RESUMO
An enhancer trap (ET) mediated by a transposon is an effective method for functional gene research. Here, an ET system based on a PB transposon that carries a mini Krt4 promoter (the keratin4 minimal promoter from zebrafish) and the green fluorescent protein gene (GFP) has been used to produce zebrafish ET lines. One enhancer trap line with eye-specific expression GFP named EYE was used to identify the trapped enhancers and genes. Firstly, GFP showed a temporal and spatial expression pattern with whole-embryo expression at 6, 12, and 24 hpf stages and eye-specific expression from 2 to 7 dpf. Then, the genome insertion sites were detected by splinkerette PCR (spPCR). The Krt4-GFP was inserted into the fourth intron of the gene itgav (integrin, alpha V) in chromosome 9 of the zebrafish genome, with the GFP direction the same as that of the itgav gene. By the alignment of homologous gene sequences in different species, three predicted endogenous enhancers were obtained. The trapped endogenous gene itgav, whose overexpression is related to hepatocellular carcinoma, showed a similar expression pattern as GFP detected by in situ hybridization, which suggested that GFP and itgav were possibly regulated by the same enhancers. In short, the zebrafish enhancer trap lines generated by the PB transposon-mediated enhancer trap technology in this study were valuable resources as visual markers to study the regulators and genes. This work provides an efficient method to identify and isolate tissue-specific enhancer sequences.
RESUMO
Objective: To measure the parameters of eye movement of young adult patients of myopia, to compare the differences in the parameters of patient groups with varied degrees of myopia, and to analyze the correlation between eye movement parameters and axial length (AL) and spherical equivalent refraction (SER). Methods: A total of 91 young adult patients of myopia were recruited. The subjects were divided into three groups of low, moderate, and high myopia according to their SER. Information on the subjects' age and sex was collected and general clinical examination was completed. The subjects' binocular fixation, reflexive saccade, and antisaccade were measured with eye tracker. One-way ANOVA or Kruskal-wallis test was used to compare the general data and eye movement parameters of the three groups of myopic patients. Pearson or Spearman correlation analysis was used to analyze AL and SER's correlation with 95% bivariate contour ellipse area (BCEA) and saccadic parameters. Results: The 95% BCEA for mild, moderate, and high myopia groups were 2.08 (0.54, 14.69) deg 2, 4.99 (0.94, 49.22) deg 2, and 5.79 (2.18, 64.91) deg 2, respectively. There was significant difference between the 95% BCEA of the three groups ( P=0.029). The 95% BCEA of the mild myopia group was significantly smaller than that of the high myopia group ( P=0.01). There were no significant differences in saccadic parameters among the three groups ( P>0.05). There was a significant positive correlation between 95% BCEA and AL ( r=0.342, P=0.001). There was a significant negative correlation between 95% BCEA and SER ( r=-0.322, P=0.002). There was no significant correlation between the parameters of saccadic movement and the AL or SER ( P>0.05). Conclusion: For young adult myopic patients, the longer the AL is, the higher the degree of the myopia and the worse the fixation stability becomes. The fixation stability of patients with high myopia was significantly worse than that of patients with mild myopia. There was no significant difference in saccadic parameters in patients with different degrees of myopia.
Assuntos
Movimentos Oculares , Miopia , Comprimento Axial do Olho , Humanos , Refração Ocular , Adulto JovemRESUMO
An unprecedented Zn(OTf)2-catalyzed asymmetric Michael addition/cyclization cascade of 3-nitro-2H-chromenes with 3-isothiocyanato oxindoles has been disclosed. This transformation provides an efficient access to various synthetically important polycyclic spirooxindoles in a highly stereoselective manner under mild conditions (7299% yields, up to >95:5 d.r. and >99% ee). The reaction leads to the formation of three consecutive stereocenters, including 1,3-nonadjacent tetrasubstituted carbon stereocenters, in a single operation. A bifunctional activation model of the chiral Zn(OTf)2/bis(oxazoline) complex was proposed based on control experiments, wherein the ZnII moiety serves as a Lewis acid and the N atom of the free NH group acts as a Lewis base by a hydrogen-bonding interaction.
