RESUMEN
The asialoglycoprotein receptor (ASGPR) is expressed in high density on hepatocytes. Multivalent variants of galactosyl carbohydrates bind ASGPR with high affinity, enabling hepatic delivery of ligand-bound cargo. Virus-like particle (VLP) conjugates of a relatively high-affinity ligand were efficiently endocytosed by ASGPR-expressing cells in a manner strongly dependent on the nature and density of ligand display, with the best formulation using a nanomolar-, but not a picomolar-level, binder. Optimized particles were taken up by HepG2 cells with greater efficiency than competing small molecules or the natural multigalactosylated ligand, asialoorosomucoid. Upon systemic injection in mice, these VLPs were rapidly cleared to the liver and were found in association with sinusoidal endothelial cells, Kupffer cells, hepatocytes, dendritic cells, and other immune cells. Both ASGPR-targeted and nontargeted particles were distributed similarly to endothelial and Kupffer cells, but targeted particles were distributed to a greater number and fraction of hepatocytes. Thus, selective cellular trafficking in the liver is difficult to achieve: even with the most potent ASGPR targeting available, barrier cells take up much of the injected particles and hepatocytes are accessed only approximately twice as efficiently in the best case.
Asunto(s)
Células Endoteliales , Hígado , Animales , Ratones , Receptor de Asialoglicoproteína , Ligandos , Células Endoteliales/metabolismo , Hígado/metabolismo , Hepatocitos/metabolismoRESUMEN
The conformational study of saccharides and glycomimetics in solution is critical for a comprehensive understanding of their interactions with biological receptors and enabling the design of optimized glycomimetics. Here, we report a nuclear magnetic resonance (NMR) study centered on the conformational properties of the hydroxymethyl group and glycosidic bond of four series of aryl S-glucosides. We found that in acetyl-protected and free aryl S-ß-glucosides, the rotational equilibrium around the C5-C6 bond (hydroxymethyl group) exhibits a linear dependence on the electronic properties of the aglycone, namely, as the aryl's substituent electron-withdrawing character increases, the dominance of the gg rotamer declines and the gt contribution rises. Likewise, the conformational equilibrium around the glycosidic C1-S bond also depends on the aglycone's electronic properties, where glucosides carrying electron-poor aglycones exhibit stiffer glycosidic bonds in comparison to their electron-rich counterparts. In the case of the α anomers, the aglycone's effect over the glycosidic bond conformation is like that observed on their ß isomers; however, we observe no aglycone's influence over the hydroxymethyl group conformation in the α-glucosides.
Asunto(s)
Glucósidos , Glicósidos , Conformación Molecular , Glicósidos/químicaRESUMEN
A voltammetric study of a series of alkyl and aryl S-glucosides unveiled the reactivity patterns of alkyl S-glucosides toward anodic oxidation and found noteworthy differences with the trends followed by aryl derivatives. The oxidation potential of alkyl S-glucosides, estimated herein from square-wave voltammetry peak potentials (Ep), depends on the steric properties of the aglycone. Glucosides substituted with bulky groups exhibit Ep values at voltages more positive than the values of those carrying small aglycones. This relationship, observed in all analyzed alkyl series, is evidenced by good linear correlations between Ep and Taft's steric parameters (ES) of the respective alkyl substituents. Moreover, the role of the aglycone's steric properties as a primary reactivity modulator is backed by poor correlations between Ep and the radical stabilization energies (RSEs) of the aglycone-derived thiyl radicals (RSâ¢). In contrast, aryl glucosides' Ep values exhibit excellent correlations with the aryl substituents' Hammett parameters (σ+) and the ArS⢠RSEs, evidencing the inherent stability of the reactive radical intermediate as the primary factor controlling aryl glucoside's electrochemical reactivity. The reactivity differences between alkyl and aryl S-glucosides also extend to the protective group's effect on Ep. Alkyl S-glucosides' reactivity proved to be more sensitive to protective group exchange.
