RESUMO
In cryogenic electron microscopy (cryo-EM) single particle analysis (SPA), high-resolution three-dimensional structures of biological macromolecules are determined by iteratively aligning and averaging a large number of two-dimensional projections of molecules. Since the correlation measures are sensitive to the signal-to-noise ratio, various parameter estimation steps in SPA will be disturbed by the high-intensity noise in cryo-EM. However, denoising algorithms tend to damage high frequencies and suppress mid- and high-frequency contrast of micrographs, which exactly the precise parameter estimation relies on, therefore, limiting their application in SPA. In this study, we suggest combining a cryo-EM image processing pipeline with denoising and maximizing the signal's contribution in various parameter estimation steps. To solve the inherent flaws of denoising algorithms, we design an algorithm named MScale to correct the amplitude distortion caused by denoising and propose a new orientation determination strategy to compensate for the high-frequency loss. In the experiments on several real datasets, the denoised particles are successfully applied in the class assignment estimation and orientation determination tasks, ultimately enhancing the quality of biomacromolecule reconstruction. The case study on classification indicates that our strategy not only improves the resolution of difficult classes (up to 5 Å) but also resolves an additional class. In the case study on orientation determination, our strategy improves the resolution of the final reconstructed density map by 0.34 Å compared with conventional strategy. The code is available at https://github.com/zhanghui186/Mscale.
Assuntos
Processamento de Imagem Assistida por Computador , Imagem Individual de Molécula , Microscopia Crioeletrônica/métodos , Processamento de Imagem Assistida por Computador/métodos , Algoritmos , Razão Sinal-RuídoRESUMO
MOTIVATION: Cryo-electron microscopy (cryo-EM) is a widely used technology for ultrastructure determination, which constructs the 3D structures of protein and macromolecular complex from a set of 2D micrographs. However, limited by the electron beam dose, the micrographs in cryo-EM generally suffer from the extremely low signal-to-noise ratio (SNR), which hampers the efficiency and effectiveness of downstream analysis. Especially, the noise in cryo-EM is not simple additive or multiplicative noise whose statistical characteristics are quite different from the ones in natural image, extremely shackling the performance of conventional denoising methods. RESULTS: Here, we introduce the Noise-Transfer2Clean (NT2C), a denoising deep neural network (DNN) for cryo-EM to enhance image contrast and restore specimen signal, whose main idea is to improve the denoising performance by correctly learning the noise distribution of cryo-EM images and transferring the statistical nature of noise into the denoiser. Especially, to cope with the complex noise model in cryo-EM, we design a contrast-guided noise and signal re-weighted algorithm to achieve clean-noisy data synthesis and data augmentation, making our method authentically achieve signal restoration based on noise's true properties. Our work verifies the feasibility of denoising based on mining the complex cryo-EM noise patterns directly from the noise patches. Comprehensive experimental results on simulated datasets and real datasets show that NT2C achieved a notable improvement in image denoising, especially in background noise removal, compared with the commonly used methods. Moreover, a case study on the real dataset demonstrates that NT2C can greatly alleviate the obstacles caused by the SNR to particle picking and simplify the identifying of particles. AVAILABILITYAND IMPLEMENTATION: The code is available at https://github.com/Lihongjia-ict/NoiseTransfer2Clean/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
Algoritmos , Redes Neurais de Computação , Microscopia Crioeletrônica/métodos , Razão Sinal-Ruído , Proteínas , Processamento de Imagem Assistida por Computador/métodosRESUMO
INTRODUCTION: Bronchoscopic navigation combined with endobronchial ultrasound-guided transbronchial lung biopsy (EBUS-TBLB) is an important approach for the diagnosis of peripheral pulmonary lesions (PPL). The fifth-generation (5G) network, characterized by low latency and high stability, has shown promising possibilities in telemedicine for remote areas. METHODS: We present two cases of PPL in primary hospitals without navigation equipment. The EBUS-TBLB was performed with the guidance of remote augmented reality virtual bronchoscopic navigation (VBN) based on a 5G network. In practice, the 5G network could enable the matching of actual/virtual bronchoscopic images and navigation paths in real time, as well as high-speed transmission at long distances (>20 km), without any visual delay (<500 ms). CONCLUSION: Both patients were successfully diagnosed with lung cancer after accurate positioning and obtaining biopsies of target lesions. This 5G-based remote VBN-guided EBUS-TBLB appears to be safe with reliable connections in both cases and shows potential for cost-effectiveness. It would be an optimal resource for undeveloped regions and/or regions lacking endoscopists with extensive experience in navigation-related procedures.
