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The SARS-CoV-2 spike protein is a critical component of vaccines and a target for neutralizing monoclonal antibodies (nAbs). Spike is also undergoing immunogenic selection with variants that increase infectivity and partially escape convalescent plasma. Here, we describe Spike Display, a high-throughput platform to rapidly characterize glycosylated spike ectodomains across multiple coronavirus-family proteins. We assayed â¼200 variant SARS-CoV-2 spikes for their expression, ACE2 binding, and recognition by 13 nAbs. An alanine scan of all five N-terminal domain (NTD) loops highlights a public epitope in the N1, N3, and N5 loops recognized by most NTD-binding nAbs. NTD mutations in variants of concern B.1.1.7 (alpha), B.1.351 (beta), B.1.1.28 (gamma), B.1.427/B.1.429 (epsilon), and B.1.617.2 (delta) impact spike expression and escape most NTD-targeting nAbs. Finally, B.1.351 and B.1.1.28 completely escape a potent ACE2 mimic. We anticipate that Spike Display will accelerate antigen design, deep scanning mutagenesis, and antibody epitope mapping for SARS-CoV-2 and other emerging viral threats.
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Mamíferos/virologia , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genética , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , COVID-19/imunologia , COVID-19/virologia , Linhagem Celular , Epitopos/genética , Epitopos/imunologia , Células HEK293 , Humanos , Mamíferos/imunologia , Ligação Proteica/genética , Ligação Proteica/imunologia , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/imunologiaRESUMO
A prototypical material for the oxidative coupling of methane (OCM) is Li/MgO, for which Li is known to be essential as a dopant to obtain high C2 selectivities. Herein, Li/MgO is demonstrated to be an effective catalyst for non-oxidative coupling of methane (NOCM). Moreover, the presence of Li is shown to favor the formation of magnesium acetylide (MgC2 ), while pure MgO promotes coke formation as evidenced by solid-state 13 Câ NMR, thus indicating that Li promotes C-C bond formation. Metadynamic simulations of the carbon mobility in MgC2 and Li2 C2 at the density functional theory (DFT) level show that carbon easily diffuses as a C2 unit at 1000 °C. These insights suggest that the enhanced C2 selectivity for Li-doped MgO is related to the formation of Li and Mg acetylides.
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Coronavirus 2019 disease (COVID-19) is associated with coagulation dysfunction that predisposes patients to an increased risk for both arterial (ATE) and venous thromboembolism (VTE) and consequent poor prognosis; in particular, the incidence of ATE and VTE in critically ill COVID-19 patients can reach 5% and 31%, respectively. The mechanism of thrombosis in COVID-19 patients is complex and still not completely clear. Recent literature suggests a link between the presence of antiphospholipid antibodies (aPLs) and thromboembolism in COVID-19 patients. However, it remains uncertain whether aPLs are an epiphenomenon or are involved in the pathogenesis of the disease.
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Anticorpos Antifosfolipídeos/imunologia , COVID-19/imunologia , SARS-CoV-2/imunologia , Tromboembolia/imunologia , Animais , Anticorpos Antifosfolipídeos/sangue , Coagulação Sanguínea , COVID-19/sangue , COVID-19/complicações , Estado Terminal , Humanos , Tromboembolia/sangue , Tromboembolia/complicações , Tromboembolia Venosa/sangue , Tromboembolia Venosa/complicações , Tromboembolia Venosa/imunologiaRESUMO
The selective conversion of methane to methanol remains one of the holy grails of chemistry, where Cu-exchanged zeolites have been shown promote this reaction under stepwise conditions. Over the years, several active sites have been proposed, ranging from mono-, di- to trimeric CuII . Herein, we report the formation of well-dispersed monomeric CuII species supported on alumina using surface organometallic chemistry and their reactivity towards the selective and stepwise conversion of methane to methanol. Extensive studies using various transition alumina supports combined with spectroscopic characterization, in particular electron paramagnetic resonance (EPR), show that the active sites are associated with specific facets, which are typically found in γ- and η-alumina phase, and that their EPR signature can be attributed to species having a tri-coordinated [(Al2 O)CuIIO(OH)]- T-shape geometry. Overall, the selective conversion of methane to methanol, a two-electron process, involves two monomeric CuII sites that play in concert.
