RESUMEN
The aim of this study was to evaluate the preclinical efficacy of [89Zr]Zr-DFO-Ab253 as a novel positron emission tomography (PET) tracer for CD146-positive malignant melanoma imaging. Considering the high expression of CD146 in malignant melanoma, this study investigated the effect of different CD146 expression levels on the tumor uptake of [89Zr]Zr-DFO-Ab253. CD146 selectivity was investigated by using the CD146-positive human melanoma cell A375 and the CD146-negative human alveolar epithelial cell A549. The cell uptake of [89Zr]Zr-DFO-Ab253 tracers was investigated, and receptor-binding affinities were measured by radioactive enzyme-linked immunosorbent assay. Biodistribution studies and micro-PET imaging of the radiotracers were performed on mice bearing A375 and A549 xenografts under baseline and blocking conditions. An immunohistochemical test was performed using A375 and A549 tissue sections for CD146 expression level analysis. [89Zr]Zr-DFO-Ab253 was obtained with a high radiochemical yield (87.86 ± 4.66%) and a satisfactory radiochemical purity (>98.0%). The specificity and affinity of [89Zr]Zr-DFO-Ab253 were confirmed in melanoma A375 cells and in vivo PET imaging of A375 tumor models. [89Zr]Zr-DFO-IgG and A549 lung tumors were prepared as control radiotracers and negative models to verify the specificity of [89Zr]Zr-DFO-Ab253 on CD146. [89Zr]Zr-DFO-Ab253 has a Kd of 4.01 ± 0.50 nM. PET imaging and biodistribution showed a higher uptake of [89Zr]Zr-DFO-Ab253 in A375 melanomas than that in A549 tumors (42.1 ± 4.04% vs 7.87 ± 1.30% ID/g at 120 h, P < 0.05). A low tumor uptake of [89Zr]Zr-DFO-IgG was observed with uptakes of 1.91 ± 0.41 and 2.80 ± 0.14 ID%/g when blocked at 120 h. The radiation-absorbed dose was calculated to be 0.13 mSv/MBq. This study demonstrates the synthesis and preclinical evaluation of [89Zr]Zr-DFO-Ab253 and indicates that the novel tracer has promising applications in malignant melanoma-specific PET imaging because of its high uptake and long-time retention in malignant melanoma. It also provides feasibility for the development of integrated molecular probes for diagnosis and treatment based on the CD146 target.
RESUMEN
In this study, we employed a combination approach for the preparative separation of constituents from Ginkgo biloba L. leaves. It involved multi-stage solvent extractions utilizing two-phase multi-solvent systems and countercurrent chromatography (CCC) separations using three different solvent systems. The n-heptane/ethyl acetate/water (1:1:2, v/v) and n-heptane/ethyl acetate/methanol/water (HepEMWat, 7:3:7:3, v/v) solvent systems were screened out as extraction systems. The polarities of the upper and lower phases in the multi-solvent systems were adjustable, enabling the effectively segmented separation of complex constituents in G. biloba L. The segmented products were subsequently directly utilized as samples and separated using CCC with the solvent systems acetate/n-butanol/water (4:1:5, v/v), HepEMWat (5:5:5:5, v/v), and HepEMWat (9:1:9:1, v/v), respectively. As a result, a total of 11 compounds were successfully isolated and identified from a 2 g methanol extract of G. biloba L through two-stage extraction and three CCC separation processes; among them, nine compounds exhibited high-performance liquid chromatography purity exceeding 85%.