Asunto(s)
Glucósidos , Electrodos , Oxidación-ReducciónRESUMEN
A compact and stable bicyclic bridged ketal was developed as a ligand for the asialoglycoprotein receptor (ASGPR). This compound showed excellent ligand efficiency, and the molecular details of binding were revealed by the first X-ray crystal structures of ligand-bound ASGPR. This analogue was used to make potent di- and trivalent binders of ASGPR. Extensive characterization of the function of these compounds showed rapid ASGPR-dependent cellular uptake in vitro and high levels of liver/plasma selectivity in vivo. Assessment of the biodistribution in rodents of a prototypical Alexa647-labeled trivalent conjugate showed selective hepatocyte targeting with no detectable distribution in nonparenchymal cells. This molecule also exhibited increased ASGPR-directed hepatocellular uptake and prolonged retention compared to a similar GalNAc derived trimer conjugate. Selective release in the liver of a passively permeable small-molecule cargo was achieved by retro-Diels-Alder cleavage of an oxanorbornadiene linkage, presumably upon encountering intracellular thiol. Therefore, the multicomponent construct described here represents a highly efficient delivery vehicle to hepatocytes.
Asunto(s)
Receptor de Asialoglicoproteína/metabolismo , Compuestos Bicíclicos con Puentes/química , Hepatocitos/metabolismo , Cetonas/química , Hígado/metabolismo , Polímeros/química , Compuestos Bicíclicos con Puentes/metabolismo , Cristalografía por Rayos X , Portadores de Fármacos/química , Humanos , Cetonas/metabolismo , Hígado/citología , Modelos Moleculares , Estructura Molecular , Polímeros/metabolismoRESUMEN
Cancer cells generate protons (H+) that are extruded to the extracellular medium mainly via the Na+/H+ exchanger 1 (NHE1), which regulates intracellular pH (pHi) and cell proliferation. In primary cultures of human ascites-derived ovarian cancer cells (haOC) we assayed whether NHE1 was required for pHi modulation and cell proliferation. Human ovary expresses NHE1, which is higher in haOC and A2780 (ovarian cancer cells) compared with HOSE cells (normal ovarian cells). Basal pHi and pHi recovery (following a NH4Cl pulse) was higher in haOC and A2780, compared with HOSE cells. Zoniporide (NHE1 inhibitor) caused intracellular acidification and pHi recovery was independent of intracellular buffer capacity, but reduced in NHE1 knockdown A2780 cells. Zoniporide reduced the maximal proliferation capacity, cell number, thymidine incorporation, and ki67 (marker of proliferation) fluorescence in haOC cells. SLC9A1 (for NHE1) amplification associated with lower overall patient survival. In conclusion, NHE1 is expressed in human ovarian cancer where it has a pro-proliferative role. Increased NHE1 expression and activity constitute an unfavourable prognostic factor in these patients.
Asunto(s)
Neoplasias Ováricas/metabolismo , Intercambiador 1 de Sodio-Hidrógeno/metabolismo , Proliferación Celular , Células Cultivadas , Femenino , Humanos , Concentración de Iones de Hidrógeno , Neoplasias Ováricas/patología , Ovario/metabolismo , Ovario/patología , Intercambiador 1 de Sodio-Hidrógeno/análisisRESUMEN
Nitric oxide plays several roles in cellular physiology, including control of the vascular tone and defence against pathogen infection. Neuronal, inducible and endothelial nitric oxide synthase (NOS) isoforms synthesize nitric oxide. Cells generate acid and base equivalents, whose physiological intracellular concentrations are kept due to membrane transport systems, including Na+ /H+ exchangers and Na+ /HCO3- transporters, thus maintaining a physiological pH at the intracellular (~7.0) and extracellular (~7.4) medium. In several pathologies, including cancer, cells are exposed to an extracellular acidic microenvironment, and the role for these membrane transport mechanisms in this phenomenon is likely. As altered NOS expression and activity is seen in cancer cells and because this gas promotes a glycolytic phenotype leading to extracellular acidosis in gynaecological cancer cells, a pro-inflammatory microenvironment increasing inducible NOS expression in this cell type is feasible. However, whether abnormal control of intracellular and extracellular pH by cancer cells regards with their ability to synthesize or respond to nitric oxide is unknown. We, here, discuss a potential link between pH alterations, pH controlling membrane transport systems and NOS function. We propose a potential association between inducible NOS induction and Na+ /H+ exchanger expression and activity in human ovary cancer. A potentiation between nitric oxide generation and the maintenance of a low extracellular pH (i.e. acidic) is proposed to establish a sequence of events in ovarian cancer cells, thus preserving a pro-proliferative acidic tumour extracellular microenvironment. We suggest that pharmacological therapeutic targeting of Na+ /H+ exchangers and inducible NOS may have benefits in human epithelial ovarian cancer.