Assuntos
Benzopiranos/química , Indóis/síntese química , Compostos Policíclicos/síntese química , Compostos de Espiro/síntese química , Catálise , Ciclização , Indóis/química , Estrutura Molecular , Oxindóis , Compostos Policíclicos/química , Compostos de Espiro/química , EstereoisomerismoRESUMO
In this study, we have successfully designed and synthesized two novel dual-emission emitters featuring phenothiazine-5-oxide and phenothiazine-5,5-dioxide motifs, characterized by highly lopsided and asymmetric conformational states. Through rigorous spectral examinations and DFT calculations, the compounds exhibit distinctive ICT phenomena, coupled with efficient emission in solid states and AIEE characteristics under high water fractions in DMF/H2O mixtures. These non-planar luminogens exhibit vibrant green and blue solid-state luminescence, with fluorescence quantum yields of 24.1 % and 15.21 %, respectively. Additionally, they both emit green fluorescence in THF solution, with notable emission quantum yields (QYs) 36.4 % and 30.4 %. Comprehensive theoretical investigations unveil well-defined electron cloud density separation between the energies of HOMO/LUMO levels within the two luminogens. Notably, the targeted molecule harboring the phenothiazine-S,S-dioxide motif also demonstrates remarkable reversible mechanofluorochromic properties. Moreover, we testify their potential in applications such as solid-state rewritable information storage and live-cell imaging in solution states. Through theoretical calculations and comparative studies, we have explored the intrinsic relationship between molecular structure and performance, effectively screening and identifying new fluorescent molecules exhibiting outstanding luminescent attributes. These discoveries establish a robust theoretical and technical foundation for the synthesis and application of efficient DSE-based MFC materials, opening new avenues in the realm of advanced luminescent materials.
RESUMO
In this work, two novel luminescent molecules containing distorted phenothiazine-S-oxide and phenothiazine-S,S dioxide skeletons were synthesized by oxidation reactions using different oxidants (m-chloroperoxybenzoic acid, acetic acid /hydrogen peroxide). The target compounds were all confirmed by 1H NMR, 13C NMR and EI-MS. Combined with the results of UV-vis absorption spectra and fluorescence emission spectra, we found that the different oxidation states of S-atom, from sulfide (+2) to sulfoxide (+4) and sulfone (+6), led to the blue, yellow-green and yellowish fluorescence of these compounds in the solid states. Subsequent studies showed that the molecule containing the phenothiazine-S-oxide skeleton exhibited obvious solvatochromism, and the increase of solvent polarity induced a red-shift in the emission wavelength. Moreover, this molecule also exhibited a rare self-recovery mechanochromatic behavior. In addition, these properties were further confirmed by theoretical calculations and X-ray single-crystal diffraction analysis.
RESUMO
Batting the ylides: A simple procedure carried out under mild conditions allows the direct and efficient synthesis of structurally diverse indoles. This approach involves a cascade reaction of sulfur ylides and N-(ortho-chloromethyl)aryl amides.
Assuntos
Amidas/química , Indóis/química , Enxofre/química , Compostos Heterocíclicos/química , Indóis/síntese química , Nitrogênio/químicaRESUMO
A series of thiourea-amine bifunctional catalysts have been developed by a rational combination of prolines with cinchona alkaloids, which are connected by a thiourea motif. The catalyst 3a, prepared from L-proline and cinchonidine, was found to be a highly efficient catalyst for the conjugate addition of ketones/aldehydes to a wide range of nitroalkenes (up to 98/2 dr and 96% ee). The privileged cinchonidine backbone and the thiourea motif are essential to the reaction activity and enantioselectivity.
Assuntos
Aldeídos/química , Alcenos/química , Aminas/química , Reagentes de Ligações Cruzadas/química , Cetonas/química , Tioureia/química , Catálise , Alcaloides de Cinchona/química , Prolina/químicaRESUMO
We report the clinical characteristics, viral shedding duration, and contact tracing for asymptomatic carriers of SARS-CoV-2 in Wuhan, China. The asymptomatic carriers were relatively young (median age: 34.5 years). Chest computed tomography showed no abnormalities. The nasopharyngeal swab was an optimum specimen for RNA testing. The median viral shedding duration was 11.5 days. Notably, 2 months of viral shedding duration were reported in two nurses, which was much longer than previously reported or than usually thought. The transmissibility of SARS-CoV-2 by asymptomatic carriers during the studied period in Wuhan appeared to be weak. Only one patient (1/12) was found to have transmitted the virus to another person. Early asymptomatic carrier detection, isolation, and contact tracing could be useful to mitigate the spread of the disease.