Assuntos
Neoplasias Pulmonares , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/patologia , Broncoscopia/métodos , Biópsia/métodos , Pulmão/diagnóstico por imagem , Pulmão/patologia , Biópsia Guiada por Imagem/métodos , Ultrassonografia , Endossonografia/métodosRESUMO
Robotic platforms have been widely used in the various fields of clinical diagnosis and therapy of diseases in the past decade. Robotic-assisted bronchoscopy (RAB) demonstrates its advantages of visibility, flexibility, and stability in comparison to conventional bronchoscopic techniques. Improving diagnostic yield and navigation yield for peripheral pulmonary lesions has been defined; however, RAB platform of treatment was not reported. In this article, we report a case of a 52-year-old woman who was diagnosed with the tracheal adenoid cystic carcinoma and recurred in the second postoperative year, leading to the involvement of the entire tracheal wall and lumen obstruction. Since the lesion was inoperable, we combined RAB and photodynamic therapy (PDT) for the patient. The potential advantages of using RAB for PDT delivery include precise light irradiation of target lesions and stable intra-operative control over the long term. This is a novel application of RAB combined with PDT for airway diseases. The case report may provide a new insight into the diagnosis and treatment of pulmonary diseases. In addition to improving the diagnostic rates, the RAB platform may also play an important role in the treatment of airway and lung disease in the future.
Assuntos
Carcinoma Adenoide Cístico , Pneumopatias , Fotoquimioterapia , Procedimentos Cirúrgicos Robóticos , Neoplasias da Traqueia , Feminino , Humanos , Pessoa de Meia-Idade , Traqueia/cirurgia , Traqueia/patologia , Broncoscopia/métodos , Carcinoma Adenoide Cístico/diagnóstico , Carcinoma Adenoide Cístico/tratamento farmacológico , Recidiva Local de Neoplasia/patologia , Neoplasias da Traqueia/diagnóstico , Neoplasias da Traqueia/tratamento farmacológico , Pneumopatias/patologiaRESUMO
Whipworms are soil-transmitted helminths that can infect a variety of animals. A Chinese serow possibly infected by whipworms was found during a wildlife disease surveillance project in Baima Snow Mountain National Nature Reserve, Yunnan, China, in 2021. As convergent evolution is common in coinhabiting parasites, a mitochondrial gene sequence (cox1) and ribosomal gene sequence (ITS1) were used to identify species similar to Trichuris from Chinese serow. The phylogenetic trees and genetic distances of ten Trichuris samples from a Chinese serow together with other Trichuris spp. that have been previously reported were analysed based on the cox1 and ITS1 sequences. The combined results of the phylogenetic tree and genetic distances based on cox1 gene showed that the whipworms in Chinese serow are T. skrjabini. However, the whipworms in the present study were divided into two apparent clades in the phylogenic trees constructed by the cox1 sequences (Clades A and B) and the ITS1 sequences (Clades C and D). In addition, the Fst and Nm values were 0.82 and 0.23 between Clade A and Clade B for the cox1 gene, and 0.30 and 0.45 between Clade C and Clade D for the ITS1 sequences; both indicators showed low gene flow among the clades. Therefore, the genetic population structure of T. skrjabini was illustrated.
