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Adipogenesis has been extensively studied using in vitro models of cellular differentiation, enabling long-term regulation of fat cell metabolism in human adipose tissue (AT) material. Many studies promote the idea that manipulation of this process could potentially reduce the prevalence of obesity and its related diseases. It has now become essential to understand the molecular basis of fat cell development to tackle this pandemic disease, by identifying therapeutic targets and new biomarkers. This review explores murine cell models and their applications for study of the adipogenic differentiation process in vitro. We focus on the benefits and limitations of different cell line models to aid in interpreting data and selecting a good cell line model for successful understanding of adipose biology.
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Adipócitos/metabolismo , Adipogenia/genética , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo/crescimento & desenvolvimento , Obesidade/genética , Tecido Adiposo/metabolismo , Tecido Adiposo Marrom/crescimento & desenvolvimento , Animais , Diferenciação Celular/genética , Humanos , Técnicas In Vitro , Camundongos , Obesidade/metabolismo , Obesidade/patologiaRESUMO
The thermal equilibration of himic anhydride [IUPAC (2-endo,3-endo)-bicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid anhydride] to (2-exo,3-exo)-bicyclo[2.2.1]hept-5-ene-2,3-dicarboxylic acid anhydride and subsequent recrystallization of the exo-product can be performed as a standard undergraduate laboratory experiment requiring minimal equipment. The interpretation of the 1H NMR spectra for these norbornene carboxylic anhydride molecules promotes an appreciation of constrained ring systems and factors that affect chemical shifts and coupling constants.
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As more evidence emerges supporting the possibility that nonsteroidal anti-inflammatory drugs, especially aspirin (acetyl salicylic acid), might have a role in the prevention and management of certain types of cancer, there have been several attempts to fabricate salicylic acid-based polymers that can be employed in the targeted therapy of tumors. The primary disadvantage so far has been in use of nontherapeutic polymeric backbones that constitute the majority of the therapeutic particle's size. The focus of this research is the creation of a biodegradable polymer consisting only of salicylic acid, and its use as the main building block in targeted nanotherapeutics that would consequently provide both high local dose and sustained release of the active moiety. This work demonstrates the synthesis and degradation of polysalicylates, and modulation of their size and hydrolytic stability through the formation of nanostructures.
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Nanopartículas/química , Polímeros/química , Ácido Salicílico/química , Materiais Biocompatíveis/química , Humanos , Hidrólise , Tamanho da PartículaRESUMO
DNA base-pairing is the central interaction in DNA assembly. However, this simple four-letter (A-T and G-C) language makes it difficult to create complex structures without using a large number of DNA strands of different sequences. Inspired by protein folding, we introduce hydrophobic interactions to expand the assembly language of DNA nanotechnology. To achieve this, DNA cages of different geometries are combined with sequence-defined polymers containing long alkyl and oligoethylene glycol repeat units. Anisotropic decoration of hydrophobic polymers on one face of the cage leads to hydrophobically driven formation of quantized aggregates of DNA cages, where polymer length determines the cage aggregation number. Hydrophobic chains decorated on both faces of the cage can undergo an intrascaffold "handshake" to generate DNA-micelle cages, which have increased structural stability and assembly cooperativity, and can encapsulate small molecules. The polymer sequence order can control the interaction between hydrophobic blocks, leading to unprecedented "doughnut-shaped" DNA cage-ring structures. We thus demonstrate that new structural and functional modes in DNA nanostructures can emerge from the synergy of two interactions, providing an attractive approach to develop protein-inspired assembly modules in DNA nanotechnology.
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DNA/química , Modelos Moleculares , Nanoestruturas/química , Polímeros/química , Pareamento de Bases , Interações Hidrofóbicas e Hidrofílicas , Micelas , Nanotecnologia , TermodinâmicaRESUMO
The recognition of anions in water remains a key challenge in modern supramolecular chemistry, and is essential if proposed applications in biological, medical, and environmental arenas that typically require aqueous conditions are to be achieved. However, synthetic anion receptors that operate in water have, in general, been the exception rather than the norm to date. Nevertheless, a significant step change towards routinely conducting anion recognition in water has been achieved in the past few years, and this Review highlights these approaches, with particular focus on controlling and using the hydrophobic effect, as well as more exotic interactions such as C-H hydrogen bonding and halogen bonding. We also look beyond the field of small-molecule recognition into the macromolecular domain, covering recent advances in anion recognition based on biomolecules, polymers, and nanoparticles.