RESUMO
INTRODUCTION: Since the coronavirus disease 2019 (COVID-19) outbreak in Wuhan in late 2019, controversy on the use of corticosteroids for COVID-19 has obtained increasing attention. We present 1 critically ill patient who had a rapid therapeutic response to moderate-dose corticosteroids. PATIENT CONCERNS: A 53-year-old critically ill woman from Wuhan suffered with COVID-19. DIAGNOSIS: The chest computed tomography scan was suggestive of COVID-19. The diagnosis was confirmed by a real-time reverse transcription polymerase chain reaction test for SARS-CoV-2. The critically ill status was characterized by worsening dyspnea, progressing bilateral lung consolidation, and poor oxygenation (SiO2/FiO2:110âmm Hg). INTERVENTIONS: The patient was treated with a moderate dose of intravenous corticosteroids and high-flow nasal cannula oxygen therapy. OUTCOMES: After the initiation of corticosteroids, the patient rapidly improved over the following 6 days. Serial chest computed tomography scans showed good absorption of the consolidations. The patient was discharged on Day 17 of hospitalization without obvious adverse effects. CONCLUSIONS: Early use of moderate-dose corticosteroids over a short period may enhance recovery from COVID-19 in critically ill patients.
Assuntos
Corticosteroides/administração & dosagem , Infecções por Coronavirus/tratamento farmacológico , Oxigenoterapia/métodos , Pneumonia Viral/tratamento farmacológico , Betacoronavirus , COVID-19 , Terapia Combinada , Infecções por Coronavirus/virologia , Feminino , Humanos , Pessoa de Meia-Idade , Pandemias , Pneumonia Viral/virologia , SARS-CoV-2 , Resultado do Tratamento , Tratamento Farmacológico da COVID-19RESUMO
Clinical data on coronavirus disease 2019 (COVID-19) with pancreatic injury are extremely limited. An acute manifestation of acute pancreatitis in COVID-19 has not been reported. We describe here 2 cases of COVID-19 with acute pancreatitis as the initial manifestation in Wuhan, China. Patient 1 died despite maximal mechanical ventilatory support and circulation support, while patient 2 was finally discharged after showing significant improvement. Low T cells in peripheral blood may indicate a poor outcome.
RESUMO
Chiral tetrahydrocarbazoles (THCs) are prevalent in numerous natural indole alkaloids as well as synthetic pharmaceuticals, and exhibit a broad spectrum of bioactivities. As such, the development of efficient synthetic methodologies for the synthesis of chiral THCs is of substantial interest. The advent of asymmetric catalysis provides a powerful platform to assemble chiral THC motifs and great efforts have been devoted to this field over the past decades. In this feature article, we summarise recent advances in catalytic asymmetric synthesis of THCs, with particular emphases on reaction types and reaction mechanism.
Assuntos
Carbazóis/síntese química , Alquilação , Carbazóis/química , Catálise , Reação de Cicloadição , Indóis/química , Ácidos de Lewis/química , Estrutura Molecular , EstereoisomerismoRESUMO
INTRODUCTION: Von Willebrand factor (VWF) plays a critical role in hemostasis by carrying factor VIII (FVIII) and binding to specific ligands on the surface of blood platelets and within the blood vessel wall. MATERIALS AND METHODS: We constructed a gene-specific phage display library containing small, random VWF fragments. Using this library, we mapped the repertoire of immune epitopes recognized by a commonly used commercial rabbit antihuman VWF polyclonal antibody. RESULTS: A total of eight discrete epitopes within the VWF protein were identified, including two dominant epitopes that account for 74% of immuno selected VWF fragments. Comparison with previously mapped epitopes for mouse monoclonal antibodies reveals four overlapping regions that may identify common antigenic determinants. The distribution of these epitopes was not readily predicted from primary amino acid sequence divergence among these mammalian species or standard algorithms for the prediction of antigenicity, hydrophobicity, or surface probability. CONCLUSION: Taken together with previous monoclonal antibody epitope mapping studies, our results suggest that a limited number of exposed domains on the surface of the human VWF protein may be the primary determinants of immunogenicity.