Asunto(s)
Distribución en Contracorriente , Ginkgo biloba , Extractos Vegetales , Hojas de la Planta , Solventes , Ginkgo biloba/química , Solventes/química , Hojas de la Planta/química , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Cromatografía Líquida de Alta Presión , Agua/química , Metanol/química , Acetatos/química , Extracto de GinkgoRESUMEN
This study is the first to investigate the effects of external resistance and electrolyte concentration on the performance of a bioelectro-Fenton (BEF) system, involving measurements of power density, H2O2 generation, and bisphenol A (BPA) removal efficiency. With optimized operating conditions (external resistance of 1.12 kΩ and cathodic NaCl concentration of 1,657 mg/L), the BEF system achieved a maximum power density of 38.59 mW/m2, which is about 3.5 times higher than with 1 kΩ external resistance and no NaCl. This system featured a 71.7 % reduction in total internal resistance. The optimized BEF also accelerated the oxygen reduction reaction rate, increasing H2O2 generation by 4.4 times compared to the unoptimized system. Moreover, it exhibited superior BPA degradation performance, removing over 99 % of BPA within 14 hs, representing a 1.1 to 3.3-fold improvement over the unoptimized BEF. By the fifth cycle (70 h), the optimized BEF still removed 70 % of BPA. Optimizing the operating conditions significantly increased the abundance of electrochemically active bacteria (Pseudomonadaceae) from 2.2 % to 20 %, facilitating rapid acclimation. The study demonstrates the strong potential of an optimized BEF system for removing persistent pollutants.
RESUMEN
Salmonella typhimurium (S. typhimurium) constitutes a major public health concern. We have previously proven that Lactobacillus crispatus 7-4 (L. crispatus 7-4) can inhibit the growth of S. typhimurium and thus can be used as a biocontrol strategy to suppress foodborne S. typhimurium infections. However, the inhibitory effect and in-depth mechanism of L. crispatus 7-4 remain to be elucidated. In this study, we found that L. crispatus 7-4 can protect against S. typhimurium-induced ileum injury by promoting intestinal barrier integrity, maintaining intestinal mucosal barrier homeostasis, and reducing intestinal inflammatory response. Furthermore, we demonstrated that this probiotic strain can increase the abundance of Lactobacillus spp. to maintain microbial homeostasis and simultaneously increase the amount of γglutamylcysteine (γ-GC) by activating the glutathione metabolic pathway. The increased γ-GC promoted the transcription of Nrf2 target genes, thereby improving the host antioxidant level, reducing reactive oxygen species (ROS) accumulation, and removing pro-inflammatory cytokines. In other words, L. crispatus 7-4 could activate the enterocyte Nrf2 pathway by improving γ-GC to protect against S. typhimurium-induced intestinal inflammation and oxidative damage.
RESUMEN
Substance use disorders (SUD) and drug addiction are major threats to public health, impacting not only the millions of individuals struggling with SUD, but also surrounding families and communities. One of the seminal challenges in treating and studying addiction in human populations is the high prevalence of co-morbid conditions, including an increased risk of contracting a human immunodeficiency virus (HIV) infection. Of the ~15 million people who inject drugs globally, 17% are persons with HIV. Conversely, HIV is a risk factor for SUD because chronic pain syndromes, often encountered in persons with HIV, can lead to an increased use of opioid pain medications that in turn can increase the risk for opioid addiction. We hypothesize that SUD and HIV exert shared effects on brain cell types, including adaptations related to neuroplasticity, neurodegeneration, and neuroinflammation. Basic research is needed to refine our understanding of these affected cell types and adaptations. Studying the effects of SUD in the context of HIV at the single-cell level represents a compelling strategy to understand the reciprocal interactions among both conditions, made feasible by the availability of large, extensively-phenotyped human brain tissue collections that have been amassed by the Neuro-HIV research community. In addition, sophisticated animal models that have been developed for both conditions provide a means to precisely evaluate specific exposures and stages of disease. We propose that single-cell genomics is a uniquely powerful technology to characterize the effects of SUD and HIV in the brain, integrating data from human cohorts and animal models. We have formed the Single-Cell Opioid Responses in the Context of HIV (SCORCH) consortium to carry out this strategy.