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Single particle analysis for structure determination in cryo-electron microscopy is traditionally applied to samples purified to near homogeneity as current reconstruction algorithms are not designed to handle heterogeneous mixtures of structures from many distinct macromolecular complexes. We extend on long established methods and demonstrate that relating two-dimensional projection images by their common lines in a graphical framework is sufficient for partitioning distinct protein and multiprotein complexes within the same data set. The feasibility of this approach is first demonstrated on a large set of synthetic reprojections from 35 unique macromolecular structures spanning a mass range of hundreds to thousands of kilodaltons. We then apply our algorithm on cryo-EM data collected from a mixture of five protein complexes and use existing methods to solve multiple three-dimensional structures ab initio. Incorporating methods to sort single particle cryo-EM data from extremely heterogeneous mixtures will alleviate the need for stringent purification and pave the way toward investigation of samples containing many unique structures.
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Microscopia Crioeletrônica , Processamento de Imagem Assistida por Computador , Substâncias Macromoleculares/ultraestrutura , Complexos Multiproteicos/ultraestrutura , Imageamento Tridimensional , Substâncias Macromoleculares/química , Complexos Multiproteicos/químicaRESUMO
Non-oxidative CH4 coupling is promoted by silica with incorporated iron sites, but the role of these sites and their speciation under reaction conditions are poorly understood. Here, silica-supported iron(II) single sites, prepared via surface organometallic chemistry and stable at 1020 °C in vacuum, are shown to rapidly initiate CH4 coupling at 1000 °C, leading to 15-22 % hydrocarbons selectivity at 3-4 % conversion. During this process, iron reduces and forms carburized iron(0) nanoparticles. This reactivity contrasts with what is observed for (iron-free) partially dehydroxylated silica, that readily converts methane, albeit with low hydrocarbon selectivity and after an induction period. This study supports that iron sites facilitate faster initiation of radical reactions and tame the surface reactivity.
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Monomeric CuII sites supported on alumina, prepared using surface organometallic chemistry, convert CH4 to CH3 OH selectively. This reaction takes place by formation of CH3 O surface species with the concomitant reduction of two monomeric CuII sites to CuI , according to mass balance analysis, infrared, solid-state nuclear magnetic resonance, X-ray absorption, and electron paramagnetic resonance spectroscopy studies. This material contains a significant fraction of Cu active sites (22 %) and displays a selectivity for CH3 OH exceeding 83 %, based on the number of electrons involved in the transformation. These alumina-supported CuII sites reveal that C-H bond activation, along with the formation of CH3 O- surface species, can occur on pairs of proximal monomeric CuII sites in a short reaction time.
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A better understanding of antitumor immune responses is the key to advancing the field of cancer immunotherapy. Endogenous immunity in cancer patients, such as circulating anticancer antibodies or tumor-reactive B cells, has been historically yet incompletely described. Here, we demonstrate that tumor-draining (sentinel) lymph node (SN) is a rich source for tumor-reactive B cells that give rise to systemic IgG anticancer antibodies circulating in the bloodstream of breast cancer patients. Using a synergistic combination of high-throughput B-cell sequencing and quantitative immunoproteomics, we describe the prospective identification of tumor-reactive SN B cells (based on clonal frequency) and also demonstrate an unequivocal link between affinity-matured expanded B-cell clones in the SN and antitumor IgG in the blood. This technology could facilitate the discovery of antitumor antibody therapeutics and conceivably identify novel tumor antigens. Lastly, these findings highlight the unique and specialized niche the SN can fill in the advancement of cancer immunotherapy.