Assuntos
Tricuríase , Trichuris , Animais , Sequência de Bases , China/epidemiologia , Filogenia , Análise de Sequência de DNA , Tricuríase/epidemiologia , Tricuríase/veterináriaRESUMO
Scabies is a common parasitic disease in many mammalian species, caused by the infestation of Sarcoptes scabiei. There is no consistent conclusion on whether Sarcoptes mites from different hosts or geographic locations have apparent genetic divergence. In this study, we collected and morphologically identified S. scabiei from Chinese serow and goral, and we described the genetic diversity of S. scabiei and other mites based on phylogenetic analyses of the ITS2 and cox1 sequence fragments, including data available in GenBank. The mites isolated from Chinese serow and goral were S. scabiei, and they were morphologically similar. The phylogenetic trees and haplotype networks showed that S. scabiei from other locations worldwide did not cluster according to host divergence or geographical distribution. Additionally, the Fst values were - 0.224 to 0.136 and - 0.045 to 1 between S. scabiei from different hosts, including humans and domestic and wild animals, based on partial ITS and cox1 sequences. Worldwide S. scabiei samples formed three clusters (with H2, H5, and H12 at their centers) in the ITS and one cluster (with C9 at the center) in the cox1 haplotype phylogenetic network. The S. scabiei collected from Chinese serow and goral were morphologically similar and had the same genotype. A study on the genetic characteristics of S. scabiei from Chinese serow and goral together with other mites from different hosts and geographic locations around the world showed no obvious divergence. These findings indicated that scabies likely is a zoonotic disease and that the global prevalence of scabies is probably related to the worldwide trade of domestic animals.
Assuntos
Sarcoptes scabiei , Escabiose , Animais , Humanos , Sarcoptes scabiei/genética , Escabiose/epidemiologia , Escabiose/veterinária , Escabiose/parasitologia , Filogenia , Ruminantes , ChinaRESUMO
BACKGROUND/AIMS: Although some studies showed that HIF-2α expression was correlated with an unfavorable prognosis in colorectal cancer (CRC), the prognostic results remain conflicting in CRC. The present study was performed to evaluate the association between HIF-2α expression and the clinicopathological features of this disease and to examine the potential prognostic role of HIF-2α expression in CRC. METHODS: Pooled odds ratios (ORs) or hazard ratios (HRs) were calculated from available publications, The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) datasets. Trial sequential analysis (TSA) was used to estimate the required sample information. RESULTS: HIF-2α protein expression was more frequent in CRC than in normal colonic tissues (OR = 150.49, P < 0.001), higher in male than female CRC patients (OR = 1.47, P = 0.008), and lower in high-grade than low-grade CRC (OR = 0.49, P = 0.029). TSA verified the reliability of the above results. HIF-2α expression was not linked to the prognosis of CRC in overall survival (OS), disease-specific survival (DSS), metastasis-free survival, and relapse-free survival, and no significant correlation was found between HIF-2α alteration and OS or disease-free survival (DFS) of CRC. Expression of both HIF-2α and vascular endothelial growth factor (VEGFA, VEGFB, or VEGFC) was associated with a poor metastasis-free survival of CRC (HR = 6.95, HR = 113.51, and HR = 8.11, respectively). No association was observed between HIF-2α expression and DFS in other cancers, but HIF-2α expression was correlated with a worse DFS of CRC (HR = 1.23, P = 0.037). Moreover, HIF-2α expression was linked to a good survival benefit in some cancers (B-cell lymphoma and lung adenocarcinoma: OS, multiple myeloma: DSS, breast cancer: distant metastasis-free survival, liposarcoma: distant recurrence-free survival) (all HRs < 1, Ps < 0.05). CONCLUSIONS: HIF-2α expression may be associated with the carcinogenesis of CRC, which is higher in males than in females, negatively linked to tumor differentiation, and correlated with a worse DFS of CRC. Additional prospective studies are needed.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias Colorretais/patologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Bases de Dados Genéticas , Intervalo Livre de Doença , Humanos , Estimativa de Kaplan-Meier , Recidiva Local de Neoplasia , Razão de Chances , Prognóstico , Modelos de Riscos Proporcionais , Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
The tumour hypoxia would trigger the angiogenesis switch for survival, and increase the ability of cancer cells to invade and metastasis. However, hypoxia regulated genes that invovled in angiogenesis in colorectal cancer (CRC) have not been explored in detail. The aim of this study was to explore angiogenic genes under hypoxia condition in CRC. Here, we found that endothelial cells tube formation and cancer cells invasion ability were promoted even under chronic hypoxia condition (72 h) in colon adenocarcinoma HCT-116 cells. Then, we explored the differentially expressed genes (DEGs) under chronic hypoxia condition by microarray from Gene Expression Omnibus (GEO) database. Subsequent bioinformatic analysis identified 17 genes that invovled in angiogenesis, blood vessel development, blood vessel morphgensis, vascular development. of these genes, VEGF-A, Smad7, Jun, IL-8, CXCR-4, PDGF-A, TGF-A, ANGPTL-4 expression levels up-regulated under hypoxia condition. Additionally, the gene expression level in acute hypoxia (24 h) was significantly higher than chronic condition (72 h). Finally, knockdown of hypoxia inducible factor (HIF-1α) by shRNA reversed the role of Smad7, CXCR-4, PDGF-A, TGF-A and ANGPTL-4 overexpression in HCT-116 cells, these findings provide the potential angiogenic targets for the treatment of colorectal cancer.