Asunto(s)
Neoplasias de los Genitales Femeninos/metabolismo , Óxido Nítrico/metabolismo , Animales , Membrana Celular/metabolismo , Femenino , Humanos , Concentración de Iones de Hidrógeno , Modelos BiológicosRESUMEN
Reduced adenosine uptake via human equilibrative nucleoside transporter 1 (hENT1) in human umbilical vein endothelial cells (HUVECs) from gestational diabetes mellitus (GDM) is reversed by insulin by restoring hENT1 expression. Insulin receptors A (IR-A) and B (IR-B) are expressed in HUVECs, and GDM results in higher IR-A mRNA expression vs. cells from normal pregnancies. We studied whether the reversal of GDM effects on transport by insulin depends on restoration of IR-A expression. We specifically measured hENT1 expression [mRNA, protein abundance, SLC29A1 (for hENT1) promoter activity] and activity (adenosine transport kinetics) and the role of IR-A/IR-B expression and signaling [total and phosphorylated 42 and 44 kDa mitogen-activated protein kinases (p44/42(mapk)) and Akt] in IR-A, IR-B, and IR-A/B knockdown HUVECs from normal (n = 33) or GDM (n = 33) pregnancies. GDM increases IR-A/IR-B mRNA expression (1.8-fold) and p44/42(mapk):Akt activity (2.7-fold) ratios. Insulin reversed GDM-reduced hENT1 expression and maximal transport capacity (V(max)/K(m)), and GDM-increased IR-A/IR-B mRNA expression and p44/42(mapk):Akt activity ratios to values in normal pregnancies. Insulin's effect was abolished in IR-A or IR-A/B knockdown cells. Thus, insulin requires normal IR-A expression and p44/42(mapk)/Akt signaling to restore GDM-reduced hENT1 expression and activity in HUVECs. This could be a protective mechanism for the placental macrovascular endothelial dysfunction seen in GDM.
Asunto(s)
Adenosina/metabolismo , Antígenos CD/metabolismo , Diabetes Gestacional/metabolismo , Insulina/metabolismo , Receptor de Insulina/metabolismo , Adolescente , Adulto , Antígenos CD/genética , Transporte Biológico Activo , Estudios de Casos y Controles , Diabetes Gestacional/genética , Tranportador Equilibrativo 1 de Nucleósido/genética , Tranportador Equilibrativo 1 de Nucleósido/metabolismo , Femenino , Expresión Génica , Técnicas de Silenciamiento del Gen , Células Endoteliales de la Vena Umbilical Humana , Humanos , Recién Nacido , Cinética , Sistema de Señalización de MAP Quinasas , Masculino , Embarazo , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptor de Insulina/antagonistas & inhibidores , Receptor de Insulina/genética , Transducción de Señal , Venas Umbilicales/metabolismo , Adulto JovenRESUMEN
NEW FINDINGS: What is the topic of this review? This review focuses on the effects of insulin therapy on fetoplacental vasculature in gestational diabetes mellitus and the potentiating effects of adenosine on this therapy. What advances does it highlight? This review highlights recent studies exploring a potential functional link between insulin receptors and their dependence on adenosine receptor activation (insulin-adenosine axis) to restore placental endothelial function in gestational diabetes mellitus. Gestational diabetes mellitus (GDM) is a disease that occurs during pregnancy and is associated with maternal and fetal hyperglycaemia. Women with GDM are treated via diet to control their glycaemia; however, a proportion of these patients do not achieve the recommended values of glycaemia and are subjected to insulin therapy until delivery. Even if a diet-treated GDM pregnancy leads to normal maternal and newborn glucose levels, fetoplacental vascular dysfunction remains evident. Thus, control of glycaemia via diet does not prevent GDM-associated fetoplacental vascular and metabolic alterations. We review the available information regarding insulin therapy in the context of its potential consequences for fetoplacental vascular function in GDM. We propose the possibility that insulin therapy to produce normoglycaemia in the mother and newborn may require additional therapeutic measures to restore the normal metabolic condition of the vascular network in GDM. A role for A1 and A2A adenosine receptors and insulin receptors A and B as well as a potential functional link in the cell signalling associated with the activation of these receptors is proposed. This possibility could be helpful for the planning of strategies, including adenosine receptor-improved insulin therapy, for the treatment of GDM patients, thereby promoting the wellbeing of the growing fetus, newborn and mother.
