RESUMO
AIMS: Lung cancer is the leading cause of cancer-related death. Unfortunately, targeted-therapies have been unsuccessful for most patients with lung adenocarcinoma (LUAD). Thus, new early biomarkers and treatment options are a pressing need. Fatty acid binding protein 5 (FABP5) has been associated with various types of cancers. Its contribution to LUAD onset, progression and metabolic reprogramming is, however, not fully understood. In this study we assessed the importance of FABP5 in LUAD and its role in cancer lipid metabolism. MAIN METHODS: By radioactive labeling and metabolite quantification, we studied the function of FABP5 in fatty acid metabolism using genetic/pharmacologic inhibition and overexpression models in LUAD cell lines. Flow cytometry, heterologous transplantation and bioinformatic analysis were used, in combination with other methodologies, to assess the importance of FABP5 for cellular proliferation in vitro and in vivo and in patient survival. KEY FINDINGS: We show that high expression of FABP5 is associated with poor prognosis in patients with LUAD. FABP5 regulates lipid metabolism, diverting fatty acids towards complex lipid synthesis, whereas it does not affect their catabolism in vitro. Moreover, FABP5 is required for de novo fatty acid synthesis and regulates the expression of enzymes involved in the pathway (including FASN and SCD1). Consistently with the changes in lipid metabolism, FABP5 is required for cell cycle progression, migration and in vivo tumor growth. SIGNIFICANCE: Our results suggest that FABP5 is a regulatory hub of lipid metabolism and tumor progression in LUAD, placing it as a new putative therapeutic target for this disease.
Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Linhagem Celular Tumoral , Proliferação de Células , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Ácidos Graxos/metabolismo , Humanos , LipogêneseRESUMO
Due to their phylogenetic proximity to humans, nonhuman primates (NHPs) are considered an adequate choice for a basic and preclinical model of sepsis. Gram-negative bacteria are the primary causative of sepsis. During infection, bacteria continuously release the potent toxin lipopolysaccharide (LPS) into the bloodstream, which triggers an uncontrolled systemic inflammatory response leading to death. Our previous research has demonstrated in vitro and in vivo using a mouse model of septic shock that Fh15, a recombinant variant of the Fasciola hepatica fatty acid binding protein, acts as an antagonist of Toll-like receptor 4 (TLR4) suppressing the LPS-induced proinflammatory cytokine storm. The present communication is a proof-of concept study aimed to demonstrate that a low-dose of Fh15 suppresses the cytokine storm and other inflammatory markers during the early phase of sepsis induced in rhesus macaques by intravenous (i.v.) infusion with lethal doses of live Escherichia coli. Fh15 was administered as an isotonic infusion 30 min prior to the bacterial infusion. Among the novel findings reported in this communication, Fh15 (i) significantly prevented bacteremia, suppressed LPS levels in plasma, and the production of C-reactive protein and procalcitonin, which are key signatures of inflammation and bacterial infection, respectively; (ii) reduced the production of proinflammatory cytokines; and (iii) increased innate immune cell populations in blood, which suggests a role in promoting a prolonged steady state in rhesus macaques even in the presence of inflammatory stimuli. This report is the first to demonstrate that a F. hepatica-derived molecule possesses potential as an anti-inflammatory drug against sepsis in an NHP model. IMPORTANCE Sepsis caused by Gram-negative bacteria affects 1.7 million adults annually in the United States and is one of the most important causes of death at intensive care units. Although the effective use of antibiotics has resulted in improved prognosis of sepsis, the pathological and deathly effects have been attributed to the persistent inflammatory cascade. There is a present need to develop anti-inflammatory agents that can suppress or neutralize the inflammatory responses and prevent the lethal consequences of sepsis. We demonstrated here that a small molecule of 14.5 kDa can suppress the bacteremia, endotoxemia, and many other inflammatory markers in an acute Gram-negative sepsis rhesus macaque model. These results reinforce the notion that Fh15 constitutes an excellent candidate for drug development against sepsis.