Assuntos
Proteínas Angiogênicas/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas de Neoplasias/metabolismo , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Hipóxia Tumoral , Doença Aguda , Doença Crônica , Neoplasias Colorretais/complicações , Perfilação da Expressão Gênica , Genes Neoplásicos , Humanos , Invasividade Neoplásica , Neovascularização Patológica/complicaçõesRESUMO
BACKGROUND: Yi Ai Fang (YAF), a traditional Chinese medicine (TCM) formula, has been identified to have anticancer activity in our previously studies. The present study aimed to explore the potential mechanism of YAF suppression of VM on colorectal cancer (CRC) in vitro and in vivo. METHODS: Cell viability was measured by CCK-8 assay. HIF-1α, E-cd(E-cadherin), Claudin-4, and VIM (Vimentin) expressions level in vitro were evaluated by Western blot or RT-PCR. In addition, Human CRC HCT-116 cells were implanted in BALB/c nude mice; mice with xenografted tumors were randomly administrated vehicle (control), 8, 16, or 32 mg/mL YAF, or 1 mg/mL fluorouracil (5-FU). HIF-1α, E-cd, Claudin-4, and VIM expression in these tumors were determined by IHC. RESULTS: YAF effectively inhibited the growth and the formation of vasculogenic mimicry (VM) of CRC cells in a dose-dependent trend. YAF restrained the formation of vasculogenic mimicry(VM) through HIF-1α/EMT pathway in CRC. YAF suppressed VM was triggered by activation of E-cd and Claudin-4,which are characteristics of endothelial cells,and inhibition of HIF-1α and VIM in vitro. In vivo data showed that YAF remarkably inhibited growth of the xenografted tumors. The YAF-treated tumor samples were analyzed by IHC for levels of HIF-1α/EMT related proteins HIF-1α, E-cd, Claudin-4, and VIM. The results indicated that YAF significantly enhanced expression of E-cd and Claudin-4,but decreased expression of HIF-1α, VIM in a dose-dependent manner. CONCLUSIONS: In conclusion, this study provided the first direct evidence that YAF inhibited the formation of VM in human CRC, suggesting that YAF may be considered as a useful target for cancer therapy.
