RESUMEN
Manipulating exciton dissociation and charge-carrier transfer processes to selectively generate free radicals of more robust photocatalytic oxidation capacity for mineralizing refractory pollutants remains challenging. Herein, we propose a strategy by simultaneously introducing the cyano-group and Na into graphitic carbon nitride (CN) to obtain CN-Cy-Na, which makes the charge-carrier transfer pathways the dominant process and consequently achieves the selective generation of free radicals. Briefly, the cyano-group intensifies the local charge density of CN, offering a potential well to attract the hole of exciton, which accelerates the exciton dissociation. Meanwhile, the separated electron transfers efficiently under the robust built-in electric field induced by the cyano-group and Na, and eventually accumulates in the heptazine ring of CN for the following O2 reduction due to the reinforced electron sink effect caused by Na. As a result, CN-Cy-Na exhibits 4.42 mmol L-1 h-1 productivity with 97.6% selectivity for free radicals and achieves 82.1% total organic carbon removal efficiency in the tetracycline photodegradation within 6 h. Additionally, CN-Cy-Na also shows outstanding photodegradation efficiency of refractory pollutants, including antibiotics, pesticide plastic additives, and dyes. This work presents an innovative approach to manipulating the exciton effect and enhancing charge-carrier mobility within two-dimensional photocatalysts, opening an avenue for precise control of free radical generation.
RESUMEN
Sunlight-responsive minerals contribute significantly to biogeochemical cycles by activating oxygen (O2) to generate reactive oxygen species (ROS). However, the role of crystal water, incorporated into minerals through hydration during rock cycles, in O2 activation remains largely unexplored. Here, we construct tungstite models containing oxygen vacancies to elucidate the modulation of mineral-based ROS dynamics by the synergy between oxygen vacancy and crystal water. Crystal water promotes the protonation process of superoxide anion radicals to produce hydrogen peroxide (H2O2) and alleviates its decomposition. This mineral-based H2O2 photosynthesis system efficiently eliminates organic pollutants in a sequential light-dark reaction. Furthermore, this synergy effect can extend to other metal oxide minerals such as TiO2, SnO2, CuO, ZnO, and Bi2O3. Our results illuminate an overlooked pathway for modulating the protonation process by immobilized water in hydrous minerals, playing a crucial role in ROS storage and migration and pollutant dynamics in a natural environment throughout the day/night cycle.
RESUMEN
As a biological macromolecule, the superantigen staphylococcal enterotoxin C2 (SEC2) is one of the most potent known T-cell activators, and it induces massive cytotoxic granule production. With this property, SEC2 and its mutants are widely regarded as immunomodulating agents for cancer therapy. In a previous study, we constructed an MHC-II-independent mutant of SEC2, named ST-4, which exhibits enhanced immunocyte stimulation and antitumor activity. However, tumor cells have different degrees of sensitivity to SEC2/ST-4. The mechanisms of immune resistance to SEs in cancer cells have not been investigated. Herein, we show that ST-4 could activate more powerful human lymphocyte granule-based cytotoxicity than SEC2. The results of RNA-seq and atomic force microscopy (AFM) analysis showed that, compared with SKOV3 cells, the softer ES-2 cells could escape from SEC2/ST-4-induced cytotoxic T-cell-mediated apoptosis by regulating cell softness through the CDC42/MLC2 pathway. Conversely, after enhancing the stiffness of cancer cells by a nonmuscle myosin-II-specific inhibitor, SEC2/ST-4 exhibited a significant antitumor effect against ES-2 cells by promoting perforin-dependent apoptosis and the S-phase arrest. Taken together, these data suggest that cell stiffness could be a key factor of resistance to SEs in ovarian cancer, and our findings may provide new insight for SE-based tumor immunotherapy.