Assuntos
Anti-Inflamatórios/administração & dosagem , Bacteriemia/tratamento farmacológico , Fasciola hepatica/metabolismo , Proteínas de Ligação a Ácido Graxo/administração & dosagem , Bactérias Gram-Negativas/fisiologia , Proteínas de Helminto/administração & dosagem , Animais , Anti-Inflamatórios/metabolismo , Bacteriemia/genética , Bacteriemia/imunologia , Bacteriemia/microbiologia , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Fasciola hepatica/química , Fasciola hepatica/genética , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Bactérias Gram-Negativas/classificação , Bactérias Gram-Negativas/genética , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , Humanos , Macaca mulatta , Masculino , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Receptor 4 Toll-Like/antagonistas & inibidores , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologiaRESUMO
Fatty Acid Binding-Protein 5 (FABP5) is a cytoplasmic protein, which binds long-chain fatty acids and other hydrophobic ligands. This protein is implicated in several physiological processes including mitochondrial ß-oxidation and transport of fatty acids, membrane phospholipid synthesis, lipid metabolism, inflammation and pain. In the present study, we used molecular docking tools to determine the possible interaction of FABP5 with six selected compounds retrieved form Drugbank. Our results showed that FABP5 binding pocket included 31 polar and non-polar amino acids, and these residues may be related to phosphorylation, acetylation, ubiquitylation, and mono-methylation. Docking results showed that the most energetically favorable compounds are NADH (-9.12 kcal/mol), 5'-O-({[(Phosphonatooxy)phosphinato]oxy}phosphinato)adenosine (-8.62 kcal/mol), lutein (-8.25 kcal/mol), (2S)-2-[(4-{[(2-Amino-4-oxo-1,4,5,6,7,8-hexahydro-6-pteridinyl)methyl]amino}benzoyl)amino]pentanedioate (-7.17 kcal/mol), Pteroyl-L-glutamate (-6.86 kcal/mol) and (1S,3R,5E,7Z)-9,10-Secocholesta-5,7,10-triene-1,3,25-triol (-6.79 kcal/mol). Common interacting residues of FABP5 with nutraceuticals included SER16, LYS24, LYS34, LYS40 and LYS17. Further, we used the SwissADME server to determine the physicochemical and pharmacokinetic characteristics and to predict the ADME parameters of the selected nutraceuticals after molecular analysis by docking with the FABP5 protein. Amongst all compounds, pteroyl-L-glutamate is the only one meeting the Lipinski's rule of five criteria, demonstrating its potential pharmacological use. Finally, our results also suggest the importance of FABP5 in mediating the anti-inflammatory activity of the nutraceutical compounds.
Assuntos
Anti-Inflamatórios , Proteínas de Ligação a Ácido Graxo , Suplementos Nutricionais , Proteínas de Ligação a Ácido Graxo/genética , Ligantes , Simulação de Acoplamento MolecularRESUMO
Pharmaceuticals and their metabolites constitute a class of xenobiotics commonly found in aquatic environments which may cause toxic effects in aquatic organisms. Several different lipophilic molecules, including some pharmaceuticals, can bind to fatty acid-binding proteins (FABPs), a group of evolutionarily related cytoplasmic proteins that belong to the intracellular lipid-binding protein (iLBP) family. An oyster FABP genome-wide investigation was not available until a recent study on gene organization, protein structure, and phylogeny of Crassostrea gigas iLBPs. Higher transcript levels of the C. gigas FABP2 gene were found after exposure to sewage and pharmaceuticals. Because of its relevance as a potential biomarker of aquatic contamination, in this study, recombinant FABP2 from C. gigas (CgFABP2) was successfully cloned, expressed, and purified, and in vitro and in silico assays were performed using lipids and pharmaceuticals. This is the first characterization of a protein from the iLBP family in C. gigas. Homology modeling and molecular docking were used to evaluate the binding affinities of natural ligands (palmitic, oleic, and arachidonic acids) and pharmaceuticals (ibuprofen, sodium diclofenac, and acetaminophen). Among the tested fatty acids, CgFABP2 showed preference for palmitic acid. The selected pharmaceuticals presented a biphasic-binding mode, suggesting a different binding affinity with a preference for diclofenac. Therefore, the approach using circular dichroism and in silico data might be useful for ligand-binding screening in an invertebrate model organism.
Assuntos
Crassostrea , Preparações Farmacêuticas , Animais , Crassostrea/genética , Proteínas de Ligação a Ácido Graxo/genética , Simulação de Acoplamento Molecular , FilogeniaRESUMO
BACKGROUND: Acute myeloid leukemia (AML) is the most common type of acute leukemia and biologically heterogeneous diseases with poor prognosis. Thus, we aimed to identify prognostic markers to effectively predict the prognosis of AML patients and eventually guide treatment. METHODS: Prognosis-associated genes were determined by Kaplan-Meier and multivariate analyses using the expression and clinical data of 173 AML patients from The Cancer Genome Atlas database and validated in an independent Oregon Health and Science University dataset. A prognostic risk score was computed based on a linear combination of 5-gene expression levels using the regression coefficients derived from the multivariate logistic regression model. The classification of AML was established by unsupervised hierarchical clustering of CALCRL, DOCK1, PLA2G4A, FCHO2 and LRCH4 expression levels. RESULTS: High FCHO2 and LRCH4 expression was related to decreased mortality. While high CALCRL, DOCK1, PLA2G4A expression was associated with increased mortality. The risk score was predictive of increased mortality rate in AML patients. Hierarchical clustering analysis of the five genes discovered three clusters of AML patients. The cluster1 AML patients were associated with lower cytogenetics risk than cluster2 or 3 patients, and better prognosis than cluster3 patients (P values < 0.05 for all cases, fisher exact test or log-rank test). CONCLUSION: The gene panel comprising CALCRL, DOCK1, PLA2G4A, FCHO2 and LRCH4 as well as the risk score may offer novel prognostic biomarkers and classification of AML patients to significantly improve outcome prediction.