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Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/imunologia , Linfócitos B/imunologia , Neoplasias da Mama/imunologia , Células Clonais/imunologia , Imunoglobulina G/imunologia , Linfonodo Sentinela/imunologia , Sequência de Aminoácidos , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Células Cultivadas , Feminino , Humanos , Homologia de SequênciaRESUMO
Mass spectrometry (MS) has emerged as a powerful tool within the growing field of immunoproteomics, which aims to understand antibody-mediated immunity at the molecular-level based on the direct determination of serological antibody repertoire. To date, these methods have relied on the use of high-resolution bottom-up proteomic strategies that require effective sampling and characterization of low abundance peptides derived from the antigen-binding domains of polyclonal antibody mixtures. Herein, we describe a method that uses restricted Lys-C enzymatic digestion to increase the average mass of proteolytic IgG peptides (≥4.5 kDa) and produce peptides which uniquely derive from single antibody species. This enhances the capacity to discriminate between very similar antibodies present within polyclonal mixtures. Furthermore, our use of 193-nm ultraviolet photodissociation (UVPD) improves spectral coverage of the antibody sequence relative to conventional collision- and electron-based fragmentation methods. We apply these methods to both a monoclonal and an antibody mixture. By identifying from a database search of approximately 15â¯000 antibody sequences those which compose the mixture, we demonstrate the analytical potential of middle-down UVPD for MS-based serological repertoire analysis.
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Imunoglobulina G/análise , Imunoproteínas/análise , Proteoma/análise , Proteômica/métodos , Raios Ultravioleta , Humanos , Espectrometria de Massas , Processos FotoquímicosRESUMO
We describe a strategy for de novo peptide sequencing based on matched pairs of tandem mass spectra (MS/MS) obtained by collision induced dissociation (CID) and 351 nm ultraviolet photodissociation (UVPD). Each precursor ion is isolated twice with the mass spectrometer switching between CID and UVPD activation modes to obtain a complementary MS/MS pair. To interpret these paired spectra, we modified the UVnovo de novo sequencing software to automatically learn from and interpret fragmentation spectra, provided a representative set of training data. This machine learning procedure, using random forests, synthesizes information from one or multiple complementary spectra, such as the CID/UVPD pairs, into peptide fragmentation site predictions. In doing so, the burden of fragmentation model definition shifts from programmer to machine and opens up the model parameter space for inclusion of nonobvious features and interactions. This spectral synthesis also serves to transform distinct types of spectra into a common representation for subsequent activation-independent processing steps. Then, independent from precursor activation constraints, UVnovo's de novo sequencing procedure generates and scores sequence candidates for each precursor. We demonstrate the combined experimental and computational approach for de novo sequencing using whole cell E. coli lysate. In benchmarks on the CID/UVPD data, UVnovo assigned correct full-length sequences to 83% of the spectral pairs of doubly charged ions with high-confidence database identifications. Considering only top-ranked de novo predictions, 70% of the pairs were deciphered correctly. This de novo sequencing performance exceeds that of PEAKS and PepNovo on the CID spectra and that of UVnovo on CID or UVPD spectra alone. As presented here, the methods for paired CID/UVPD spectral acquisition and interpretation constitute a powerful workflow for high-throughput and accurate de novo peptide sequencing.
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Peptídeos/química , Análise de Sequência de Proteína , Processos Fotoquímicos , Espectrometria de Massas em Tandem , Raios UltravioletaRESUMO
Most vaccines confer protection via the elicitation of serum antibodies, yet more than 100 y after the discovery of antibodies, the molecular composition of the human serum antibody repertoire to an antigen remains unknown. Using high-resolution liquid chromatography tandem MS proteomic analyses of serum antibodies coupled with next-generation sequencing of the V gene repertoire in peripheral B cells, we have delineated the human serum IgG and B-cell receptor repertoires following tetanus toxoid (TT) booster vaccination. We show that the TT(+) serum IgG repertoire comprises â¼100 antibody clonotypes, with three clonotypes accounting for >40% of the response. All 13 recombinant IgGs examined bound to vaccine antigen with Kd â¼ 10(-8)-10(-10) M. Five of 13 IgGs recognized the same linear epitope on TT, occluding the binding site used by the toxin for cell entry, suggesting a possible explanation for the mechanism of protection conferred by the vaccine. Importantly, only a small fraction (<5%) of peripheral blood plasmablast clonotypes (CD3(-)CD14(-)CD19(+)CD27(++)CD38(++)CD20(-)TT(+)) at the peak of the response (day 7), and an even smaller fraction of memory B cells, were found to encode antibodies that could be detected in the serological memory response 9 mo postvaccination. This suggests that only a small fraction of responding peripheral B cells give rise to the bone marrow long-lived plasma cells responsible for the production of biologically relevant amounts of vaccine-specific antibodies (near or above the Kd). Collectively, our results reveal the nature and dynamics of the serological response to vaccination with direct implications for vaccine design and evaluation.