Assuntos
Neoplasias Colorretais/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Células HCT116 , Humanos , Camundongos , Camundongos Nus , Neovascularização Patológica/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The hemicellulose 4-O-methyl glucuronoxylan is one of the principle components present in the secondary cell walls of eudicotyledonous plants. However, the biochemical mechanisms leading to the formation of this polysaccharide and the effects of modulating its structure on the physical properties of the cell wall are poorly understood. We have identified and functionally characterized an Arabidopsis glucuronoxylan methyltransferase (GXMT) that catalyzes 4-O-methylation of the glucuronic acid substituents of this polysaccharide. AtGXMT1, which was previously classified as a domain of unknown function (DUF) 579 protein, specifically transfers the methyl group from S-adenosyl-L-methionine to O-4 of α-D-glucopyranosyluronic acid residues that are linked to O-2 of the xylan backbone. Biochemical characterization of the recombinant enzyme indicates that GXMT1 is localized in the Golgi apparatus and requires Co(2+) for optimal activity in vitro. Plants lacking GXMT1 synthesize glucuronoxylan in which the degree of 4-O-methylation is reduced by 75%. This result is correlated to a change in lignin monomer composition and an increase in glucuronoxylan release during hydrothermal treatment of secondary cell walls. We propose that the DUF579 proteins constitute a previously undescribed family of cation-dependent, polysaccharide-specific O-methyl-transferases. This knowledge provides new opportunities to selectively manipulate polysaccharide O-methylation and extends the portfolio of structural targets that can be modified either alone or in combination to modulate biopolymer interactions in the plant cell wall.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Ácido Glucurônico/metabolismo , Metiltransferases/metabolismo , Xilanos/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Catálise , Cátions/metabolismo , Parede Celular/enzimologia , Éteres/metabolismo , Complexo de Golgi/metabolismo , Lignina/metabolismo , Metilação , Metiltransferases/química , Metiltransferases/genética , Mutagênese/fisiologia , Polissacarídeos/metabolismo , Estrutura Terciária de Proteína/fisiologia , Xilanos/biossínteseRESUMO
In dilute acid pretreatment of lignocellulosic biomass, lignin has been shown to form droplets that deposit on the cellulose surface and retard enzymatic digestion of cellulose (Donohoe et al., 2008; Selig et al., 2007). However, studies of this nature are limited for hydrothermal pretreatment, with the result that the corresponding mechanisms that inhibit cellulosic enzymes are not well understood. In this study, scanning electron microscope (SEM) and wet chemical analysis of solids formed by hydrothermal pretreatment of a mixture of Avicel cellulose and poplar wood showed that lignin droplets from poplar wood relocated onto the Avicel surface. In addition, nuclear magnetic resonance (NMR) showed higher S/G ratios in deposited lignin than the initial lignin in poplar wood. Furthermore, the lignin droplets deposited on Avicel significantly impeded cellulose hydrolysis. A series of tests confirmed that blockage of the cellulose surface by lignin droplets was the main cause of cellulase inhibition. The results give new insights into the fate of lignin in hydrothermal pretreatment and its effects on enzymatic hydrolysis.
Assuntos
Celulases/metabolismo , Celulose/química , Celulose/metabolismo , Lignina/química , Lignina/metabolismo , Celulose/efeitos da radiação , Celulose/ultraestrutura , Temperatura Alta , Hidrólise , Lignina/efeitos da radiação , Lignina/ultraestrutura , Espectroscopia de Ressonância Magnética , Microscopia Eletrônica de Varredura , Populus , MadeiraRESUMO
The content of the asarone submicro emulsion injection was determind by HPLC method, and thereby a quality evaluation method was established based on indexes of pH value, particle size, peroxide value, methoxy aniline values, free fatty acid, lysophosphatidylcholine, visible foreign substances, insoluble particle, sterility, bacterial endotoxin and impurities, etc. The results showed that the injection exhibited uniform physical appearance and all the products were in milkwhite liquid. The content of the three batches products were respectively 102.9%, 100.8%, 97.70% of the labeled amount, with mean particle size of 210-250 nm, and other indexes all met with the standards. The reserved samples showed no obvious change in terms of detection indexes and indicated good stability after the accelerated stability test and long-term stability for 12 months. The quality evaluation method established in this study could be applied to quality control and stability investigation of asarone submicron emulsion injection, which laid a basis for further clinical research and application.