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Six cyclometalated iridium(III) phenanthroimidazole complexes with different modifications to the imidazole phenanthroline ligand exhibit enhanced luminescence when bound to guanine (G-) quadruplex DNA sequences. The complexes bind with low micromolar affinity to human telomeric and c-myc sequences in a 1:1 complex:quadruplex stoichiometry. Due to the luminescence enhancement upon binding to G-quadruplex DNA, the complexes can be used as selective quadruplex indicators. In addition, the electrogenerated chemiluminescence of all complexes increases in the presence of specific G-quadruplex sequences, demonstrating potential for the development of an ECL-based G-quadruplex assay.
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Complexos de Coordenação/química , Quadruplex G , Imidazóis/química , Irídio/química , Substâncias Luminescentes/química , Fenantrolinas/química , Sítios de Ligação , Complexos de Coordenação/metabolismo , Guanina/química , Guanina/metabolismo , Imidazóis/metabolismo , Irídio/metabolismo , Ligantes , Luminescência , Substâncias Luminescentes/metabolismo , Medições Luminescentes , Modelos Moleculares , Fenantrolinas/metabolismoRESUMO
Polymer self-assembly and DNA nanotechnology have both proved to be powerful nanoscale techniques. To date, most attempts to merge the fields have been limited to placing linear DNA segments within a polydisperse block copolymer. Here we show that, by using hydrophobic polymers of a precisely predetermined length conjugated to DNA strands, and addressable 3D DNA prisms, we are able to effect the formation of unprecedented monodisperse quantized superstructures. The structure and properties of larger micelles-of-prisms were probed in depth, revealing their ability to participate in controlled release of their constituent nanostructures, and template light-harvesting energy transfer cascades, mediated through both the addressability of DNA and the controlled aggregation of the polymers.
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DNA/química , Nanoestruturas , Polímeros/química , MicelasRESUMO
Inspired by biological polymers, sequence-controlled synthetic polymers are highly promising materials that integrate the robustness of synthetic systems with the information-derived activity of biological counterparts. Polymer-biopolymer conjugates are often targeted to achieve this union; however, their synthesis remains challenging. We report a stepwise solid-phase approach for the generation of completely monodisperse and sequence-defined DNA-polymer conjugates using readily available reagents. These polymeric modifications to DNA display self-assembly and encapsulation behavior-as evidenced by HPLC, dynamic light scattering, and fluorescence studies-which is highly dependent on sequence order. The method is general and has the potential to make DNA-polymer conjugates and sequence-defined polymers widely available.
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Materiais Biocompatíveis/química , DNA/química , Polímeros/química , Técnicas de Síntese em Fase Sólida , Cromatografia Líquida de Alta Pressão , DNA/metabolismo , Microscopia de Força Atômica , Modelos Moleculares , Estrutura Molecular , Polímeros/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Zebrafish have been utilized for many years as a model animal for pharmacological studies on diabetes and obesity. High-fat diet (HFD), streptozotocin and alloxan injection, and glucose immersion have all been used to induce diabetes and obesity in zebrafish. Currently, studies commonly used both male and female zebrafish, which may influence the outcomes since male and female zebrafish are biologically different. This study was designed to investigate the difference between the metabolites of male and female diabetic zebrafish, using limonene - a natural product which has shown several promising results in vitro and in vivo in treating diabetes and obesity-and provide new insights into how endogenous metabolites change following limonene treatment. Using HFD-fed male and female zebrafish, we were able to develop an animal model of T2D and identify several endogenous metabolites that might be used as diagnostic biomarkers for diabetes. The endogenous metabolites in males and females were different, even though both genders had high blood glucose levels and a high BMI. Treatment with limonene prevented high blood glucose levels and improved in diabesity zebrafish by limonene, through reversal of the metabolic changes caused by HFD in both genders. In addition, limonene was able to reverse the elevated expression of AKT during HFD.