Asunto(s)
Antineoplásicos , Enterotoxinas , Humanos , Enterotoxinas/farmacología , Enterotoxinas/metabolismo , Superantígenos/farmacología , Antineoplásicos/farmacología , Linfocitos T , Activación de LinfocitosRESUMEN
CD8+ T cells can switch between fatty acid catabolism and mitochondrial energy metabolism to sustain expansion and their cytotoxic functions. ST-4 is a TCR-enhanced mutant derived from superantigen staphylococcal enterotoxin C2 (SEC2), which can hyperactivate CD4+ T cells without MHC class II molecules. However, whether ST-4/SEC2 can enhance metabolic reprogramming in CD8+ T cells remains poorly understood. In this study, we found that ST-4, but not SEC2, could induce proliferation of purified CD8+ T cell from BALB/c mice in Vß8.2- and -8.3-specific manners. Results of gas chromatography-mass spectroscopy analysis showed that fatty acid contents in CD8+ T cells were increased after ST-4 stimulation. Flow cytometry and Seahorse analyses showed that ST-4 significantly promoted mitochondrial energy metabolism in CD8+ T cells. We also observed significantly upregulated levels of gene transcripts for fatty acid uptake and synthesis, and significantly increased protein expression levels of fatty acid and mitochondrial metabolic markers of mTOR/PPARγ/SREBP1 and p38-MAPK signaling pathways in ST-4-activated CD8+ T cells. However, blocking mTOR, PPARγ, SREBP1, or p38-MAPK signals with specific inhibitors could significantly relieve the enhanced fatty acid catabolism and mitochondrial capacity induced by ST-4. In addition, blocking these signals inhibited ST-4-stimulated CD8+ T cell proliferation and effector functions. Taken together, our findings demonstrate that ST-4 enhanced fatty acid and mitochondria metabolic reprogramming through mTOR/PPARγ/SREBP and p38-MAPK signaling pathways, which may be important regulatory mechanisms of CD8+ T cell activation. Understanding the effects of ST-4-induced regulatory metabolic networks on CD8+ T cells provide important mechanistic insights to superantigen-based tumor therapy.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Metabolismo Energético , Enterotoxinas , Ácidos Grasos/inmunología , Activación de Linfocitos/efectos de los fármacos , Mitocondrias/inmunología , Mutación , Animales , Metabolismo Energético/efectos de los fármacos , Metabolismo Energético/inmunología , Enterotoxinas/genética , Enterotoxinas/inmunología , Enterotoxinas/toxicidad , Femenino , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/inmunología , Ratones , Ratones Endogámicos BALB CRESUMEN
Chlorimuron-ethyl is a widely used herbicide in agriculture. However, uncontrolled chlorimuron-ethyl application causes serious environmental problems. Chlorimuron-ethyl can be effectively degraded by microbes, but the underlying molecular mechanisms are not fully understood. In this study, we identified the possible pathways and key genes involved in chlorimuron-ethyl degradation by the Chenggangzhangella methanolivorans strain CHL1, a Methylocystaceae strain with the ability to degrade sulfonylurea herbicides. Using a metabolomics method, eight intermediate degradation products were identified, and three pathways, including a novel pyrimidine-ring-opening pathway, were found to be involved in chlorimuron-ethyl degradation by strain CHL1. Transcriptome sequencing indicated that three genes (atzF, atzD, and cysJ) are involved in chlorimuron-ethyl degradation by strain CHL1. The gene knock-out and complementation techniques allowed for the functions of the three genes to be identified, and the enzymes involved in the different steps of chlorimuron-ethyl degradation pathways were preliminary predicted. The results reveal a previously unreported pathway and the key genes of chlorimuron-ethyl degradation by strain CHL1, which have implications for attempts to enrich the biodegradation mechanism of sulfonylurea herbicides and to construct engineered bacteria in order to remove sulfonylurea herbicide residues from environmental media.