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Anticorpos Antibacterianos/biossíntese , Toxoide Tetânico/administração & dosagem , Sequência de Aminoácidos , Anticorpos Antibacterianos/química , Linfócitos B/imunologia , Cromatografia Líquida , Humanos , Imunofenotipagem , Dados de Sequência Molecular , Espectrometria de Massas em TandemRESUMO
De novo peptide sequencing by mass spectrometry represents an important strategy for characterizing novel peptides and proteins, in which a peptide's amino acid sequence is inferred directly from the precursor peptide mass and tandem mass spectrum (MS/MS or MS(3)) fragment ions, without comparison to a reference proteome. This method is ideal for organisms or samples lacking a complete or well-annotated reference sequence set. One of the major barriers to de novo spectral interpretation arises from confusion of N- and C-terminal ion series due to the symmetry between b and y ion pairs created by collisional activation methods (or c, z ions for electron-based activation methods). This is known as the "antisymmetric path problem" and leads to inverted amino acid subsequences within a de novo reconstruction. Here, we combine several key strategies for de novo peptide sequencing into a single high-throughput pipeline: high-efficiency carbamylation blocks lysine side chains, and subsequent tryptic digestion and N-terminal peptide derivatization with the ultraviolet chromophore AMCA yield peptides susceptible to 351 nm ultraviolet photodissociation (UVPD). UVPD-MS/MS of the AMCA-modified peptides then predominantly produces y ions in the MS/MS spectra, specifically addressing the antisymmetric path problem. Finally, the program UVnovo applies a random forest algorithm to automatically learn from and then interpret UVPD mass spectra, passing results to a hidden Markov model for de novo sequence prediction and scoring. We show this combined strategy provides high-performance de novo peptide sequencing, enabling the de novo sequencing of thousands of peptides from an Escherichia coli lysate at high confidence.
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Algoritmos , Espectrometria de Massas , Peptídeos/química , Raios Ultravioleta , Sequência de Aminoácidos , Escherichia coli/química , Íons/química , Processos Fotoquímicos , Análise de Sequência de ProteínaRESUMO
We have developed and validated a methodology for determining the antibody composition of the polyclonal serum response after immunization. Pepsin-digested serum IgGs were subjected to standard antigen-affinity chromatography, and resulting elution, wash, and flow-through fractions were analyzed by bottom-up, liquid chromatography-high-resolution tandem mass spectrometry. Identification of individual monoclonal antibodies required the generation of a database of IgG variable gene (V-gene) sequences constructed by NextGen sequencing of mature B cells. Antibody V-gene sequences are characterized by short complementarity determining regions (CDRs) of high diversity adjacent to framework regions shared across thousands of IgGs, greatly complicating the identification of antigen-specific IgGs from proteomically observed peptides. By mapping peptides marking unique V(H) CDRH3 sequences, we identified a set of V-genes heavily enriched in the affinity chromatography elution, constituting the serum polyclonal response. After booster immunization in a rabbit, we find that the antigen-specific serum immune response is oligoclonal, comprising antibodies encoding 34 different CDRH3s that group into 30 distinct antibody V(H) clonotypes. Of these 34 CDRH3s, 12 account for â¼60% of the antigen-specific CDRH3 peptide mass spectral counts. For comparison, antibodies with 18 different CDRH3s (12 clonotypes) were represented in the antigen-specific IgG fraction from an unimmunized rabbit that fortuitously displayed a moderate titer for BSA. Proteomically identified antibodies were synthesized and shown to display subnanomolar affinities. The ability to deconvolute the polyclonal serum response is likely to be of key importance for analyzing antibody responses after vaccination and for more completely understanding adaptive immune responses in health and disease.