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Diabetes Mellitus , Hiperglicemia , Animais , Feminino , Masculino , Hipoglicemiantes/farmacologia , Limoneno , Peixe-Zebra/metabolismo , Glicemia/metabolismo , Espectroscopia de Prótons por Ressonância Magnética , Obesidade/metabolismo , Dieta Hiperlipídica , Hiperglicemia/complicaçõesRESUMO
A series of 14 conjugates of 2α,3ß,23-triacetyl-madecassic acid and silybin were designed and synthesized. The madecassic acid unit was linked to silybin either directly at position C-7 or C-3; or through an amino acid linker (glycine, ß-alanine, or 11-aminoundecanoic acid) at position C-3. The conjugates were tested in vitro for their cytotoxic effect on HepG2 cells using the MTT assay. The results confirmed that the conjugated compounds demonstrated a stronger cytotoxic effect compared to the parent compounds. Of these compounds, the most promising conjugate, compound 8, was evaluated for cytotoxic activity in the additional Hep3B, Huh7, and Huh7R human hepatocellular carcinoma cell lines and also for cell cycle changes and induction of apoptosis in HepG2 cells. This compound caused a rapid and significant induction of caspase 3 activity and induced cell cycle arrest in the S phase - effects distinct from the activity of madecassic acid. This is the first study on the synthesis and cytotoxicity of madecassic acid-silybin conjugates, and of their testing against liver cancer cell lines and provides evidence for a distinct biological profile versus madecassic acid alone.
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The chemistry of aptamers is largely limited to natural nucleotides, and although modifications of nucleic acids can enhance target aptamer affinity, there has not yet been a technology for selecting the right modifications in the right locations out of the vast number of possibilities, because enzymatic amplification does not transmit sequence-specific modification information. Here we show the first method for the selection of specific nucleoside modifications that increase aptamer binding efficacy, using the oncoprotein EGFR as a model target. Using fluorescence-activated bead sorting (FABS), we have successfully selected optimized aptamers from a library of >65 000 variations. Hits were identified by tandem mass spectrometry and validated by using an EGFR binding assay and computational docking studies. Our results provide proof of concept for this novel strategy for the selection of chemically optimised aptamers and offer a new method for rapidly synthesising and screening large aptamer libraries to accelerate diagnostic and drug discovery.
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In this study, the anti-obesity effects of 5,7,3',4',5-pentamethoxyflavone (PMF) and 6,2',4'-trimethoxyflavone (TMF) were evaluated through two distinct mechanisms of action: inhibition of crude porcine pancreatic lipase (PL), and inhibition of adipogenesis in 3T3-L1 pre-adipocytes. Both flavones show dose dependent, competitive inhibition of PL activity. Molecular docking studies revealed binding of the flavones to the active site of PL. In 3T3-L1 adipocytes, both flavones reduced the accumulation of lipids and triglycerides. PMF and TMF also lowered the expression of adipogenic and lipogenic genes. They both reduced the expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ), CCAAT/enhancer-binding protein α and ß (C/EBP α and ß), sterol regulatory element-binding protein 1 (SREBF 1), fatty acid synthase (FASN), adipocyte binding protein 2 (aP2), and leptin gene. In addition, these flavones enhanced adiponectin mRNA expression, increased lipolysis and enhanced the expression of lipolytic genes: adipose triglycerides lipase (ATGL), hormone sensitive lipase (HSL) and monoglycerides lipase (MAGL) in mature 3T3-L1 adipocytes. Overall, PMF was seen to be a more potent inhibitor of both PL activity and adipogenesis versus TMF. These results suggest that PMF and TMF possess anti-obesity activities and can be further evaluated for their anti-obesity effects.