Asunto(s)
Herbicidas , Methylocystaceae , Contaminantes del Suelo , Biodegradación Ambiental , Herbicidas/metabolismo , Methylocystaceae/metabolismo , Pirimidinas/metabolismo , Contaminantes del Suelo/metabolismo , Compuestos de Sulfonilurea/metabolismoRESUMEN
Bacterial superantigens potently activate conventional T-cells to induce massive cytokine production and mediate tumor cell death. To engineer superantigens for immunotherapy against tumors in clinic, we previously generated SAM-1, a staphylococcal enterotoxins C2 (SEC2) mutant, that exhibited significantly reduced toxicity but maintained the superantigen activity in animal models. This present study aimed to investigate whether SAM-1 activates T cells and induces apoptosis in human tumor cells. We found that SAM-1 induced the maturation of dendritic cells (DCs) with upregulating expression of the surface markers CD80, CD86 and HLA-DR, which secreted high levels of IL-12p70 by activating TLR2-NF-κB signaling pathways. SAM-1 could activate human CD4+ subgroup T cells and CD8+ subgroup T cells in the presence of mature dendritic cells (DCs), leading to the productions of cytokines TRAIL, IL-2, IFN-γ and TNF-α. We observed that TRAIL mediated the apoptosis and S-phase and G2/M-phase arrest in HGC-27 tumor cells via binding to upregulated death receptors DR4 and DR5. Using shRNA knockdown in HGC-27 cells or constitutive overexpression in ES2 cells for DR4 and DR5, we demonstrated the vital requirement of DR4 and DR5 in apoptosis of tumor cells in response to TRAIL secreted from SAM-1-activated T cells. Collectively, our results will facilitate better understanding of SAM-1-based immunotherapies for cancer.
Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Enterotoxinas/farmacología , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/biosíntesis , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Células A549 , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD8-positivos/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Enterotoxinas/genética , Células HeLa , Humanos , Células K562 , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiologíaRESUMEN
PURPOSE: To develop an automated framework for sleep stage scoring from PSG via a deep neural network. METHODS: An automated deep neural network was proposed by using a multi-model integration strategy with multiple signal channels as input. All of the data were collected from one single medical center from July 2017 to April 2019. Model performance was evaluated by overall classification accuracy, precision, recall, weighted F1 score, and Cohen's Kappa. RESULTS: Two hundred ninety-four sleep studies were included in this study; 122 composed the training dataset, 20 composed the validation dataset, and 152 were used in the testing dataset. The network achieved human-level annotation performance with an average accuracy of 0.8181, weighted F1 score of 0.8150, and Cohen's Kappa of 0.7276. Top-2 accuracy (the proportion of test samples for which the true label is among the two most probable labels given by the model) was significantly improved compared to the overall classification accuracy, with the average being 0.9602. The number of arousals affected the model's performance. CONCLUSION: This research provides a robust and reliable model with the inter-rater agreement nearing that of human experts. Determining the most appropriate evaluation parameters for sleep staging is a direction for future research.
Asunto(s)
Redes Neurales de la Computación , Apnea Obstructiva del Sueño/fisiopatología , Fases del Sueño/fisiología , Adulto , Anciano , Aprendizaje Profundo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polisomnografía , Adulto JovenRESUMEN
SEC2, a major histocompatibility complex class II (MHC II)-dependent T-cell mitogen, binds MHC II and T-cell receptor (TCR) Vßs to induce effective co-stimulating signals for clonal T-cell expansion. We previously characterized a SEC2 mutant with increased recognition of TCR Vßs, ST-4, which could intensify NF-κB signaling transduction, leading to IL-2 production and T-cell activation. In this study, we found that in contrast to SEC2, ST-4 could induce murine CD4+ T-cell proliferation in a Vß8.2- and Vß8.3-specific manner in the absence of MHC II+ antigen-presenting cells (APCs). Furthermore, although IL-2 secretion in response to either SEC2 or ST-4 stimulation was accompanied by up-regulation of protein kinase Cθ (PKCθ), inhibitor of κB (IκB), α and ß IκB kinase (IKKα/ß), IκBα, and NF-κB in mouse splenocytes, only ST-4 could activate CD4+ T cells in the absence of MHC II+ APCs through the PKCθ/NF-κB signaling pathway. The PKCθ inhibitor AEB071 significantly suppressed SEC2/ST-4-induced T-cell proliferation, CD69 and CD25 expression, and IL-2 secretion with or without MHC II+ APCs. Further, SEC2/ST-4-induced changes in PKCθ/NF-κB signaling were significantly relieved by AEB071 in a dose-dependent manner. Using Lck siRNA, we found that Lck controlled SEC2/ST-4-induced phosphorylation of PKCθ. We also demonstrated that the IL-2R/STAT5 pathway is essential for SEC2/ST-4-induced T-cell activation. Collectively, our data demonstrate that an enhanced ST-4-TCR interaction can compensate for lack of MHC II and stimulate MHC II-free CD4+ T-cell proliferation via PKCθ/NF-κB and IL-2R/STAT5 signaling pathways. Compared with SEC2, intensified PKCθ/NF-κB and IL-2R/STAT5 signals induced by ST-4 lead to enhanced T-cell activation. The results of this study will facilitate better understanding of TCR-based immunotherapies for cancer.