RESUMEN
Coronavirus disease 2019 (COVID-19) has caused a global pandemic since its onset in 2019, and the development of effective vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to induce potent and long-lasting immunity remains a priority. Herein, we prepared two Lactobacillus exopolysaccharide (EPS) nanoparticle adjuvants (NPs 7-4 and NPs 8-2) that were constructed by using sulfation-modified EPS and quaternization-modified chitosan. These two NPs displayed a spherical morphology with sizes of 39 and 47 nm. Furthermore, the zeta potentials of NPs 7-4 and NPs 8-2 were 50.40 and 44.40 mV, respectively. In vitro assays demonstrated that NPs could effectively adsorb antigenic proteins and exhibited a sustained release effect. Mouse immunization tests showed that the NPs induced the expression of cytokines and chemokines at the injection site and promoted the uptake of antigenic proteins by macrophages. Mechanically, the NPs upregulated the expression of pattern recognition receptors (toll-like receptors and nod-like receptors) and activated the immune response of T cells and the production of neutralizing antibodies. In addition, the NP adjuvants had favorable immune-enhancing effects in cats, which are of great significance for controlling the trans-host transmission and re-endemicity of SARS-CoV-2. Overall, we demonstrated that NP-adjuvanted SARS-CoV-2 receptor binding domain proteins could induce robust specific humoral and cellular immunity.
Asunto(s)
COVID-19 , Nanopartículas , Animales , Ratones , Gatos , Vacunas contra la COVID-19 , SARS-CoV-2 , Sulfatos/farmacología , Adyuvantes Inmunológicos/química , Nanopartículas/química , Adyuvantes Farmacéuticos/farmacología , Inmunidad Celular , Vacunas de Subunidad/farmacologíaRESUMEN
Membrane-less single-medium sediment microbial fuel cells (single-SMFC) can remove Cu2+ from sediment through electromigration. However, the high mass transfer resistance of the sediment and amount of oxygen at the cathode of the SMFC limit its Cu2+ removal ability. Therefore, this study used an oxygen-releasing bead (ORB) for slow oxygen release to increase oxygen at the SMFC cathode and improve the mass transfer property of the sediment. Resultantly, the copper removal efficiency of SMFC increased significantly. Response surface methodology was used to optimize the nano zero-valent iron (nZVI)-modified biochar as the catalyst to enhance the ability of the modified ORB (ORBm) to remove Cu2+ and slow release of O2. The maximum Cu2+ removal (95 %) and the slowest O2 release rate (0.41 mg O2/d·g ORBm) were obtained when the CaO2 content and ratio of nZVI-modified biochar to unmodified biochar were 0.99 g and 4.95, respectively. When the optimized ORBm was placed at the single-SMFC cathode, the voltage output and copper removal increased by 4.6 and 2.1 times, respectively, compared with the system without ORBm. This shows that the ORBm can improve the migration of Cu2+ in the sediment, providing a promising remediation method for Cu-contaminated sediments.
Asunto(s)
Fuentes de Energía Bioeléctrica , Carbón Orgánico , Cobre , Electrodos , Sedimentos Geológicos , Hierro , Oxígeno , Fuentes de Energía Bioeléctrica/microbiología , Carbón Orgánico/química , Cobre/química , Oxígeno/química , Hierro/química , Sedimentos Geológicos/química , Sedimentos Geológicos/microbiologíaRESUMEN
PM2.5 is an important risk factor for the development and progression of cognitive impairment-related diseases. Ferroptosis, a new form of cell death driven by iron overload and lipid peroxidation, is proposed to have significant implications. To verify the possible role of ferroptosis in PM2.5-induced neurotoxicity, we investigated the cytotoxicity, intracellular iron content, iron metabolism-related genes, oxidative stress indices and indicators involving in Nrf2 and ferroptosis signaling pathways. Neurotoxicity biomarkers as well as the ferroptotic cell morphological changes were determined by Western Blot and TEM analysis. Our results revealed that PM2.5 induced cytotoxicity, lipid peroxidation, as indicated by MDA content, and neurotoxicity via Aß deposition in a dose-related manner. Decreased cell viability and excessive iron accumulation in HT-22 cells can be partially blocked by ferroptosis inhibitors. Interestingly, GPX activity, Nrf2, and its regulated ferroptotic-related proteins (i.e. GPX4 and HO-1) were significantly up-regulated by PM2.5. Moreover, gene expression of DMT1, TfR1, IRP2 and FPN1 involved in iron homeostasis and NCOA4-dependent ferritinophagy were activated after PM2.5 exposure. The results demonstrated that PM2.5 triggered ferritinophagy-dependent ferroptotic cell death due to iron overload and redox imbalance. Activation of Nrf2 signaling pathways may confer a protective mechanism for PM2.5-induced oxidative stress and ferroptosis.