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Anticorpos Monoclonais/genética , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Cromatografia de Afinidade , Regiões Determinantes de Complementaridade , Imunoglobulina G/genética , Imunoglobulina G/imunologia , Espectrometria de Massas , Dados de Sequência Molecular , Proteômica , CoelhosRESUMO
The use of ultraviolet photodissociation (UVPD) for the activation and dissociation of peptide anions is evaluated for broader coverage of the proteome. To facilitate interpretation and assignment of the resulting UVPD mass spectra of peptide anions, the MassMatrix database search algorithm was modified to allow automated analysis of negative polarity MS/MS spectra. The new UVPD algorithms were developed based on the MassMatrix database search engine by adding specific fragmentation pathways for UVPD. The new UVPD fragmentation pathways in MassMatrix were rigorously and statistically optimized using two large data sets with high mass accuracy and high mass resolution for both MS(1) and MS(2) data acquired on an Orbitrap mass spectrometer for complex Halobacterium and HeLa proteome samples. Negative mode UVPD led to the identification of 3663 and 2350 peptides for the Halo and HeLa tryptic digests, respectively, corresponding to 655 and 645 peptides that were unique when compared with electron transfer dissociation (ETD), higher energy collision-induced dissociation, and collision-induced dissociation results for the same digests analyzed in the positive mode. In sum, 805 and 619 proteins were identified via UVPD for the Halobacterium and HeLa samples, respectively, with 49 and 50 unique proteins identified in contrast to the more conventional MS/MS methods. The algorithm also features automated charge determination for low mass accuracy data, precursor filtering (including intact charge-reduced peaks), and the ability to combine both positive and negative MS/MS spectra into a single search, and it is freely open to the public. The accuracy and specificity of the MassMatrix UVPD search algorithm was also assessed for low resolution, low mass accuracy data on a linear ion trap. Analysis of a known mixture of three mitogen-activated kinases yielded similar sequence coverage percentages for UVPD of peptide anions versus conventional collision-induced dissociation of peptide cations, and when these methods were combined into a single search, an increase of up to 13% sequence coverage was observed for the kinases. The ability to sequence peptide anions and cations in alternating scans in the same chromatographic run was also demonstrated. Because ETD has a significant bias toward identifying highly basic peptides, negative UVPD was used to improve the identification of the more acidic peptides in conjunction with positive ETD for the more basic species. In this case, tryptic peptides from the cytosolic section of HeLa cells were analyzed by polarity switching nanoLC-MS/MS utilizing ETD for cation sequencing and UVPD for anion sequencing. Relative to searching using ETD alone, positive/negative polarity switching significantly improved sequence coverages across identified proteins, resulting in a 33% increase in unique peptide identifications and more than twice the number of peptide spectral matches.