Asunto(s)
Enterotoxinas/genética , Activación de Linfocitos , Mutación , Receptores de Antígenos de Linfocitos T/metabolismo , Transducción de Señal , Linfocitos T/inmunología , Animales , Proliferación Celular , Enterotoxinas/metabolismo , Femenino , Regulación de la Expresión Génica/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Interleucina-2/metabolismo , Ratones , Ratones Endogámicos BALB C , FN-kappa B/metabolismo , Fosforilación , Proteína Quinasa C-theta/metabolismo , Receptores de Interleucina-2/metabolismo , Factor de Transcripción STAT5/metabolismo , Bazo/inmunología , Linfocitos T/citologíaRESUMEN
As a kind of superantigen, staphylococcal enterotoxin C2 (SEC2) is well known as a powerful immunomodulator. However, most previous studies about SEC2 focus on its T cell activating characters. But the direct effect of SEC2 on antigen-presenting cells (APCs) which are important for the T cell activation is not clearly. In this study, we investigated the effect of SEC2 on murine bone marrow-derived dendritic cells (BMDCs) which are known as the specialized professional APCs. Contrary to its effects on T cells, SEC2 could not induce proliferation or cytotoxicity to BMDCs even in high concentrations. While SEC2 could promote the mature of BMDCs with increased expression of co-stimulatory molecules on cell membrane such as CD80, CD86, and MHC II. The production of pro-inflammatory cytokines such as TNF-α, IFN-γ and IL-6 were also increased in BMDCs treated with SEC2. We also found that SEC2 enhanced the genes expression of pattern recognition receptors including toll-like receptors 2 (TLR2) and TLR4 in BMDCs, and up-regulated the key signal molecule MyD88 in both mRNA and protein levels. In addition, SEC2 also caused IκBα degradating and NFκB p65 translocating from the cytoplasm to the nucleus in BMDCs. The siRNAs for both TLR2 and TLR4, as well as NFκB specific inhibitor BAY 11-7085 could inhibit the co-stimulatory molecules expression and pro-inflammatory cytokines releasing induced by SEC2. Moreover, TLR2/4 specific siRNAs inhibited p65 and MyD88 upregulation induced by SEC2. In summary, all our results indicated that SEC2 could stimulate BMDCs maturation through TLR-NFκB signaling pathways.
Asunto(s)
Médula Ósea/efectos de los fármacos , Enterotoxinas/farmacología , FN-kappa B/efectos de los fármacos , Receptor Toll-Like 4/efectos de los fármacos , Animales , Médula Ósea/metabolismo , Células de la Médula Ósea/citología , Citocinas/metabolismo , Células Dendríticas , Femenino , Activación de Linfocitos/efectos de los fármacos , Ratones Endogámicos BALB C , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos , Receptor Toll-Like 4/metabolismoRESUMEN
Staphylococcal enterotoxin C2 (SEC2), a superantigen, causes rapid clonal expansion of lymphocytes and secretion of T cell growth factors, leading to a severe inflammatory response within tissues. Although previous studies have shown that ST-4, a SEC2 mutant with enhanced recognition of Vß regions of T-cell receptors (TCRVß), can activate an increased number of T cells and produce more cytokines than SEC2. However, the signaling mechanisms of SEC2/ST-4-mediated immune activation have not been addressed. In this study, we showed that the phosphatidylinositide-3-kinase (PI-3K) inhibitor LY294002, mammalian target of rapamycin (mTOR) inhibitor rapamycin, and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) inhibitor Bay11-7085 could suppress SEC2/ST-4-induced proliferation, CD69/CD25 expression, cell-cycle progression, and IL-2 production in BALB/c mouse splenocytes. In addition, we observed significantly upregulated expression of p70S6K, cyclin E, cyclin D3, and NF-ĸB/p65, but downregulated expression of p27kip during SEC2/ST-4-driven T cells activation. However, SEC2/ST-4-induced changes in cell cycle and PI3K/mTOR signaling were significantly relieved by either LY294002 or rapamycin, and the induction of NF-ĸB/p65 induced was significantly downregulated by Bay11-7085. Moreover, we found that IL-2 secretion was positively associated with p65 expression in a time- and dose-dependent manner. Taken together, our findings demonstrate the involvement of PI3K/mTOR and NF-κB signaling pathways in SEC2/ST-4-induced T cell activation. ST-4 intensifies PI3K/mTOR and NF-ĸB signaling transduction, ultimately leading to enhance T cell activation. These results provide a theoretical mechanism for future immunotherapy using ST-4.