Asunto(s)
Ferroptosis , Sobrecarga de Hierro , Humanos , Factor 2 Relacionado con NF-E2/genética , Oxidación-Reducción , Hierro , Material Particulado/toxicidadRESUMEN
Clinical pancreatic ductal adenocarcinoma (PDAC) treatment is severely limited by lack of effective KRAS suppression strategies. To address this dilemma, a reactive oxygen species (ROS)-responsive and PDAC-targeted nanodrug named Z/B-PLS was constructed to confront KRAS through dual-blockade of its downstream PI3K/AKT/mTOR and RAF/MEK/ERK for enhanced PDAC treatment. Specifically, photosensitizer zinc phthalocyanine (ZnPc) and PI3K/mTOR inhibitor BEZ235 (BEZ) were co-loaded into PLS which was constructed by click chemistry conjugating MEK inhibitor selumetinib (SEL) to low molecular weight heparin with ROS-responsive oxalate bond. The BEZ and SEL blocked PI3K/AKT/mTOR and RAF/MEK/ERK respectively to remodel glycolysis and non-canonical glutamine metabolism. ZnPc mediated photodynamic therapy (PDT) could enhance drug release through ROS generation, further facilitating KRAS downstream dual-blockade to create treatment-promoting drug delivery-therapeutic positive feedback. Benefiting from this broad metabolic modulation cascade, the metabolic symbiosis between normoxic and hypoxic tumor cells was also cut off simultaneously and effective tumor vascular normalization effects could be achieved. As a result, PDT was dramatically promoted through glycolysis-non-canonical glutamine dual-metabolism regulation, achieving complete elimination of tumors in vivo. Above all, this study achieved effective multidimensional metabolic modulation based on integrated smart nanodrug delivery, helping overcome the therapeutic challenges posed by KRAS mutations of PDAC.
Asunto(s)
Carcinoma Ductal Pancreático , Nanopartículas , Neoplasias Pancreáticas , Humanos , Glutamina/farmacología , Glutamina/metabolismo , Glutamina/uso terapéutico , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/uso terapéutico , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/uso terapéutico , Especies Reactivas de Oxígeno/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Serina-Treonina Quinasas TOR/metabolismo , Serina-Treonina Quinasas TOR/uso terapéutico , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/uso terapéutico , Glucólisis , Fototerapia , Línea Celular TumoralRESUMEN
The unmarked potential drug molecule azamulin has been found to be a specific inhibitor of CYP3A4 and CYP3A5 in recent years, but this molecule also shows different binding ability and affinity to the two CYP3A isoforms. In order to explore the microscopic mechanism, conventional molecular dynamics (MD) simulation methods were performed to study the dynamic interactions between two isoforms and azamulin. The simulation results show that the binding of the ligand leads to different structural properties of two CYP3A proteins. First of all, compared with apo-CYP3A4, the binding of the ligand azamulin can lead to changes in the structural flexibility of CYP3A4, i.e., holo-CYP3A4 is more flexible than apo-CYP3A4. The structural changes of CYP3A5 are just the opposite. The ligand binding increases the rigidity of CYP3A5. Furthermore, the representative structures of the production phase in the MD simulation were in details analyzed to obtain the microscopic interactions between the ligand azamulin and two CYP3A isoforms at the atomic level. It is speculated that the difference of composition and interaction of the active sites is the fundamental cause of the change of structural properties of the two proteins.Communicated by Ramaswamy H. Sarma.
RESUMEN
The utilization of downhole optical cables has significantly enhanced the efficiency and reliability of oilfield production operations; however, the challenging high-temperature and high-pressure conditions prevalent in oil-gas fields markedly reduce the service lifespan of these optical cables. This limitation severely impedes their application and further development in subterranean environments. In this study, a qualitative analysis was conducted on the structural materials utilized in two types of optical cables to identify these materials and assess the high-temperature tolerance and aging resistance properties of the optical fibers incorporated within. It was discovered that hydrogen infiltration into the subterranean optical cables predominantly accounts for their operational failure. To address this issue, an optical loss testing platform was established, facilitating the execution of a high-temperature and high-pressure hydrogen permeation aging experiment on the optical fibers, allowing for the evaluation of the hydrogen resistance capabilities of the two types of optical fibers. The findings from this study provide a theoretical foundation and methodological guidance for the optimization of optical fibers, aiming to enhance their durability and functional performance in adverse environmental conditions encountered in oil-gas field applications.