Assuntos
Anticorpos/imunologia , Fator de von Willebrand/imunologia , Animais , Anticorpos Monoclonais/imunologia , Ensaio de Imunoadsorção Enzimática , Mapeamento de Epitopos , Humanos , Camundongos , CoelhosRESUMO
BACKGROUND: For patients with non-small-cell lung cancer (NSCLC) and multiple brain metastases, whole-brain irradiation (WBI) is a standard-of-care treatment, but its effects on neurocognition are complex and concerning. We compared the efficacy of an epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI), icotinib, versus WBI with or without chemotherapy in a phase 3 trial of patients with EGFR-mutant NSCLC and multiple brain metastases. METHODS: We did a multicentre, open-label, parallel randomised controlled trial (BRAIN) at 17 hospitals in China. Eligible participants were patients with NSCLC with EGFR mutations, who were naive to treatment with EGFR-TKIs or radiotherapy, and had at least three metastatic brain lesions. We randomly assigned participants (1:1) to either icotinib 125 mg orally (three times per day) or WBI (30 Gy in ten fractions of 3 Gy) plus concurrent or sequential chemotherapy for 4-6 cycles, until unacceptable adverse events or intracranial disease progression occurred. The randomisation was done by the Chinese Thoracic Oncology Group with a web-based allocation system applying the Pocock and Simon minimisation method; groups were stratified by EGFR gene mutation status, treatment line (first line or second line), brain metastases only versus both intracranial and extracranial metastases, and presence or absence of symptoms of intracranial hypertension. Clinicians and patients were not masked to treatment assignment, but individuals involved in the data analysis did not participate in the treatments and were thus masked to allocation. Patients receiving icotinib who had intracranial progression only were switched to WBI plus either icotinib or chemotherapy until further progression; those receiving icotinib who had extracranial progression only were switched to icotinib plus chemotherapy. Patients receiving WBI who progressed were switched to icotinib until further progression. Icotinib could be continued beyond progression if a clinical benefit was observed by the investigators (eg, an improvement in cognition or intracranial pressure). The primary endpoint was intracranial progression-free survival (PFS), defined as the time from randomisation to either intracranial disease progression or death from any cause. We assessed efficacy and safety in the intention-to-treat population (all participants who received at least one dose of study treatment), hypothesising that intracranial PFS would be 40% longer (hazard ratio [HR] 0·60) with icotinib compared with WBI. This trial is registered with ClinicalTrials.gov, number NCT01724801. FINDINGS: Between Dec 10, 2012, and June 30, 2015, we assigned 176 participants to treatment: 85 to icotinib and 91 to WBI. 18 withdrew from the WBI group before treatment, leaving 73 for assessment. Median follow-up was 16·5 months (IQR 11·5-21·5). Median intracranial PFS was 10·0 months (95% CI 5·6-14·4) with icotinib versus 4·8 months (2·4-7·2) with WBI (equating to a 44% risk reduction with icotinib for an event of intracranial disease progression or death; HR 0·56, 95% CI 0·36-0·90; p=0·014). Adverse events of grade 3 or worse were reported in seven (8%) of 85 patients in the icotinib group and 28 (38%) of 73 patients in the WBI group. Raised concentrations of alanine aminotransferase and rash were the most common adverse events of any grade in both groups, occurring in around 20-30% of each group. At the time of final analysis, 42 (49%) patients in the icotinib group and 37 (51%) in the WBI group had died. 78 of these patients died from disease progression, and one patient in the WBI group died from thrombogenesis related to chemotherapy. INTERPRETATION: In patients with EGFR-mutant NSCLC and multiple brain metastases, icotinib was associated with significantly longer intracranial PFS than WBI plus chemotherapy, indicating that icotinib might be a better first-line therapeutic option for this patient population. FUNDING: Guangdong Provincial Key Laboratory of Lung Cancer Translational Medicine, National Health and Family Planning Commission of China, Guangzhou Science and Technology Bureau, Betta Pharmaceuticals, and the Chinese Thoracic Oncology Group.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/radioterapia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Irradiação Craniana/métodos , Éteres de Coroa/uso terapêutico , Receptores ErbB/genética , Neoplasias Pulmonares , Quinazolinas/uso terapêutico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/secundário , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/secundário , Intervalo Livre de Doença , Receptores ErbB/antagonistas & inibidores , Feminino , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Mutação , Modelos de Riscos Proporcionais , Resultado do TratamentoRESUMO
In this study, five microalgae strains were cultured for their ability to survive in biogas slurry, remove nitrogen resources and accumulate carbohydrates. It was proved that five microalgae strains adapted in biogas slurry well without ammonia inhibition. Among them, Chlorella vulgaris ESP-6 showed the best performance on carbohydrate accumulation, giving the highest carbohydrate content of 61.5% in biogas slurry and the highest ammonia removal efficiency and rate of 96.3% and 91.7mg/L/d respectively in biogas slurry with phosphorus and magnesium added. Additionally, the absence of phosphorus and magnesium that can be adverse for biomass accumulation resulted in earlier timing of carbohydrate accumulation and magnesium was firstly recognized and proved as the influence factor for carbohydrate accumulation. Microalgae that cultured in biogas slurry accumulated more carbohydrate in cell, making biogas slurry more suitable medium for the improvement of carbohydrate content, thus can be regarded as a new strategy to accumulate carbohydrate.
