oncogenes in various tumors, including neuroblastoma (NB), one of the most prevalent malignant solid tumors in children originating from the neural crest. Cyclin-dependent kinase 5 regulatory subunit-associated protein 3 (CDK5RAP3) is a newly identified super-enhancer-driven gene regulated by master TFs in NB; however, its function in NB remains unclear. Through an integrated study of publicly available datasets and microarrays, we observed a significantly elevated CDK5RAP3 expression level in NB, associated with poor patient prognosis. Further research demonstrated that CDK5RAP3 promotes the growth of NB cells, both in vitro and in vivo. Mechanistically, defective CDK5RAP3 interfered with the UFMylation system, thereby triggering endoplasmic reticulum (ER) phagy. Additionally, we provide evidence that CDK5RAP3 maintains the stability of MEIS2, a master TF in NB, and in turn, contributes to the high expression of CDK5RAP3. Overall, our findings shed light on the molecular mechanisms by which CDK5RAP3 promotes tumor progression and suggest that its inhibition may represent a novel therapeutic strategy for NB.
Assuntos
Proteínas de Ciclo Celular , Regulação Neoplásica da Expressão Gênica , Neuroblastoma , Fatores de Transcrição , Animais , Humanos , Camundongos , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Retículo Endoplasmático/metabolismo , Elementos Facilitadores Genéticos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos Nus , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neuroblastoma/genética , Neuroblastoma/patologia , Neuroblastoma/metabolismo , Prognóstico , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
BACKGROUND: Ranibizumab addition may benefit to improve the efficacy in patients with diabetic retinopathy than only photocoagulation, and this meta-analysis aims to explore the impact of ranibizumab addition on efficacy for diabetic retinopathy. METHODS: PubMed, EMbase, Web of science, EBSCO, and Cochrane library databases were systematically searched, and we included randomized controlled trials assessing the effect of ranibizumab addition on patients with diabetic retinopathy for this meta-analysis. RESULTS: Six randomized controlled trials were finally included in the meta-analysis. Overall, compared with control intervention for diabetic retinopathy, ranibizumab addition showed significantly increased number of neovascularization area reduction (OR = 4.20; 95% CI = 1.47-12.02; P = .007) and reduced fluorescein leakage (MD = -2.53; 95% CI = -3.31 to -1.75; P < .00001), but showed no obvious impact on neovascularization area (MD = -1.80; 95% CI = -3.68 to 0.08; P = .06), photocoagulation retreatment (OR = 1.03; 95% CI = 0.47-2.27; P = .94) or adverse events (OR = 1.45; 95% CI = 0.49-4.29; P = .50). CONCLUSIONS: Ranibizumab combined with photocoagulation is effective to improve efficacy for diabetic retinopathy.
Assuntos
Diabetes Mellitus , Retinopatia Diabética , Humanos , Ranibizumab/uso terapêutico , Retinopatia Diabética/complicações , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/cirurgia , Inibidores da Angiogênese/uso terapêutico , Bevacizumab/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/uso terapêutico , Fotocoagulação a Laser , Injeções Intravítreas , Diabetes Mellitus/tratamento farmacológicoRESUMO
AIM: The association of bone turnover with the incidence and progression of nonalcoholic fatty liver disease (NAFLD) is unclear. We aimed to evaluate serum levels of bone turnover markers in relation to NAFLD and nonalcoholic hepatic steatohepatitis (NASH). METHODS: Two cohorts were involved in our study. For the first cohort, 370 participants without NAFLD were retrospectively recruited and followed up for incident NAFLD according to ultrasound. For the second cohort, 562 subjects who underwent liver biopsy were included and grouped into non-NAFLD, non-NASH or NASH according to the NASH Clinical Research Network system. The bone turnover markers osteocalcin, C-terminal telopeptide (CTX) and N-terminal propeptide of type-1 procollagen (P1NP) were measured. RESULTS: Baseline osteocalcin was significantly lower in subjects who developed NAFLD (13.93 [11.03;16.39] versus 18.24 [15.45;22.47] ng/ml, P < 0.001), with a median of 26.4 months of follow-up. Low levels of osteocalcin, but not CTX or P1NP, was an independent predictor of incident NAFLD (OR 0.755 [95%CI 0.668; 0.855] P < 0.001). Moreover, the osteocalcin level was negatively associated with the degree of liver steatosis. Furthermore, subjects with NASH had significantly lower osteocalcin than non-NASH and non-NAFLD group (13.28 [10.49;16.59] versus 14.91 [12.45;18.09] versus 18.21 [15.04;22.05] ng/ml, all P < 0.001). A low osteocalcin level was an independent risk factor for NASH (OR for highest versus lowest quartile: 0.282 [0.147;0.543] P < 0.001). CONCLUSION: Low level of osteocalcin, but not CTX or P1NP, was associated with NAFLD and NASH, indicating its potential role as an important endocrine regulator of hepatic energy metabolism.