RESUMEN
Silver nanoparticles (AgNPs) are widely used in daily life and medical fields owing to their unique physicochemical properties. Daily exposure to AgNPs has become a great concern regarding their potential toxicity to human beings, especially to the central nervous system. Ferroptosis, a newly recognized programmed cell death, was recently reported to be associated with the neurodegenerative process. However, whether and how ferroptosis contributes to AgNPs-induced neurotoxicity remain unclear. In this study, we investigated the role of ferroptosis in neurotoxic effects induced by AgNPs using in vitro and in vivo models. Our results showed that AgNPs induced a notable dose-dependent cytotoxic effect on HT-22 cells and cognitive impairment in mice as indicated by a decline in learning and memory and brain tissue injuries. These findings were accompanied by iron overload caused by the disruption of the iron transport system and activation of NCOA4-mediated autophagic degradation of ferritin. The excessive free iron subsequently induced GSH depletion, loss of GPX and SOD activities, differential expression of Nrf2 signaling pathway elements, down-regulation of GPX4 protein and production of lipid peroxides, initiating ferroptosis cascades. The mitigating effects of ferrostatin-1 and deferoxamine on iron overload, redox imbalance, neuronal cell death, impairment of mice learning and memory, Aß deposition and synaptic plasticity reduction suggested ferroptosis as a potential molecular mechanism in AgNPs-induced neurotoxicity. Taken together, these results demonstrated that AgNPs induced neuronal cell death and cognitive impairment with Aß deposition and reduction of synaptic plasticity, which were mediated by ferroptosis caused by iron-mediated lipid peroxidation. Our study provides new insights into the underlying mechanisms of AgNPs-induced neurotoxicity and predicts potential preventive strategies.
Asunto(s)
Disfunción Cognitiva , Ferroptosis , Sobrecarga de Hierro , Nanopartículas del Metal , Ratones , Humanos , Animales , Plata/toxicidad , Ferroptosis/fisiología , Nanopartículas del Metal/toxicidad , Hierro/metabolismo , Disfunción Cognitiva/inducido químicamenteRESUMEN
Wheat sheath blight caused by the necrotic fungal pathogen Rhizoctonia cerealis is responsible for severe damage to bread wheat. Reactive oxygen species (ROS) are vital for stress resistance by plants and their homeostasis plays an important role in wheat resistance to sheath blight. Valine-glutamine (VQ) proteins play important roles in plant growth and development, and responses to biotic and abiotic stresses. However, the functional mechanism mediated by wheat VQ protein in response to sheath blight via ROS homeostasis regulation is unclear. In this study, we identified TaVQ22 protein containing the VQ motif and clarified the functional mechanisms involved in the defense of wheat against R. cerealis. TaVQ22 silencing reduced the accumulation of ROS and enhanced the resistance of wheat to R. cerealis. In addition, we showed that TaVQ22 regulated ROS generation by interacting with the WRKY transcription factor TaWRKY19-2B, thereby indicating that TaVQ22 and TaWRKY19-2B formed complexes in the plant cell nucleus. Yeast two-hybrid analysis showed that the VQ motif in TaVQ22 is crucial for the interaction, where it inhibits the transcriptional activation function of TaWRKY19-2B. In summary, TaVQ22 interacts with TaWRKY19-2B to regulate ROS homeostasis and negatively regulate the defense response to R. cerealis infection. This study provides novel insights into the mechanism that allows VQ protein to mediate the immune response in plants.