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Cromatografia Líquida/métodos , Bases de Dados de Proteínas , Ensaios de Triagem em Larga Escala , Proteoma/metabolismo , Proteômica/métodos , Espectrometria de Massas em Tandem/métodos , Raios Ultravioleta , Algoritmos , Ânions , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Halobacterium/metabolismo , Células HeLa , Humanos , Concentração de Íons de Hidrogênio , Peso Molecular , Peptídeos/metabolismo , Proteoma/química , Curva ROC , Reprodutibilidade dos Testes , Análise de Sequência de ProteínaRESUMO
Characterizing the in vivo dynamics of the polyclonal antibody repertoire in serum, such as that which might arise in response to stimulation with an antigen, is difficult due to the presence of many highly similar immunoglobulin proteins, each specified by distinct B lymphocytes. These challenges have precluded the use of conventional mass spectrometry for antibody identification based on peptide mass spectral matches to a genomic reference database. Recently, progress has been made using bottom-up analysis of serum antibodies by nanoflow liquid chromatography/high-resolution tandem mass spectrometry combined with a sample-specific antibody sequence database generated by high-throughput sequencing of individual B cell immunoglobulin variable domains (V genes). Here, we describe how intrinsic features of antibody primary structure, most notably the interspersed segments of variable and conserved amino acid sequences, generate recurring patterns in the corresponding peptide mass spectra of V gene peptides, greatly complicating the assignment of correct sequences to mass spectral data. We show that the standard method of decoy-based error modeling fails to account for the error introduced by these highly similar sequences, leading to a significant underestimation of the false discovery rate. Because of these effects, antibody-derived peptide mass spectra require increased stringency in their interpretation. The use of filters based on the mean precursor ion mass accuracy of peptide-spectrum matches is shown to be particularly effective in distinguishing between "true" and "false" identifications. These findings highlight important caveats associated with the use of standard database search and error-modeling methods with nonstandard data sets and custom sequence databases.
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Anticorpos Monoclonais/sangue , Proteômica/métodos , Sequência de Aminoácidos , Animais , Reações Antígeno-Anticorpo , Cromatografia Líquida de Alta Pressão , Cisteína/química , Humanos , Dados de Sequência Molecular , Coelhos , Espectrometria de Massas em TandemRESUMO
PURPOSE: A hemodynamic event such as hypertension after interscalene block (ISB) is a complication that is often overlooked. The irregular spread of local anesthetic would cause a blockade of carotid sinus baroreceptors leading to the adverse event. The purpose of the present study is to compare ultrasound and neurostimulation technique in preventing hypertension after ISB. METHODS: Thirty patients without hypertension history who underwent arthroscopic shoulder surgery for a rotator cuff tear were enrolled. After preoperative administration of the State Trait Anxiety Inventory questionnaire, patients were allocated to receive ultrasound-guided ISB with 20 ml levobupivacaine-HCl 0.5 % (group US) and 40 ml levobupivacaine-HCl 0.5 % with neurostimulation (group NS). The need for antihypertensive drug was recorded. Block onset sensory and motor times were assessed. Systolic and diastolic blood pressures, and heart rate and pulse oximetry (SpO2), were evaluated before the block as well as 2, 5, 10, 15, 20, and 30 min after. RESULTS: No differences in patient characteristics and anxiety were found in the two groups. Block onset times were similar. At 15 min after block placement, group NS showed significantly higher systolic and diastolic blood pressures compared to group US. No differences in heart rate and SpO2 were found between the two groups. Three patients of group NS required urapidil administration because of hypertension. CONCLUSIONS: Ultrasound-guided ISB permits the use of a low volume of local anesthetic and seems to reduce the incidence of hypertension.
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Anestésicos Locais/uso terapêutico , Bupivacaína/análogos & derivados , Hemodinâmica , Hipertensão/etiologia , Bloqueio Nervoso/efeitos adversos , Bloqueio Nervoso/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Anestésicos Locais/administração & dosagem , Artroscopia/métodos , Plexo Braquial/diagnóstico por imagem , Bupivacaína/administração & dosagem , Bupivacaína/uso terapêutico , Feminino , Humanos , Levobupivacaína , Masculino , Pessoa de Meia-Idade , Ombro/cirurgia , Ultrassonografia/efeitos adversos , Ultrassonografia/métodosRESUMO
We present the case of a 57-year-old female who initially presented with a chief complaint of left-sided orbital headaches and associated left eyelid swelling. Initial imaging work-up with CT head/orbit revealed soft tissue enhancement of the left orbital roof, concerning for neoplastic process (primary lymphoma versus extracranial primary tumor versus metastatic tumor). Further imaging studies with CT chest/abdomen/pelvis revealed an anterior mediastinal mass, concerning for possible thymoma versus lymphoma. The patient underwent further consultation with the Hematology/Oncology and Ophthalmology Departments, which recommended definitive biopsies from both sites, which showed matching histologic findings of moderately differentiated enteric-type adenocarcinoma with positive staining for CDX2, an intestinal marker. Thymic carcinomas are rare cancers that account for approximately 0.06% of all malignancies and require a high degree of clinical suspicion. Extrathoracic metastases from thymic carcinomas, especially to the orbit, is a rare occurrence and the exact incidence of this phenomenon is unknown. This case represents the diagnostic challenges associated with a rare cancer type and underscores the importance of a multidisciplinary approach to patient care.