Asunto(s)
Diabetes Gestacional/tratamiento farmacológico , Diabetes Gestacional/fisiopatología , Insulina/uso terapéutico , Circulación Placentaria/fisiología , Femenino , Humanos , Recién Nacido , Pruebas de Función Placentaria/métodos , EmbarazoRESUMEN
A focused library of virtual heterobifunctional ligands was generated in silico and a set of ligands with recombined fragments was synthesized and evaluated for binding to Clostridium difficile toxins. The position of the trisaccharide fragment was used as a reference for filtering docked poses during virtual screening to match the trisaccharide ligand in a crystal structure. The peptoid, a diversity fragment probing the protein surface area adjacent to a known binding site, was generated by a multi-component Ugi reaction. Our approach combines modular fragment-based design with in silico screening of synthetically feasible compounds and lays the groundwork for future efforts in development of composite bifunctional ligands for large clostridial toxins.
Asunto(s)
Clostridioides difficile , Simulación por Computador , Bibliotecas de Moléculas Pequeñas/metabolismo , Toxinas Biológicas/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Sitios de Unión , Metabolismo de los Hidratos de Carbono , Técnicas Químicas Combinatorias , Cristalografía por Rayos X , Ligandos , Modelos Moleculares , Datos de Secuencia Molecular , Conformación Proteica , Toxinas Biológicas/químicaRESUMEN
Helicobacter pylori is the etiologic agent of a series of gastric pathologies that may culminate in the development of gastric adenocarcinoma. An initial step in this process is the loss of glandular structures in the gastric mucosa, presumably as the consequence of increased apoptosis and reduced cellular regeneration, which may be attributed to the combination of several bacterial and host factors and to an unfavorable proinflammatory environment. In a previous study, we showed that survivin, a member of the inhibitor of apoptosis protein family, is expressed in the normal human gastric mucosa and that its levels decrease in the mucosa of infected patients and in gastric cells exposed in culture to the bacteria, coincident with increased cell death in the latter case. We investigated the bacterial factors responsible for loss of survivin in gastric cells exposed to H. pylori. The results of this study indicated that the loss of survivin due to H. pylori infection involves proteasome-mediated degradation of the protein. Studies with isogenic mutants deficient in either CagA, VacA, lipopolysaccharide, or gamma-glutamyl transpeptidase (GGT) implicated the latter in H. pylori-induced loss of survivin and cell viability. Moreover, experiments with the GGT inhibitor 6-diazo-5-oxo-l-norleucine and purified recombinant GGT protein indicated that secreted bacterial GGT activity was required and sufficient to induce these effects.
Asunto(s)
Infecciones por Helicobacter/microbiología , Infecciones por Helicobacter/patología , Helicobacter pylori/enzimología , Helicobacter pylori/patogenicidad , Proteínas Inhibidoras de la Apoptosis/metabolismo , Factores de Virulencia/metabolismo , gamma-Glutamiltransferasa/metabolismo , Línea Celular , Eliminación de Gen , Humanos , Viabilidad Microbiana , Survivin , Factores de Virulencia/genética , gamma-Glutamiltransferasa/genéticaRESUMEN
PROteolysis Targeting Chimeras (PROTACs) showed tremendous therapeutic potential in degrading several oncoproteins including undruggable proteins. PROTACs are bifunctional molecules where one-part binds to target protein while the other end recruits protein degradation machinery. With the unveiling advancements in the field of PROTACs, we explored a combinatorial approach by developing antibody-based PROTAC (ABTAC) which may effectively degrade one of the key oncoprotein driving proliferation and progression of cancer - Epidermal growth factor receptor (EGFR). The objective of current research was to synthesize and characterize an EGFR degrading ABTAC for the treatment of non-small cell lung cancer (NSCLC). Cetuximab and pomalidomide (E3 ligase recruiting ligand) were conjugated using lysine conjugation and copper free azide-alkyne cycloaddition (CuAAC) click chemistry. Analytical characterization using reverse-phase liquid chromatography and mass spectrometry suggested conjugation of five E3-ligase inhibitor molecules/antibody. Nearly 10-30 folds reduction in IC50 was observed with ABTAC in HCC827 (EGFR sensitive) and H1650 (EGFR resistant) cells compared to cetuximab. Multicellular 3D spheroid assay strongly suggested that ABTAC induced significant apoptosis and also inhibited cell proliferation compared to control and antibody alone. Circular dichroism and surface plasmon resonance (SPR) confirmed minor alterations in the structure and receptor binding efficacy of the antibody post-conjugation.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Cetuximab/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Quimera Dirigida a la Proteólisis , Regulación hacia Abajo , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Receptores ErbB/metabolismo , Mutación , ProteolisisRESUMEN
The Galß(1â4)Man disaccharide, found in the cell surface lipophosphoglycan (LPG) of Leishmania species, has been synthesized by a Ferrier glycosylation/cis-dihydroxylation strategy. This stereoselective method proved efficient for synthesizing the target saccharide in good yield. In addition, we prepared two clickable O-glycoside and phospho-glycoside versions of Galß(1â4)Man to enable conjugation to protein carriers for further immunological and antibody-binding studies.