Asunto(s)
Enfermedades de las Plantas , Triticum , Triticum/genética , Especies Reactivas de Oxígeno , Homeostasis , Desarrollo de la Planta , Saccharomyces cerevisiaeRESUMEN
Antibiotic-resistant bacteria are prevalent in husbandry around the world due to the abuse of antibiotic growth promoters (AGPs); therefore, it is necessary to find alternatives to AGPs in animal feed. Among all the candidates, probiotics are promising alternatives to AGPs against Salmonella infection. The anti-Salmonella effects of three probiotic strains, namely, Lactobacillus crispatus 7-4, Lactobacillus johnsonii 3-1, and Pediococcus acidilactici 20-1, have been demonstrated in our previous study. In this study, we further obtained the alginate beads containing compound probiotics, namely, microencapsulate probiotics (MP), and evaluated its regulatory effect on the health of broilers. We incubated free and microencapsulate probiotics in simulated gastric and intestinal juice for 2 h, and the results showed that compared to free probiotics, encapsulation increased tolerance of compound probiotics in the simulated gastrointestinal condition. We observed that the application of probiotics, especially MP, conferred protective effects against Salmonella typhimurium (S.Tm) infection in broilers. Compared to the S.Tm group, the MP could promote the growth performance (p < 0.05) and reduce the S.Tm load in intestine and liver (p < 0.05). In detail, MP pretreatment could modulate the cecal microflora and upregulate the relative abundance of Lactobacillus and Enterobacteriaceae. Besides, MP could reduce the inflammation injury of the intestine and liver, reduce the pro-inflammatory cytokines (IL-6, TNF-α, IL-1ß) expression, and induce of anti-inflammatory cytokine (IL-10) expression. Furthermore, MP could inhibit NLRP3 pathway in ileum, thereby attenuating S.Tm-induced inflammation. In conclusion, MP could be a new feeding supplementation strategy to substitute AGPs in poultry feeding.
Asunto(s)
Probióticos , Salmonelosis Animal , Animales , Salmonella typhimurium/fisiología , Pollos , Salmonelosis Animal/prevención & control , Salmonelosis Animal/microbiología , Probióticos/farmacología , Citocinas , Inflamación , AntibacterianosRESUMEN
Analysis of cell-cell communication (CCC) in the tumor micro-environment helps decipher the underlying mechanism of cancer progression and drug tolerance. Currently, single-cell RNA-Seq data are available on a large scale, providing an unprecedented opportunity to predict cellular communications. There have been many achievements and applications in inferring cell-cell communication based on the known interactions between molecules, such as ligands, receptors and extracellular matrix. However, the prior information is not quite adequate and only involves a fraction of cellular communications, producing many false-positive or false-negative results. To this end, we propose an improved hierarchical variational autoencoder (HiVAE) based model to fully use single-cell RNA-seq data for automatically estimating CCC. Specifically, the HiVAE model is used to learn the potential representation of cells on known ligand-receptor genes and all genes in single-cell RNA-seq data, respectively, which are then utilized for cascade integration. Subsequently, transfer entropy is employed to measure the transmission of information flow between two cells based on the learned representations, which are regarded as directed communication relationships. Experiments are conducted on single-cell RNA-seq data of the human skin disease dataset and the melanoma dataset, respectively. Results show that the HiVAE model is effective in learning cell representations, and transfer entropy could be used to estimate the communication scores between cell types.