Assuntos
Anisóis/química , Medicamentos de Ervas Chinesas/química , Emulsões/química , Derivados de Alilbenzenos , Cromatografia Líquida de Alta Pressão , Estabilidade de Medicamentos , Tamanho da Partícula , Controle de QualidadeRESUMO
Cryo-electron microscopy (cryo-EM) has emerged as a potent technique for determining the structure and functionality of biological macromolecules. However, limited by the physical imaging conditions, such as low electron beam dose, micrographs in cryo-EM typically contend with an extremely low signal-to-noise ratio (SNR), impeding the efficiency and efficacy of subsequent analyses. Therefore, there is a growing demand for an efficient denoising algorithm designed for cryo-EM micrographs, aiming to enhance the quality of macromolecular analysis. However, owing to the absence of a comprehensive and well-defined dataset with ground truth images, supervised image denoising methods exhibit limited generalization when applied to experimental micrographs. To tackle this challenge, we introduce a simulation-aware image denoising (SaID) pretrained model designed to enhance the SNR of cryo-EM micrographs where the training is solely based on an accurately simulated dataset. First, we propose a parameter calibration algorithm for simulated dataset generation, aiming to align simulation parameters with those of experimental micrographs. Second, leveraging the accurately simulated dataset, we propose to train a deep general denoising model that can well generalize to real experimental cryo-EM micrographs. Comprehensive experimental results demonstrate that our pretrained denoising model achieves excellent denoising performance on experimental cryo-EM micrographs, significantly streamlining downstream analysis.
Assuntos
Algoritmos , Microscopia Crioeletrônica , Processamento de Imagem Assistida por Computador , Razão Sinal-Ruído , Microscopia Crioeletrônica/métodos , Processamento de Imagem Assistida por Computador/métodos , Simulação por ComputadorRESUMO
Cryoelectron tomography (cryo-ET) has become an indispensable technology for visualizing in situ biological ultrastructures, where the tilt series alignment is the key step to obtain a high-resolution three-dimensional reconstruction. Specifically, with the advent of high-throughput cryo-ET data collection, there is an increasing demand for high-accuracy and fully automatic tilt series alignment, to enable efficient data processing. Here, we propose Markerauto2, a fast and robust fully automatic software that enables accurate fiducial marker-based tilt series alignment. Markerauto2 implements the following novel pipelines: (1) an accelerated high-precision fiducial marker detection with wavelet multiscale template, (2) an ultra-fast and robust fiducial marker tracking supported by hashed geometric features, (3) a high-angle fiducial marker supplementation strategy to produce more complete tracks, and (4) a precise and robust calibration of projection parameters with group-weighted parameter optimization. Comprehensive experiments conducted on both simulated and real-world datasets demonstrate the robustness, efficiency, and effectiveness of the proposed software.
RESUMO
Slow transit constipation (STC) is a long-lasting and prevalent intestinal condition, marked by hard, dry feces. The primary cause of STC may be attributed to an imbalance in the gut's microbial community and alterations in its metabolic byproducts. Tongbian formula (TB), a traditional Chinese medicinal formula, has been used to treat STC and shows a great effect on relieving constipation. The role of TB in regulating intestinal microbiota has not been fully elucidated. Herein, we investigated the potential effect of TB on gut microbiota and further explored the potential mechanism behind its effects. Our study demonstrated that TB significantly increased fecal water content and intestinal ink propulsion rate in loperamide (Lope)-induced STC rats. 5-HT signaling was suppressed in STC colon tissue, and the abundance of butyric acid (BA) in colonic contents was significantly down-regulated after Lope treatment. Notably, TB administration led to the restoration of microbial dysbiosis and the up-regulation of BA content, subsequently activating 5-HT signaling pathways. When BA was combined with a tryptophan hydroxylase-1 (TPH1) inhibitor, which is crucial for 5-HT synthesis, its therapeutic efficacy for treating STC was compromised. TB alleviates STC by reversing the intestinal microbiota imbalance and activating the 5-HT signaling in the colon through increasing BA levels. These findings suggest that TB is an ideal candidate for STC treatment.