Assuntos
Proteína Semelhante a Receptor de Calcitonina/genética , Proteínas de Ligação a Ácido Graxo/genética , Expressão Gênica , Fosfolipases A2 do Grupo IV/genética , Leucemia Mieloide Aguda/genética , Proteínas de Membrana/genética , Proteínas do Tecido Nervoso/genética , Proteínas rac de Ligação ao GTP/genética , Bases de Dados Genéticas , Perfilação da Expressão Gênica , Humanos , Estimativa de Kaplan-Meier , Leucemia Mieloide Aguda/mortalidade , Análise Multivariada , Prognóstico , Fatores de Risco , Resultado do TratamentoRESUMO
The intestinal fatty acid binding protein (FABP) is a small protein expressed along the small intestine that bind long-chain fatty acids and other hydrophobic ligands. Several lines of evidence suggest that, once in the nucleus, it interacts with nuclear receptors, activating them and thus transferring the bound ligand into the nucleus. Previous work by our group suggests that FABP2 would participate in the cytoplasm-nucleus translocation of fatty acids. Because the consensus NLS is absent in the sequence of FABP2, we propose that a 3D signal could be responsible for its nuclear translocation. The results obtained by transfection assays of recombinant wild type and mutated forms of Danio rerio Fabp2 in Caco-2 cell cultures, showed that lysine 17, arginine 29 and lysine 30 residues, which are located in the helix-turn-helix region, would constitute a functional non-classical three-dimensional NLS.
Assuntos
Núcleo Celular/metabolismo , Proteínas de Ligação a Ácido Graxo/metabolismo , Sequência de Aminoácidos , Animais , Células CACO-2 , Proteínas de Ligação a Ácido Graxo/química , Proteínas de Ligação a Ácido Graxo/genética , Ácidos Graxos/metabolismo , Humanos , Microscopia Confocal , Mutagênese , Sinais de Localização Nuclear/química , Ligação Proteica , Conformação Proteica em alfa-Hélice , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/química , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
Obesity is an independent risk factor for the development of chronic kidney disease. The pathophysiology of the obesity-induced kidney injury is complex, but evidence suggests the involvement of reduced adiponectin levels and signaling. We investigated the extent by which adiponectin contributes to the establishment and progression of renal disease in wild type (WT) and adiponectin null (adipoKO) mice fed a control or a high-fat diet (HFD) for 16 weeks. HFD induced obesity, kidney hypertrophy, albuminuria, renal lipid accumulation and decreased nephrin expression in both mice genotypes. Notably, HFD in adipoKO mice exacerbated progression of albuminuria in comparison to WT mice. In addition, lack of adiponectin per se increased kidney weight, reduced nephrin levels, up-regulated Fabp4 expression, reduced Cpt1a expression and increased miR-130 levels in kidney. Our results demonstrate that lack of adiponectin combined with a HFD contributes to accelerated kidney dysfunction.
Assuntos
Adiponectina/genética , Albuminúria/fisiopatologia , Dieta Hiperlipídica/efeitos adversos , Obesidade/complicações , Insuficiência Renal Crônica/fisiopatologia , Albuminúria/genética , Animais , Carnitina O-Palmitoiltransferase/genética , Modelos Animais de Doenças , Progressão da Doença , Proteínas de Ligação a Ácido Graxo/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/genética , Insuficiência Renal Crônica/genéticaRESUMO
INTRODUCTION AND OBJECTIVES: Non-alcoholic fatty liver disease (NAFLD) is multistage with heterogeneous outcomes. We studied the influence of insulin resistance (IR) on the hepatic transcriptome of early NAFLD stages, to understand disease development. MATERIALS AND METHODS: In this cross-sectional study, possible clinicopathological risk factors were compared between mild-NAFL (Nâ¯=â¯72) and non-alcoholic steatohepatitis (NASH; Nâ¯=â¯51) patients. Liver tissue-transcriptome difference was studied between a subset of 25 mild-NAFL and 20 NASH biopsies and validated on another subset of 12 mild-NAFL and 13 NASH biopsies, using RT-PCR. The relationship between IR driven gene expression changes with fibrosis in NASH was investigated. RESULTS: Significantly higher weight (pâ¯=â¯0.005) and elevated levels of HbA1c (pâ¯=â¯0.009), FBG (pâ¯=â¯0.03) and HOMA-IR (pâ¯=â¯0.009) were found in NASH patients. Five differentially expressed genes (DEGs, fold changeâ¯>â¯1.5) were identified in NASH-FABP4, FABP5L2, CD24, PRAP1, and SPP1. The DEGs were positively associated with disease severity and HOMA-IR, and were found to be efficient classifiers of mild-NAFL and NASH. Additional 1218 genes identified related to IR (IrCGs), which can classify NASH-with-fibrosis patients separately from mild-NAFL and NASH patients. IrCGs can promote intra-hepatic fat accumulation, dysregulation of the lipid metabolism, lipotoxicity, and activation of cell survival pathways including activation of cell proliferation and differentiation pathways. CONCLUSIONS: Hepatic expression of genes associated with insulin resistance may drive NAFLD development and progression.