Asunto(s)
Enterotoxinas/genética , Activación de Linfocitos/inmunología , FN-kappa B/inmunología , Fosfatidilinositol 3-Quinasas/inmunología , Linfocitos T/inmunología , Serina-Treonina Quinasas TOR/inmunología , Animales , Células Cultivadas , Femenino , Interleucina-2/inmunología , Ratones Endogámicos BALB C , Mutación , Transducción de Señal , Bazo/citologíaRESUMEN
Superantigens can induce cell-mediated cytotoxicity preferentially against MHC II-positive target cells with large amounts of inflammatory cytokines releasing. In this study, superantigen staphylococcal enterotoxin C (SEC) 1 was investigated to evaluate its potential in bladder cancer immunotherapy in vitro and in vivo. Our results revealed that SEC1 could stimulate the proliferation of human peripheral blood mononuclear cells (PBMCs) in a dose-dependent manner, accompanied with the release of interleukin-2, interferon-γ, and tumor necrosis factor-α, and increased the population of CD4+ T cells and CD8+ T cells. PBMCs stimulated by SEC1 could initiate significant cytotoxicity towards human bladder cancer cells in vitro. The results of in vivo antitumor experiment indicated that SEC1 could decrease the rate of tumor formation and prolong the survival time of tumor-bearing mice. Our study demonstrated that SEC1 inhibited the growth of bladder cancer. And it is also suggested that SEC1 may become a candidate for bladder cancer immunotherapy.
Asunto(s)
Enterotoxinas/farmacología , Superantígenos/farmacología , Neoplasias de la Vejiga Urinaria/patología , Animales , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/efectos de los fármacos , Recuento de Células , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Interferón gamma/metabolismo , Interleucina-2/metabolismo , Ratones , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Staphylococcal enterotoxin C2 (SEC2), a member of bacterial superantigen, is one of the most potent known activators of T lymphocytes. With this property, SEC2 has already been used in clinic as a tumor immunotherapy agent in China. To increase the antitumor activity, a SEC2 mutant named ST-4 (GKVTG102-106WWH) with amino acid substitutions in T cell receptor (TCR)-binding domain was generated by site-directed mutagenesis, and the molecular mechanism of the enhanced antitumor activity was investigated. Results showed that ST-4 could activate much more Vß 8.2 and 8.3 T cells and NK cells compared with SEC2, and exhibited significantly enhanced immunocyte stimulation and antitumor activity in vitro. The synthetic peptide sequencing the residues of mutant TCR-binding domain could competitively inhibit the immunocyte stimulation activity of ST-4. Most importantly, ST-4 up-regulated granzyme B and perforin at both mRNA and protein levels. We also found that expression of proapoptotic proteins cytochrome c, BAX and activation of caspase-3, 9 was up-regulated, and antiapoptotic protein Bcl-xL was down-regulated in the treatment with either ST-4 or SEC2. When granzyme B inhibitor or perforin inhibitor is presented, tumor cell viability was significantly rescued. Taken together, we demonstrate that increased ST-4-TCR recognition contributed to massive T cells and NK cells activation. These activated cells released up-regulated granzyme B and perforin, which induced the enhanced tumor cells apoptosis by mitochondrial apoptotic pathway, and ultimately led to enhanced tumor cell growth inhibition. ST-4 may be a promising candidate for antitumor clinic usage in future.