Assuntos
Biocombustíveis/microbiologia , Carboidratos/química , Chlorella vulgaris , Amônia/química , Biomassa , Microalgas , Nitrogênio/química , Fósforo/químicaRESUMO
Genetic factors increase the risk to coronary artery disease (CAD). To date, a limited number of genes that potentially contribute to development of CAD have been identified. In this study, we have performed large-scale gene expression analysis of approximately 12,000 human genes in nine severely atherosclerotic and six nonatherosclerotic human coronary arteries using oligonucleotide microarrays. Fifty-six genes showed differential expression in atherosclerotic coronary artery tissues; expression of 55 genes was increased in atherosclerotic coronary arteries, whereas only one gene, GST, encoding a reducing agent, showed downregulated expression. The expression data of selected genes were validated by quantitative RT-PCR analysis as well as immunostaining. The associations of 49 genes with CAD appear to be novel, and they include genes encoding ICAM-2, PIM-2, ECGF1, fusin, B cell activator (BL34, GOS8), Rho GTPase activating protein-4, retinoic acid receptor responder, beta2-arrestin, membrane aminopeptidase, cathepsins K and H, MIR-7, TNF-alpha-induced protein 2 (B94), and flavocytochrome 558. In conclusion, we have identified 56 genes whose expression is associated with CAD, and 49 of them may represent new genes linked to CAD.
Assuntos
Doença da Artéria Coronariana/genética , Perfilação da Expressão Gênica , Apoptose/genética , Adesão Celular/genética , Divisão Celular/genética , Vasos Coronários , Matriz Extracelular/metabolismo , Imunofluorescência , Regulação da Expressão Gênica , Humanos , Inflamação/genética , Metabolismo dos Lipídeos , Necrose , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: The aim of this study was to evaluate how CYP2C19 affects icotinib and metabolite' exposure, and to determine whether the exposure and EGFR genotype influences survival time, tumor metastasis and adverse drug reactions. MATERIALS AND METHODS: 274 NSCLC patients who accepted 125 mg icotinib/t.i.d. were chosen from a phase III study. Blood samples were obtained in 672 nd (4th week) and 1,680 th hours (10th week), and plasma was used to quantify the concentration of icotinib and blood cells were sampled to check the genotypes. Clinical data were also collected at the same time, including EGFR genotypes. Plasma concentrations were assessed by HPLC-MS/MS and genotype by sequencing. All data were analyzed through SPSS 17.0 and SAS 9.2. RESULTS: CYP 2C19 genotypes affected bio-transformation from icotinib to M24 and M26, especially in poor-metabolisers. Higher icotinib concentrations (>1000 ng/mL) not only increased patient PFS and OS but also reduced tumor metastasis. Patients with mutant EGFR experienced a higher median PFS and OS (234 and 627 days), especially those with the 19del genotype demonstrating higher PR ratio. Patients who suffered grade II skin toxicity had a higher icotinib exposure than those with grade I skin toxicity or no adverse effects. Liver toxic reactions might occur in patients with greater M20 and M23 plasma concentrations. CONCLUSIONS: CYP2C19 polymorphisms significantly affect icotinib, M24 and M26 exposure. Patients with mutant EGFR genotype and higher icotinib concentration might have increased PFS and OS and lower tumor metastasis. Liver ADR events and serious skin effects might be respectively induced by greater M20, M23 and icotinib concentrations.