Assuntos
Ácido Butírico , Constipação Intestinal , Medicamentos de Ervas Chinesas , Microbioma Gastrointestinal , Serotonina , Transdução de Sinais , Constipação Intestinal/tratamento farmacológico , Constipação Intestinal/metabolismo , Animais , Ácido Butírico/farmacologia , Transdução de Sinais/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Ratos , Serotonina/metabolismo , Masculino , Ratos Sprague-Dawley , Trânsito Gastrointestinal/efeitos dos fármacos , Loperamida , Modelos Animais de Doenças , Colo/metabolismo , Colo/efeitos dos fármacosRESUMO
Immune checkpoint blockade (ICB) therapies function by alleviating immunosuppression on tumor-infiltrating lymphocytes (TILs) but are often insufficient to fully reactivate these dysfunctional TILs. Although interleukin 12 (IL-12) has been used in combination with ICB to improve efficacy, this remains limited by severe toxicity associated with systemic administration of this cytokine. Here, we engineer a fusion protein composed of an anti-PD-1 antibody and a mouse low-affinity IL-12 mutant-2 (αPD1-mIL12mut2). Systemic administration of αPD1-mIL12mut2 displays robust antitumor activities with undetectable toxicity. Mechanistically, αPD1-mIL12mut2 preferentially activates tumor-infiltrating PD-1+CD8+T cells via high-affinity αPD-1 mediated cis-binding of low-affinity IL-12. Additionally, αPD1-mIL12mut2 treatment exerts an abscopal effect to suppress distal tumors, as well as metastasis. Collectively, αPD1-mIL12mut2 treatment induces robust systemic antitumor responses with reduced side effects.
Assuntos
Linfócitos T CD8-Positivos , Interleucina-12 , Linfócitos do Interstício Tumoral , Receptor de Morte Celular Programada 1 , Animais , Interleucina-12/metabolismo , Interleucina-12/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Receptor de Morte Celular Programada 1/metabolismo , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Camundongos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Linhagem Celular Tumoral , Feminino , Inibidores de Checkpoint Imunológico/farmacologia , Humanos , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/genéticaRESUMO
Background: Escherichia coli is an important intestinal flora, of which pathogenic E. coli is capable of causing many enteric and extra-intestinal diseases. Antibiotics are essential for the treatment of bacterial infections caused by pathogenic E. coli; however, with the widespread use of antibiotics, drug resistance in E. coli has become particularly serious, posing a global threat to human, animal, and environmental health. While the drug resistance and pathogenicity of E. coli carried by tigers and leopards in captivity have been studied intensively in recent years, there is an extreme lack of information on E. coli in these top predators in the wild environment. Methods: Whole genome sequencing data of 32 E. coli strains collected from the feces of wild Amur tiger (Panthera tigris altaica, n = 24) and North China leopard (Panthera pardus japonensis, n = 8) were analyzed in this article. The multi-locus sequence types, serotypes, virulence and resistance genotypes, plasmid replicon types, and core genomic SNPs phylogeny of these isolates were studied. Additionally, antimicrobial susceptibility testing (AST) was performed on these E. coli isolates. Results: Among the E. coli isolates studied, 18 different sequence types were identified, with ST939 (21.9%), ST10 (15.6%), and ST3246 (9.4%) being the most prevalent. A total of 111 virulence genes were detected, averaging about 54 virulence genes per sample. They contribute to invasion, adherence, immune evasion, efflux pump, toxin, motility, stress adaption, and other virulence-related functions of E. coli. Sixty-eight AMR genes and point mutations were identified. Among the detected resistance genes, those belonging to the efflux pump family were the most abundant. Thirty-two E. coli isolates showed the highest rate of resistance to tetracycline (14/32; 43.8%), followed by imipenem (4/32; 12.5%), ciprofloxacin (3/32; 9.4%), doxycycline (2/32; 6.3%), and norfloxacin (1/32; 3.1%). Conclusions: Our results suggest that E. coli isolates carried by wild Amur tigers and North China leopards have potential pathogenicity and drug resistance.