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Adipogenia , Flavonas , Camundongos , Suínos , Animais , Lipase/metabolismo , Simulação de Acoplamento Molecular , Células 3T3-L1 , Proteína alfa Estimuladora de Ligação a CCAAT/genética , PPAR gama/genética , PPAR gama/metabolismo , Flavonas/farmacologia , Triglicerídeos/metabolismo , Obesidade , Diferenciação CelularRESUMO
Hydroxylated polymethoxyflavones (HPMFs) have been shown to possess various anti-disease effects, including against obesity. This study investigates the anti-obesity effects of HPMFs in further detail, aiming to gain understanding of their mechanism of action in this context. The current study demonstrates that two HPMFs; 3'-hydroxy-5,7,4',5'-tetramethoxyflavone (3'OH-TetMF) and 4'-hydroxy-5,7,3',5'-tetramethoxyflavone (4'OH-TetMF) possess anti-obesity effects. They both significantly reduced pancreatic lipase activity in a competitive manner as demonstrated by molecular docking and kinetic studies. In cell studies, it was revealed that both of the HPMFs suppress differentiation of 3T3-L1 mouse embryonic fibroblast cells during the early stages of adipogenesis. They also reduced expression of key adipogenic and lipogenic marker genes, namely peroxisome proliferator-activated receptor-gamma (PPARγ), CCAAT/enhancer-binding protein α and ß (C/EBP α and ß), adipocyte binding protein 2 (aP2), fatty acid synthase (FASN), and sterol regulatory element-binding protein 1 (SREBF 1). They also enhanced the expression of cell cycle genes, i.e., cyclin D1 (CCND1) and C-Myc, and reduced cyclin A2 expression. When further investigated, it was also observed that these HPMFs accelerate lipid breakdown (lipolysis) and enhance lipolytic genes expression. Moreover, they also reduced the secretion of proteins (adipokines), including pro-inflammatory cytokines, from mature adipocytes. Taken together, this study concludes that these HPMFs have anti-obesity effects, which are worthy of further investigation.
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Adipogenia , Lipólise , Animais , Camundongos , Lipase/metabolismo , Lipase/farmacologia , Células 3T3-L1 , Cinética , Simulação de Acoplamento Molecular , Fibroblastos/metabolismo , Diferenciação Celular , Obesidade/metabolismo , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , PPAR gama/genética , PPAR gama/metabolismoRESUMO
Obesity is a major health crisis affecting over a third of the global population. This multifactorial disease is regulated via interoceptive neural circuits in the brain, whose alteration results in excessive body weight. Certain central neuronal populations in the brain are recognised as crucial nodes in energy homeostasis; in particular, the hypothalamic arcuate nucleus (ARC) region contains two peptide microcircuits that control energy balance with antagonistic functions: agouti-related peptide/neuropeptide-Y (AgRP/NPY) signals hunger and stimulates food intake; and pro-opiomelanocortin (POMC) signals satiety and reduces food intake. These neuronal peptides levels react to energy status and integrate signals from peripheral ghrelin, leptin, and insulin to regulate feeding and energy expenditure. To manage obesity comprehensively, it is crucial to understand cellular and molecular mechanisms of information processing in ARC neurons, since these regulate energy homeostasis. Importantly, a specific strategy focusing on ARC circuits needs to be devised to assist in treating obese patients and maintaining weight loss with minimal or no side effects. The aim of this review is to elucidate the recent developments in the study of AgRP-, NPY- and POMC-producing neurons, specific to their role in controlling metabolism. The impact of ghrelin, leptin, and insulin signalling via action of these neurons is also surveyed, since they also impact energy balance through this route. Lastly, we present key proteins, targeted genes, compounds, drugs, and therapies that actively work via these neurons and could potentially be used as therapeutic targets for treating obesity conditions.
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Pró-OpiomelanocortinaRESUMO
We have created sequenced phosphoester-linked trimers of aromatic donor/acceptors which participate in charge-transfer interactions. Each sequence displays characteristic self-assembly, and complementary sequences interact with each other to produce new nanostructures and thermochromism. This paves the way towards new functional nanomaterials which make bio-analogous use of sequence to tune structure.