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Transition metal carbides have numerous applications and are known to excel in terms of hardness, thermal stability and conductivity. In particular, the Pt-like behavior of Mo and W carbides has led to the popularization of metal carbides in catalysis, ranging from electrochemically-driven reactions to thermal methane coupling. Herein, we show the active participation of carbidic carbon in the formation of C2 products during methane coupling at high temperature that is associated with the dynamics of Mo and W carbides. A detailed mechanistic study reveals that the catalyst performance of these metal carbides can be traced back to its carbon diffusivity and exchange capability upon interaction with methane (gas phase carbon). A stable C2 selectivity over time on stream for Mo carbide (Mo2C) can be rationalized by fast carbon diffusion dynamics, while W carbide (WC) shows loss of selectivity due to slow diffusion leading to surface carbon depletion. This finding showcases that the bulk carbidic carbon of the catalyst plays a crucial role and that the metal carbide is not only responsible for methyl radical formation. Overall, this study evidences the presence of a carbon equivalent to the Mars-Van Krevelen type mechanism for non-oxidative coupling of methane.
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The ongoing evolution of SARS-CoV-2 into more easily transmissible and infectious variants has provided unprecedented insight into mutations enabling immune escape. Understanding how these mutations affect the dynamics of antibody-antigen interactions is crucial to the development of broadly protective antibodies and vaccines. Here we report the characterization of a potent neutralizing antibody (N3-1) identified from a COVID-19 patient during the first disease wave. Cryogenic electron microscopy revealed a quaternary binding mode that enables direct interactions with all three receptor-binding domains of the spike protein trimer, resulting in extraordinary avidity and potent neutralization of all major variants of concern until the emergence of Omicron. Structure-based rational design of N3-1 mutants improved binding to all Omicron variants but only partially restored neutralization of the conformationally distinct Omicron BA.1. This study provides new insights into immune evasion through changes in spike protein dynamics and highlights considerations for future conformationally biased multivalent vaccine designs.
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COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/genética , Anticorpos NeutralizantesRESUMO
Systemic coagulation abnormalities including clotting activation and inhibition of anticoagulant factors have been observed in patients with pneumonia. In severe coronavirus disease 2019 (COVID-19) the alteration of coagulation parameters was associated with poor prognosis. We evaluated the difference in coagulopathy between critically ill patients with COVID-19 pneumonia (COVID group) and non-COVID-19 pneumonia (non-COVID group), using traditional coagulation markers and rotational thromboelastometry (ROTEM). Standard laboratory and ROTEM parameters were evaluated in 45 patients (20 COVID group patients and 25 non-COVID group patients) at time of admission to the Intensive Care Unit (ICU) (T0) and at 5 (T5) and 10 days (T10) later. In all evaluations times, platelet count was found higher in COVID group rather than in non-COVID group. At T0, COVID group revealed a fibrinogen value greater than non-COVID group. d-Dimer values were high in both groups and they were not statistically different. At T0 COVID group showed a significant reduction of clot formation time in INTEM and in EXTEM and a significant increase of maximum clot firmness in INTEM, EXTEM and FIBTEM respect to non-COVID group. Moreover, COVID group demonstrated a coagulability state with ROTEM profiles higher than non-COVID group at T5 and T10. Coagulation profiles showed that critically ill patients with COVID-19 pneumonia are characterized by a higher coagulable state than others; this greater procoagulative state persists over time.