RESUMEN
Locally delivered pre-exposure prophylaxis (PrEP) has proven to be a promising strategy to combat Human immunodeficiency virus (HIV) transmission but several findings encountered toxicities or proved to be marginally effective in clinical settings. Therefore, innovative, multifunctional, and safer alternatives are being progressively investigated. Herein, we explored negatively charged carbohydrate, anionic pullulan (AP) as a rapidly soluble film-former and novel anti-HIV agent. Additionally, Bictegravir (BCT), an HIV integrase inhibitor was co-delivered in the form of nanomicelles for sustained antiviral activity. BCT-loaded PLGA-PEG polymeric nanomicelles (BN) were incorporated into PVA/pullulan-based film matrix comprising of 2 % w/v AP (BN-AP film). In cell-based assays, biocompatibility and TEER values for BN-AP films were similar to control while the commercial vaginal contraceptive film (VCF®) showed severe cytotoxicity and drastically reduced the tight junction integrity. Rapid disintegration of BN-AP film with >85 % drug release was observed in simulated vaginal and seminal fluid. Most importantly, AP and BN-AP film significantly inhibited HIV-1 replication with IC50 at as low as 91 µg/mL and 0.708 nM, respectively. Therefore, this study entails successful development of BN-AP film that functioned as an effective, biocompatible dual-acting PrEP formulation.
Asunto(s)
Fármacos Anti-VIH , Infecciones por VIH , Profilaxis Pre-Exposición , Femenino , Humanos , Fármacos Anti-VIH/farmacología , Administración Intravaginal , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/prevención & controlRESUMEN
Systemically delivered lipid nanoparticles are preferentially taken up by hepatocytes. This hinders the development of effective, non-viral means of editing genes in tissues other than the liver. Here we show that lipid-nanoparticle-mediated gene editing in the lung and spleen of adult mice can be enhanced by reducing Cas9-mediated insertions and deletions in hepatocytes via oligonucleotides disrupting the secondary structure of single-guide RNAs (sgRNAs) and also via their combination with short interfering RNA (siRNA) targeting Cas9 messenger RNA (mRNA). In SpCas9 mice with acute lung inflammation, the systemic delivery of an oligonucleotide inhibiting an sgRNA targeting the intercellular adhesion molecule 2 (ICAM-2), followed by the delivery of the sgRNA, reduced the fraction of ICAM-2 indels in hepatocytes and increased that in lung endothelial cells. In wild-type mice, the lipid-nanoparticle-mediated delivery of an inhibitory oligonucleotide, followed by the delivery of Cas9-degrading siRNA and then by Cas9 mRNA and sgRNA, reduced the fraction of ICAM-2 indels in hepatocytes but not in splenic endothelial cells. Inhibitory oligonucleotides and siRNAs could be used to modulate the cell-type specificity of Cas9 therapies.