Asunto(s)
Citotoxicidad Inmunológica/efectos de los fármacos , Enterotoxinas/toxicidad , Granzimas/metabolismo , Mutación , Perforina/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Animales , Línea Celular Tumoral , Electroforesis en Gel de Poliacrilamida , Enterotoxinas/genética , Femenino , Ratones , Ratones Endogámicos BALB CRESUMEN
Staphylococcal enterotoxins C2 (SEC2) is a classical model of superantigens (SAg), which has the powerful ability to activate T cells as well as induce massive cytokine production. This property makes SEC2 and its mutants well concerned as a potential new immune-regulatory agent for cancer therapy. We previously constructed a SEC2 mutant named SAM-3, which had prominently antitumor activity in BALB/c mice model. But, the underlying molecular mechanism for stimulation of human peripheral blood mononuclear cells (PBMCs) and antitumor effect on human tumor cells induced by SAM-3 is not clear. Here, we showed that SAM-3 could activate human TCR Vß 12, 13A, 14, 15, 17, and 20 CD8(+) subgroup T cells, which secreted the cytokines IL-2, IFN-γ, and TNF-α, and exhibit stimulation activity in a dose-dependent manner. TNF-α secreted from activated T cells could induce apoptosis and G1-phase arrest and lead to the antitumor effect in HepG2 cells. Meanwhile, SAM-3 upregulated the expression of tumor necrosis factor receptor 1 (TNFR1) mRNA and activity of caspase-3 and caspase-8. We also found that the antitumor activity and activity of caspase-3 and caspase-8 were decreased when the neutralizing TNF-α monoclonal antibody presented. These data suggest that TNF-α secreted by SAM-3-activated T cells is an important factor in inducing apoptosis in HepG2 cells.
Asunto(s)
Antineoplásicos/metabolismo , Apoptosis , Enterotoxinas/metabolismo , Células Hep G2/fisiología , Proteínas Mutantes/metabolismo , Linfocitos T/fisiología , Factor de Necrosis Tumoral alfa/metabolismo , Enterotoxinas/genética , Células Hep G2/efectos de los fármacos , Humanos , Activación de Linfocitos , Proteínas Mutantes/genética , Linfocitos T/efectos de los fármacosRESUMEN
A network of inflammation factors is related to pulmonary fibrosis induced by paraquat (PQ) poisoning. At high doses, the superantigen staphylococcal enterotoxin C1 (SEC1) can induce immunological unresponsiveness and inhibit release of inflammation factors. In this study, site-directed mutagenesis was performed at the H118 and H122 amino acid residues of SEC1 to reduce SEC1 toxicity. The SEC1 mutant showed significantly decreased pyrogenic toxicity, but retained clonal anergy at high dosages in vitro. Pretreatment with the SEC1 mutant prior to PQ poisoning in mice reduced symptom duration and severity, prolonged survival time, and decreased the splenocyte response to ConA induction. The SEC1 mutant also down-regulated several important cytokines related to fibrosis in the plasma after PQ poisoning. SEC1 decreased the expression of genes related to pulmonary fibrosis, including α-SMA, COL1a1, COL3 and TGF-ß1, in PQ poisoned mice. Histomorphological observation indicated alleviation of pathological changes in the lungs after SEC1 pretreatment compared to mice in the PQ group. In conclusion, the SEC1 mutant reduced pulmonary interstitial fibrosis induced by PQ poisoning.
Asunto(s)
Antígenos Bacterianos/uso terapéutico , Enterotoxinas/uso terapéutico , Herbicidas/toxicidad , Paraquat/toxicidad , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/prevención & control , Animales , Antígenos Bacterianos/genética , Antígenos Bacterianos/toxicidad , Proliferación Celular , Citocinas/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Enterotoxinas/genética , Enterotoxinas/toxicidad , Escherichia coli/genética , Femenino , Expresión Génica/efectos de los fármacos , Herbicidas/antagonistas & inhibidores , Ratones , Ratones Endogámicos BALB C , Mutagénesis Sitio-Dirigida , Paraquat/antagonistas & inhibidores , Fibrosis Pulmonar/genética , Conejos , Bazo/citología , Bazo/efectos de los fármacos , Superantígenos , Análisis de SupervivenciaRESUMEN
The aim of this study was to construct a bifunctional fusion protein consisting of staphylococcal enterotoxin C2 (SEC2) and enhanced green fluorescent protein (EGFP). We inserted EGFP and SEC2 fragments into the pET-28a(+) vector to create the expression plasmid vector, pET-28a(+)-SEC2-EGFP, using a two-step method. After verification of the plasmid, successful isolation of the fusion protein, SEC2-EGFP, was achieved by Ni+-affinity chromatography. Fluorescence microscopy, methylthiazol tetrazolium, and flow cytometry assays demonstrated that the constructed fusion protein not only retained the fluorescence signal of EGFP but also exhibited SEC2 bioactivity. Therefore, SEC2-EGFP is a promising tool for the study of the detailed temporal and spatial distributions of SEC2 in cells. Future studies with this vector may help uncover novel therapeutic strategies to treat or manage SEC2-associated diseases and be a new clinical tool for exploiting SEC2 in immunotherapy.