Asunto(s)
Neoplasias , Análisis de Expresión Génica de una Sola Célula , Humanos , Análisis de la Célula Individual/métodos , Comunicación Celular , Secuenciación del Exoma , Análisis de Secuencia de ARN/métodos , Perfilación de la Expresión Génica/métodos , Microambiente TumoralRESUMEN
In bacteria, the second messenger cyclic di-GMP (c-di-GMP) is synthesized and degraded by multiple diguanylate cyclases (DGCs) and phosphodiesterases. A high level of c-di-GMP induces biofilm formation and represses motility. WspR, a hybrid response regulator DGC, produces c-di-GMP when it is phosphorylated. FlhF, a signal recognition particle-type GTPase, is initially localized to the cell poles and is indispensable for polar flagellar localization in Pseudomonas aeruginosa. In this study, we report that deletion of flhF affected biofilm formation and the c-di-GMP level in P. aeruginosa. Phenotypic analysis of a flhF knockout mutant revealed increased biofilm formation, wrinkled colonies on Congo red agar, and an elevated c-di-GMP level compared to the wild-type strain, PAO1. Yeast and bacterial two-hybrid systems showed that FlhF binds to the response regulator HsbR, and HsbR binds to WspR. Deletion of hsbR or wspR in the ΔflhF background abolished the phenotype of ΔflhF. In addition, confocal microscopy demonstrated that WspR-GFP was distributed throughout the cytoplasm and formed a visible cluster at one cell pole in PAO1 and ΔhsbR, but it was mainly distributed as visible clusters at the lateral side of the periplasm and with visible clusters at both cell poles in ΔflhF. These findings suggest that FlhF influences the subcellular cluster and localization of WspR and negatively modulates WspR DGC activity in a manner dependent on HsbR. Together, our findings demonstrate a novel mechanism for FlhF modulating the lifestyle transition between motility and biofilm via HsbR to regulate the DGC activity of WspR.IMPORTANCECyclic di-GMP (c-di-GMP) is a second messenger that controls flagellum biosynthesis, adhesion, virulence, motility, exopolysaccharide production, and biofilm formation in bacteria. Recent research has shown that distinct diguanylate cyclases (DGCs) or phosphodiesterases (PDEs) produce highly specific outputs. Some DGCs and PDEs contribute to the total global c-di-GMP concentration, but others only affect local c-di-GMP in a microenvironment. However, the underlying mechanisms are unclear. Here, we report that FlhF affects the localization and DGC activity of WspR via HsbR and is implicated in local c-di-GMP signaling in Pseudomonas aeruginosa. This study establishes the link between the c-di-GMP signaling system and the flagellar localization and provides insight for understanding the complex regulatory network of c-di-GMP signaling.
Asunto(s)
Dietilestilbestrol/análogos & derivados , Proteínas de Escherichia coli , Pseudomonas aeruginosa , Pseudomonas aeruginosa/genética , Proteínas de Escherichia coli/genética , GMP Cíclico/metabolismo , Biopelículas , Liasas de Fósforo-Oxígeno/genética , Hidrolasas Diéster Fosfóricas/metabolismo , Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión GénicaRESUMEN
Acquiring big-size datasets to raise the performance of deep models has become one of the most critical problems in representation learning (RL) techniques, which is the core potential of the emerging paradigm of federated learning (FL). However, most current FL models concentrate on seeking an identical model for isolated clients and thus fail to make full use of the data specificity between clients. To enhance the classification performance of each client, this study introduces the FDRL, a federated discriminative RL model, by partitioning the data features of each client into a global subspace and a local subspace. More specifically, FDRL learns the global representation for federated communication between those isolated clients, which is to capture common features from all protected datasets via model sharing, and local representations for personalization in each client, which is to preserve specific features of clients via model differentiating. Toward this goal, FDRL in each client trains a shared submodel for federated communication and, meanwhile, a not-shared submodel for locality preservation, in which the two models partition client-feature space by maximizing their differences, followed by a linear model fed with combined features for image classification. The proposed model is implemented with neural networks and optimized in an iterative manner between the server of computing the global model and the clients of learning the local classifiers. Thanks to the powerful capability of local feature preservation, FDRL leads to more discriminative data representations than the compared FL models. Experimental results on public datasets demonstrate that our FDRL benefits from the subspace partition and achieves better performance on federated image classification than the state-of-the-art FL models.
RESUMEN
CYP 3A4 and CYP 3A5 are two important members of the human cytochrome P450 family. Although their overall structures are similar, the local structures of the active site are different, which directly leads to obvious individual differences in drug metabolic efficacy and toxicity. In this work, midazolam (MDZ) was selected as the probe substrate, and its interaction with two proteins, CYP 3A4 and CYP 3A5, was studied by molecular dynamics simulation (MD) along with the calculation of the binding free energy. The results show that two protein-substrate complexes have some similarities in enzyme-substrate binding; that is, in both complexes, Ser119 forms a high occupancy hydrogen bond with MDZ, which plays a key role in the stability of the interaction between MDZ and the enzymes. However, the complex formed by CYP 3A4 and MDZ is more stable, which may be attributed to the sandwich structure formed by the fluorophenyl group of the substrate with Leu216 and Leu482. Our study interprets the binding differences between two isoform-substrate complexes and reveals a structure-function relationship from the atomic perspective, which is expected to provide a theoretical basis for accurately measuring the effectiveness and toxicity of drugs for individuals in the era of precision medicine.