Assuntos
Escherichia coli , Fezes , Panthera , Tigres , Sequenciamento Completo do Genoma , Animais , Tigres/microbiologia , Escherichia coli/genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/patogenicidade , Escherichia coli/isolamento & purificação , Panthera/microbiologia , Fezes/microbiologia , Infecções por Escherichia coli/veterinária , Infecções por Escherichia coli/microbiologia , Filogenia , Antibacterianos/farmacologia , Genoma Bacteriano/genética , Testes de Sensibilidade Microbiana , China , Virulência/genética , Farmacorresistência Bacteriana/genética , Polimorfismo de Nucleotídeo Único/genética , Tipagem de Sequências MultilocusRESUMO
Because conventional approaches for evaluating sugar release from the coupled operations of pretreatment and enzymatic hydrolysis are extremely time and material intensive, high throughput (HT) pretreatment and enzymatic hydrolysis systems have become vital for screening large numbers of lignocellulosic biomass samples to identify feedstocks and/or processing conditions that significantly improve performance and lower costs. Because dilute acid pretreatment offers many important advantages in rendering biomass highly susceptible to subsequent enzymatic hydrolysis, a high throughput pretreatment and co-hydrolysis (HTPH) approach was extended to employ dilute acid as a tool to screen for enhanced performance. First, a single-step neutralization and buffering method was developed to allow effective enzymatic hydrolysis of the whole pretreated slurry. Switchgrass and poplar were then pretreated with 0.5% and 1% acid loadings at a 5% solids concentration, the resulting slurry conditioned with the buffering approach, and the entire mixture enzymatically hydrolyzed. The resulting sugar yields demonstrated that single-step neutralizing and buffering was capable of adjusting the pH as needed for enzymatic saccharification, as well as overcoming enzyme inhibition by compounds released in pretreatment. In addition, the effects of pretreatment conditions and biomass types on susceptibility of pretreated substrates to enzymatic conversion were clearly discernible, demonstrating the method to be a useful extension of HTPH systems.
Assuntos
Ácidos/metabolismo , Biomassa , Biotecnologia/métodos , Ensaios de Triagem em Larga Escala/métodos , Lignina/metabolismo , Hidrólise , Panicum/efeitos dos fármacos , Populus/efeitos dos fármacosRESUMO
High throughput pretreatment (HTPH) and enzymatic hydrolysis systems are now vital for screening large numbers of biomass samples to investigate biomass recalcitrance over various pretreatment and enzymatic hydrolysis conditions. Although hydrothermal pretreatment is currently being employed in most high throughput applications, thermochemical pretreatment at low and high pH conditions can offer additional insights to better understand the roles of hemicellulose and lignin, respectively, in defining biomass recalcitrance. Thus, after successfully applying the HTPH approach to dilute acid pretreatment [Gao et al. (2012) Biotechnol. Bioeng. 110(3): 754-762], extension to dilute alkali pretreatment was also achieved using a similar single-step neutralization and buffering concept. In the latter approach, poplar and switchgrass were pretreated with 1 wt% sodium hydroxide at 120°C for different reaction times. Following pretreatment, an H2Citâ»/HCit²â» buffer with a pH of 4.5 was used to condition the pretreatment slurry to a pH range of 4.69-4.89, followed by enzymatic hydrolysis for 72 h of the entire mixture. Sugar yields showed different trends for poplar and switchgrass with increases in pretreatment times, demonstrating the method provided a clearly discernible screening tool at alkali conditions. This method was then applied to selected Populus tremuloides samples to follow ring-by-ring sugar release patterns. Observed variations were compared to results from hydrothermal pretreatments, providing new insights in understanding the influence of biomass structural differences on recalcitrance.
Assuntos
Álcalis/metabolismo , Hidroliases/metabolismo , Lignina/metabolismo , Polissacarídeos/metabolismo , Carboidratos/análise , Temperatura Alta , Hidrólise , Panicum/metabolismo , Populus/metabolismoRESUMO
Xylooligosaccharides released in hydrothermal pretreatment of lignocellulosic biomass can be purified for high-value products or further hydrolyzed into sugars for fermentation or chemical conversion. In addition, characterization of xylooligosaccharides is vital to understand hemicellulose structure and removal mechanisms in pretreatment of cellulosic biomass. In this study, gel permeation chromatography was applied to fractionate xylooligosaccharides produced from birchwood xylan according to their specific degree of polymerization (DP). Then, each fraction was identified by high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS); and their concentrations were determined by a downscaled post-hydrolysis method. Based on PAD responses and sugar concentrations for each fraction, a series of response factors were developed that can be used to quantify xylooligosaccharides of DP from 2 to 14 without standards. The resulting approach can profile xylooligosaccharides and help gain new insights into biomass deconstruction.