Asunto(s)
Edición Génica , Nanopartículas , Animales , Antígenos CD , Sistemas CRISPR-Cas , Moléculas de Adhesión Celular/genética , Células Endoteliales , Lípidos/química , Liposomas , Hígado , Pulmón , Ratones , Nanopartículas/química , BazoRESUMEN
Aggresomes are dynamic structures formed when the ubiquitin-proteasome system is overwhelmed with aggregation-prone proteins. In this process, small protein aggregates are actively transported towards the microtubule-organizing center. A functional role for autophagy in the clearance of aggresomes has also been proposed. In the present work we investigated the molecular mechanisms involved on aggresome formation in cultured rat cardiac myocytes exposed to glucose deprivation. Confocal microscopy showed that small aggregates of polyubiquitinated proteins were formed in cells exposed to glucose deprivation for 6 h. However, at longer times (18 h), aggregates formed large perinuclear inclusions (aggresomes) which colocalized with gamma-tubulin (a microtubule-organizing center marker) and Hsp70. The microtubule disrupting agent vinblastine prevented the formation of these inclusions. Both small aggregates and aggresomes colocalized with autophagy markers such as GFP-LC3 and Rab24. Glucose deprivation stimulates reactive oxygen species (ROS) production and decreases intracellular glutathione levels. ROS inhibition by N-acetylcysteine or by the adenoviral overexpression of catalase or superoxide dismutase disrupted aggresome formation and autophagy induced by glucose deprivation. In conclusion, glucose deprivation induces oxidative stress which is associated with aggresome formation and activation of autophagy in cultured cardiac myocytes.
Asunto(s)
Autofagia/fisiología , Glucosa/deficiencia , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Estrés Oxidativo , Multimerización de Proteína , Acetilcisteína/farmacología , Adenosina Trifosfato/metabolismo , Animales , Autofagia/efectos de los fármacos , Catalasa/metabolismo , Células Cultivadas , Glucosa/metabolismo , Glutatión/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Cuerpos de Inclusión/metabolismo , Microscopía Electrónica de Transmisión , Centro Organizador de los Microtúbulos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Complejo de la Endopetidasa Proteasomal/metabolismo , Transporte de Proteínas , Ratas , Especies Reactivas de Oxígeno/metabolismo , Superóxido Dismutasa/metabolismo , Tubulina (Proteína)/metabolismo , Ubiquitina/metabolismoAsunto(s)
Proteínas de Transporte de Catión/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Óxido Nítrico/metabolismo , Neoplasias Ováricas/metabolismo , Ovario/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismo , Femenino , Humanos , Óxido Nítrico Sintasa de Tipo II/genética , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Ovario/patología , Intercambiador 1 de Sodio-Hidrógeno , Regulación hacia ArribaRESUMEN
A series of alkyl ß-glucosyl sulfoxides were synthesized and characterized in order to study their stereochemical properties. The dependence of the aglycon, solvent and absolute configuration of the sulfinyl group on the conformational properties around the glucosidic and C5-C6 (hydroxymethyl group) bonds were studied. The results for R(S) sulfoxides show linear correlations between the rotamer populations of the hydroxymethyl group and the corresponding Taft's steric parameter (E(S)) of the alkyl group attached to the sulfinyl group in polar and apolar solvents, an increase in the absolute value of E(S) leading to an increase in the gt population. In addition, NOE experiments reveal that as the bulkiness of the alkyl group increases the population of the g- rotamer increases, the latter stabilized by the exo-anomeric effect. These results are in complete agreement with the participation of the exo-anomeric effect in both conformational properties of R(S) sulfoxides. Sulfoxides with the S(S) configuration show different behavior to their R(S) epimers; thus, an increase in the E(S) value of the alkyl group leads to similar or lower gt populations in apolar solvents and to increases in gt in polar solvents. Their NOE studies reveal a conformational equilibrium (in polar and apolar solvents) between g- and g+, dependent on the size of the alkyl group R attached to the sulfinyl group. All these results for both epimers support the general hypothesis that the exo-anomeric effect modifies the conformation of the hydroxymethyl group, fulfills the stereoelectronic requirements, and shows dependence on the solvent.