Assuntos
Antineoplásicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Éteres de Coroa/uso terapêutico , Citocromo P-450 CYP2C19/genética , Genes erbB-1/genética , Neoplasias Pulmonares/tratamento farmacológico , Quinazolinas/uso terapêutico , Adulto , Idoso , Antineoplásicos/metabolismo , Povo Asiático/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Doença Hepática Induzida por Substâncias e Drogas/genética , China , Cromatografia Líquida de Alta Pressão , Éteres de Coroa/metabolismo , Citocromo P-450 CYP2C19/metabolismo , Intervalo Livre de Doença , Toxidermias/etiologia , Toxidermias/genética , Feminino , Genótipo , Humanos , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Quinazolinas/metabolismo , Espectrometria de Massas em TandemRESUMO
The present study was to synthesize a novel multi-targeted kinase inhibitor and evaluated its anticancer effects on a hepatocellular carcinoma xenograft model. In our study, in vivo efficacy was determined in nude mice bearing HuH7 human HCC xenografts. The mice were randomly divided into the following five groups with the use of a randomization chart (n = 8 in each group): high-dose BZG-4000 group, medium-dose BZG-4000 group, low-dose BZG-4000 group, sorafenib group, and model group. Tumor size measurements included the length (L) and width (W) measured with calipers, and tumor volume was calculated as (LWâ§2)/2. Tumor tissues slides were hematoxylin and eosin (HE) stained for histopathological examination. Immunohistochemistry detected CD31 expression, and Western blotting measured VEGF protein expression. We found that when BZG-4000 was administered orally to xenograft HuH7 nude mice, tumor growth was inhibited and significant tumor shrinkage was evident. After oral administration of BZG-4000 at 40â mg/kg/day, the tumor weight and volume were significantly lower than tumors of the sorafenib group. BZG-4000 considerably decreased the expression of CD31 and VEGF in tumors compared to tumors treated with positive control drug. It was concluded that BZG-4000 has the potential to inhibit the tumorigenesis of hepatocellular carcinoma in vivo by decreasing the expression of CD31 and VEGF.
Assuntos
Inibidores da Angiogênese/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Neovascularização Patológica/prevenção & controle , Compostos de Fenilureia/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Administração Oral , Inibidores da Angiogênese/síntese química , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Esquema de Medicação , Expressão Gênica , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Niacinamida/análogos & derivados , Niacinamida/farmacologia , Compostos de Fenilureia/síntese química , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Inibidores de Proteínas Quinases/síntese química , Sorafenibe , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Through nearly 50 years of development, sulphur ylides have been established as versatile and powerful reagents for the construction of carbocycles and heterocycles. Despite advances, two important and yet elusive bottlenecks continue to inhibit the advancement of this chemistry: a limited number of reagents with polar groups to react with sulphur ylides, and the wide utilization of chiral auxiliaries or substrates to achieve asymmetric cycloaddition processes in the majority of known reports. Herein, we apply an asymmetric palladium catalysis strategy to the chemistry of sulphur ylides to address these two fundamental problems. We thus achieve an unprecedented decarboxylation-cycloaddition sequence of cyclic allylic esters with sulphur ylides through the enantioselective trapping of Pd-stabilized zwitterionic intermediates by the ylides. As a result, a series of biologically and synthetically important 3-vinyl indolines are rapidly assembled with a high reaction efficiency and stereoselectivity.
RESUMO
von Willebrand factor (VWF) tethers platelets to sites of vascular injury via interaction with the platelet surface receptor, GPIb. To further define the VWF sequences required for VWF-platelet interaction, a phage library displaying random VWF protein fragments was screened against formalin-fixed platelets. After 3 rounds of affinity selection, DNA sequencing of platelet-bound clones identified VWF peptides mapping exclusively to the A1 domain. Aligning these sequences defined a minimal, overlapping segment spanning P1254-A1461, which encompasses the C1272-C1458 cystine loop. Analysis of phage carrying a mutated A1 segment (C1272/1458A) confirmed the requirement of the cystine loop for optimal binding. Four rounds of affinity maturation of a randomly mutagenized A1 phage library identified 10 and 14 unique mutants associated with enhanced platelet binding in the presence and absence of botrocetin, respectively, with 2 mutants (S1370G and I1372V) common to both conditions. These results demonstrate the utility of filamentous phage for studying VWF protein structure-function and identify a minimal, contiguous peptide that bind to formalin-fixed platelets, confirming the importance of the VWF A1 domain with no evidence for another independently platelet-binding segment within VWF. These findings also point to key structural elements within the A1 domain that regulate VWF-platelet adhesion.