Assuntos
Perfilação da Expressão Gênica , Resistência à Insulina/genética , Cirrose Hepática/genética , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/genética , Transcriptoma , Adulto , Glicemia/metabolismo , Antígeno CD24/genética , Antígeno CD24/metabolismo , Estudos Transversais , Progressão da Doença , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Feminino , Hemoglobinas Glicadas/metabolismo , Humanos , Insulina/sangue , Fígado/patologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Osteopontina/genética , Osteopontina/metabolismo , Proteínas da Gravidez/genética , Proteínas da Gravidez/metabolismo , Índice de Gravidade de DoençaRESUMO
Fatty acid-binding proteins (FABPs) are small intracellular proteins that reversibly bind fatty acids and other hydrophobic ligands. In cestodes, due to their inability to synthesise fatty acids de novo, FABPs have been proposed as essential proteins, and thus, as possible drug targets and/or carriers against these parasites. We performed data mining in Echinococcus multilocularis and Echinococcus granulosus genomes in order to test whether this family of proteins is more complex than previously reported. By exploring the genomes of E. multilocularis and E. granulosus, six genes coding for FABPs were found in each organism. In the case of E. granulosus, all of them have different coding sequences, whereas in E. multilocularis, two of the genes code for the same protein. Remarkably, one of the genes (in both cestodes) encodes a FABP with a C-terminal extension unusual for this family of proteins. The newly described genes present variations in their structure in comparison with previously described FABP genes in Echinococcus spp. The coding sequences for E. multilocularis were validated by cloning and sequencing. Moreover, differential expression patterns of FABPs were observed at different stages of the life cycle of E. multilocularis by exploring transcriptomic data from several sources. In summary, FABP family in cestodes is far more complex than previously thought and includes new members that seem to be only present in flatworms.
Assuntos
Echinococcus granulosus/genética , Echinococcus multilocularis/genética , Proteínas de Ligação a Ácido Graxo/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA de Protozoário/genética , Ácidos Graxos/metabolismo , Genoma de Protozoário/genética , Análise de Sequência , Análise de Sequência de DNA , Transcriptoma/genéticaRESUMO
Nitro-fatty acids (NO2-FA) are electrophilic lipid mediators derived from unsaturated fatty acid nitration. These species are produced endogenously by metabolic and inflammatory reactions and mediate anti-oxidative and anti-inflammatory responses. NO2-FA have been postulated as partial agonists of the Peroxisome Proliferator-Activated Receptor gamma (PPARγ), which is predominantly expressed in adipocytes and myeloid cells. Herein, we explored molecular and cellular events associated with PPARγ activation by NO2-FA in monocytes and macrophages. NO2-FA induced the expression of two PPARγ reporter genes, Fatty Acid Binding Protein 4 (FABP4) and the scavenger receptor CD36, at early stages of monocyte differentiation into macrophages. These responses were inhibited by the specific PPARγ inhibitor GW9662. Attenuated NO2-FA effects on PPARγ signaling were observed once cells were differentiated into macrophages, with a significant but lower FABP4 upregulation, and no induction of CD36. Using in vitro and in silico approaches, we demonstrated that NO2-FA bind to FABP4. Furthermore, the inhibition of monocyte FA binding by FABP4 diminished NO2-FA-induced upregulation of reporter genes that are transcriptionally regulated by PPARγ, Keap1/Nrf2 and HSF1, indicating that FABP4 inhibition mitigates NO2-FA signaling actions. Overall, our results affirm that NO2-FA activate PPARγ in monocytes and upregulate FABP4 expression, thus promoting a positive amplification loop for the downstream signaling actions of this mediator.