Asunto(s)
Citocromo P-450 CYP3A , Midazolam , Humanos , Midazolam/química , Citocromo P-450 CYP3A/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Dominio Catalítico , Isoformas de Proteínas/metabolismoRESUMEN
Recent advances in targeted therapy and immunotherapy have substantially improved the treatment of melanoma. However, therapeutic strategies are still needed for unresponsive or treatment-relapsed patients with melanoma. To discover antibody-drug conjugate (ADC)-tractable cell surface targets for melanoma, we developed an atlas of melanoma cell surface-binding antibodies (pAb) using a proteome-scale antibody array platform. Target identification of pAbs led to development of melanoma cell killing ADCs against LGR6, TRPM1, ASAP1, and MUC18, among others. MUC18 was overexpressed in both tumor cells and tumor-infiltrating blood vessels across major melanoma subtypes, making it a potential dual-compartment and universal melanoma therapeutic target. AMT-253, an MUC18-directed ADC based on topoisomerase I inhibitor exatecan and a self-immolative T moiety, had a higher therapeutic index compared with its microtubule inhibitor-based counterpart and favorable pharmacokinetics and tolerability in monkeys. AMT-253 exhibited MUC18-specific cytotoxicity through DNA damage and apoptosis and a strong bystander killing effect, leading to potent antitumor activities against melanoma cell line and patient-derived xenograft models. Tumor vasculature targeting by a mouse MUC18-specific antibody-T1000-exatecan conjugate inhibited tumor growth in human melanoma xenografts. Combination therapy of AMT-253 with an antiangiogenic agent generated higher efficacy than single agent in a mucosal melanoma model. Beyond melanoma, AMT-253 was also efficacious in a wide range of MUC18-expressing solid tumors. Efficient target/antibody discovery in combination with the T moiety-exatecan linker-payload exemplified here may facilitate discovery of new ADC to improve cancer treatment. SIGNIFICANCE: Discovery of melanoma-targeting antibodies using a proteome-scale array and use of a cutting-edge linker-payload system led to development of a MUC18-targeting antibody-exatecan conjugate with clinical potential for treating major melanoma subtypes.
Asunto(s)
Inmunoconjugados , Melanoma , Canales Catiónicos TRPM , Humanos , Ratones , Animales , Inmunoconjugados/farmacología , Proteoma , Inhibidores de Topoisomerasa I/farmacología , Inmunoterapia , Ensayos Antitumor por Modelo de Xenoinjerto , Línea Celular TumoralRESUMEN
We have previously proven that the environmental toxin could accelerate the development and progression of nonalcoholic steatohepatitis (NASH). However, the underlying mechanism associated with such excessive inflammation hasn't been fully illustrated. Although Genistein has been well accepted for its capability in anti-inflammation and anti-oxidation, its effect in ameliorating contaminants-induced NASH still needs to be identified. In this study, using chickens and primary chicken hepatocytes as models, we found that NOD-like receptor pyrin domain-containing 3 (NLRP3) inflammasome were over-activated in bromoacetic acid (BAA, one of the typical environmental toxins)-induced NASH, characterized by the infiltration of inflammatory cell, and the increase of NLRP3, Caspase-1 p20, and cytokines (IL-1ß, IL-18) expressions. Interestingly, genistein treatment could recover these changes, with the signs of restored activities of anti-oxidases, decreased expressions of NLRP3 inflammasome components, and increased levels of elements in phase I metabolic system. The detailed mechanism was that, via up-regulating aryl hydrocarbon receptor (AHR), genistein lifted mRNA levels of Cyp1-related genes to reconstruct cytochrome P450 (CYP450) systems, and the raised AHR negatively regulated NLRP3 inflammasome activity to relieve inflammation. More important, the interaction and co-localization between AHR and NLRP3 was first proved, and genistein could promote the levels of AHR that interacted with NLRP3, which thereafter blocked the activation of NLRP3 inflammasome. Conclusively, in this research, we confirmed the AHR-dependent protective role of genistein in environmental toxin-linked NASH, which shed light on the potential precautions for contaminants-induced NASH.