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Nanoestruturas , Nanoestruturas/químicaRESUMO
The presence of amyloid fibrils is a hallmark of more than 50 human disorders, including neurodegenerative diseases and systemic amyloidoses. A key unresolved challenge in understanding the involvement of amyloid in disease is to explain the relationship between individual structural polymorphs of amyloid fibrils, in potentially mixed populations, and the specific pathologies with which they are associated. Although cryo-electron microscopy (cryo-EM) and solid-state nuclear magnetic resonance (ssNMR) spectroscopy methods have been successfully employed in recent years to determine the structures of amyloid fibrils with high resolution detail, they rely on ensemble averaging of fibril structures in the entire sample or significant subpopulations. Here, we report a method for structural identification of individual fibril structures imaged by atomic force microscopy (AFM) by integration of high-resolution maps of amyloid fibrils determined by cryo-EM in comparative AFM image analysis. This approach was demonstrated using the hitherto structurally unresolved amyloid fibrils formed in vitro from a fragment of tau (297-391), termed 'dGAE'. Our approach established unequivocally that dGAE amyloid fibrils bear no structural relationship to heparin-induced tau fibrils formed in vitro. Furthermore, our comparative analysis resulted in the prediction that dGAE fibrils are structurally closely related to the paired helical filaments (PHFs) isolated from Alzheimer's disease (AD) brain tissue characterised by cryo-EM. These results show the utility of individual particle structural analysis using AFM, provide a workflow of how cryo-EM data can be incorporated into AFM image analysis and facilitate an integrated structural analysis of amyloid polymorphism.
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Doença de Alzheimer , Amiloide , Amiloidose , Doença de Alzheimer/patologia , Amiloide/química , Proteínas Amiloidogênicas/química , Amiloidose/patologia , Microscopia Crioeletrônica/métodos , Humanos , Microscopia de Força Atômica , Estrutura Secundária de ProteínaRESUMO
DNA-peptide conjugates offer an opportunity to marry the benefits of both biomolecular classes, combining the high level of programmability found with DNA, with the chemical diversity of peptides. These hybrid systems offer potential in fields such as therapeutics, nanotechnology, and robotics. Using the first DNA-ß-turn peptide conjugate, we present three studies investigating the self-assembly of DNA-peptide conjugates over a period of 28 days. Time-course studies, such as these have not been previously conducted for DNA-peptide conjugates, although they are common in pure peptide assembly, for example in amyloid research. By using aging studies to assess the structures produced, we gain insights into the dynamic nature of these systems. The first study explores the influence varying amounts of DNA-peptide conjugates have on the self-assembly of our parent peptide. Study 2 explores how DNA and peptide can work together to change the structures observed during aging. Study 3 investigates the presence of orthogonality within our system by switching the DNA and peptide control on and off independently. These results show that two orthogonal self-assemblies can be combined and operated independently or in tandem within a single macromolecule, with both spatial and temporal effects upon the resultant nanostructures.
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The chloride anion templated synthesis of an electrochemical anion sensory interlocked host system, prepared by the integration of redox-active 1,2,3,4,5-pentaphenylferrocene stopper groups into the structure of a rotaxane capable of binding anionic guests is described. Extensive (1)H NMR and electrochemical titration investigations were used to probe the anion recognition and sensing properties of the rotaxane, compared to the axle and model system components. A characteristic electrochemical response was observed for chloride binding by the rotaxane, which was attributed to the topologically constrained cavity of the interlocked host molecule.
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Ânions/química , Cloretos/química , Compostos Ferrosos/química , Rotaxanos/química , Cristalografia por Raios X , Eletroquímica , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Estrutura MolecularRESUMO
The interaction between imidazolium cations and coordinating anions is investigated through the anion-templated assembly of interpenetrated and interlocked structures. The orientation of the imidazolium motif with respect to anion binding, and hence the hydrogen bond donor arrangement, was varied in acyclic receptors, interpenetrated assemblies, and the first mono-imidazolium interlocked systems. Their anion recognition properties and co-conformations were studied by solution-phase 1Hâ NMR investigations, solid-state structures, molecular dynamics simulations, and density functional theory calculations. Our findings suggest that the imidazolium-anion binding interaction is dominated by electrostatics with hydrogen-bonding contributions having weak orientational dependence.