Asunto(s)
Glucosa/química , Sulfóxidos/química , Dicroismo Circular , Espectroscopía de Resonancia Magnética , Conformación Molecular , Soluciones , Solventes/química , EstereoisomerismoRESUMEN
Dendritic cells (DCs) are highly effective antigen-presenting cells that shape immune responses. Vaccines that deliver antigen to the DCs can harness their power. DC surface lectins recognize glycans not typically present on host tissue to facilitate antigen uptake and presentation. Vaccines that target these surface lectins should offer improved antigen delivery, but their efficacy will depend on how lectin targeting influences the T cell subtypes that result. We examined how antigen structure influences uptake and signaling from the C-type lectin DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin or CD209). Virus-like particles (VLPs) were engineered from bacteriophage Qß to present an array of mannoside ligands. The VLPs were taken up by DCs and efficiently trafficked to endosomes. The signaling that ensued depended on the ligand displayed on the VLP: only those particles densely functionalized with an aryl mannoside, Qß-Man540, elicited DC maturation and induced the expression of the proinflammatory cytokines characteristic of a T helper type 1 (TH1)-like immune response. This effect was traced to differential binding to DC-SIGN at the acidic pH of the endosome. Mice immunized with a VLP bearing the aryl mannoside, and a peptide antigen (Qß-Ova-Man540) had antigen-specific responses, including the production of CD4+ T cells producing the activating cytokines interferon-γ and tumor necrosis factor-α. A TH1 response is critical for intracellular pathogens (e.g., viruses) and cancer; thus, our data highlight the value of targeting DC lectins for antigen delivery and validate the utility of DC-targeted VLPs as vaccine vehicles that induce cellular immunity.
Asunto(s)
Células Dendríticas , Polisacáridos , Animales , Antígenos , Inmunidad Celular , Ratones , Linfocitos TRESUMEN
BACKGROUND: Outbreaks of Chikungunya virus (CHIKV) occurred in Brazil during 2015-2017. Fortaleza was the city that reported the most cases. METHODS: The first round of a cohort study was conducted among women aged 15-39 y in Fortaleza, Brazil, in 2018 (Zika in Fortaleza). We collected sera to detect CHIKV IgG and IgM antibodies. Factors for CHIKV infection were identified using a Poisson regression model. RESULTS: We evaluated 1466 serum samples and 13.8% and 37.2% of women were found positive for CHIKV IgM and IgG antibodies, respectively. Living with more than four others in the same house and having an abandoned house nearby were associated with CHIKV infection. Being currently pregnant was associated with a decreased probability of CHIKV infection, which was also associated with pregnant women reporting using more repellent, both inside and outside the house, than non-pregnant women. CONCLUSIONS: Crowding in households and abandoned houses nearby can increase potential transmission. Policies providing better living conditions and regulation of abandoned sites and buildings are necessary to control the mosquito population. Programmes providing repellant at low or no cost to pregnant women should be implemented in the neighbourhoods where arbovirus infections are endemic.
Asunto(s)
Fiebre Chikungunya , Virus Chikungunya , Infección por el Virus Zika , Virus Zika , Animales , Anticuerpos Antivirales , Brasil/epidemiología , Fiebre Chikungunya/epidemiología , Estudios de Cohortes , Femenino , Humanos , Embarazo , Infección por el Virus Zika/epidemiologíaRESUMEN
Caveolin-1 (CAV1) is a well-established nitric oxide synthase inhibitor, whose function as a tumor suppressor is favored by, but not entirely dependent on, the presence of E-cadherin. Tumors are frequently hypoxic and the activation of the hypoxia-inducible factor-1α (HIF1α) promotes tumor growth. HIF1α is regulated by several post-translational modifications, including S-nitrosylation. Here, we evaluate the mechanisms underlying tumor suppression by CAV1 in cancer cells lacking E-cadherin in hypoxia. Our main findings are that CAV1 reduced HIF activity and Vascular Endothelial Growth Factor expression in vitro and in vivo. This effect was neither due to reduced HIF1α protein stability or reduced nuclear translocation. Instead, HIF1α S-nitrosylation observed in hypoxia was diminished by the presence of CAV1, and nitric oxide synthase (NOS) inhibition by Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) reduced HIF1α transcriptional activity in cells to the same extent as observed upon CAV1 expression. Additionally, arginase inhibition by (S)-(2-Boronoethyl)-L-cysteine (BEC) partially rescued cells from the CAV1-mediated suppression of HIF1α transcriptional activity. In vivo, CAV1-mediated tumor suppression was dependent on NOS activity. In summary, CAV1-dependent tumor suppression in the absence of E-cadherin is linked to reduced HIF1α transcriptional activity via diminished NOS-mediated HIF1α S-nitrosylation.