Asunto(s)
Enterotoxinas/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Ingeniería de Proteínas/métodos , Proteínas Recombinantes de Fusión/metabolismo , Animales , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Enterotoxinas/química , Enterotoxinas/genética , Escherichia coli , Femenino , Proteínas Fluorescentes Verdes/química , Proteínas Fluorescentes Verdes/genética , Linfocitos , Ratones , Ratones Endogámicos BALB C , Plásmidos , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/farmacologíaRESUMEN
BACKGROUND: To accurately measure students' science, technology, engineering and mathematics (STEM) career interest, researchers must get inside the 'black box' to understand students' conceptualizations of STEM careers. AIMS: The aim of Study 1 was to explore whether students' conceptualizations of STEM included medical careers. The aim of Study 2 was to explore whether predictors of STEM career interest (e.g., gender and motivation) varied by STEM definition (inclusion/exclusion of medical careers). SAMPLES: In Study 1, the sample was US college students (N = 125) who were mostly White (80%). In Study 2, the sample was US 10th-grade high school students (N = 455) who were mostly Black (79%). METHODS: In Study 1, students completed an online questionnaire. In Study 2, students completed various measures of math achievement, motivation (science and math expectancies of success, interest and importance value) and career interest with an importance. RESULTS: In Study 1, medical careers were less often classified as STEM careers than traditional STEM careers, but more often classified as STEM than non-STEM careers. In Study 2, science importance value was the only motivational predictor of students' STEM+Medicine career interest, and no motivation constructs predicted traditional STEM career interest. Boys expressed greater interest in traditional STEM careers, while girls expressed greater interest in STEM+Medicine careers. CONCLUSIONS: Students' conceptualizations of STEM are not binary. Thus, we recommend researchers are explicit about their definition of STEM with study participants, in their coding and in their publications.
Asunto(s)
Ingeniería , Matemática , Ciencia , Tecnología , Femenino , Humanos , Masculino , Selección de Profesión , MotivaciónRESUMEN
AIM: To develop a homogeneous assay for high-throughput screening (HTS) of inhibitors of phosphodiesterase 10 (PDE10). METHODS: Purified human PDE10 enzyme derived from E coli, [(3)H]-cAMP and yttrium silicate microbeads were used to develop an HTS assay based on the scintillation proximity assay (SPA) technology. This method was applied to a large-scale screening campaign against a diverse compound library and subsequent confirmation studies. Preliminary structure-activity relationship (SAR) studies were initiated through limited structural modifications of the hits. RESULTS: The IC50 value of the control compound (papaverine) assessed with the SPA approach was comparable and consistent with that reported in the literature. Signal to background (S/B) ratio and Z' factor of the assay system were evaluated to be 5.24 and 0.71, respectively. In an HTS campaign of 71 360 synthetic and natural compounds, 67 hits displayed reproducible PDE10 inhibition, of which, 8 were chosen as the scaffold for structural modifications and subsequent SAR analysis. CONCLUSION: The homogeneous PDE10 SPA assay is an efficient and robust tool to screen potential PDE10 inhibitors. Preliminary SAR studies suggest that potent PDE10 inhibitors could be identified and developed through this strategy.
Asunto(s)
Descubrimiento de Drogas/métodos , Ensayos Analíticos de Alto Rendimiento/métodos , Papaverina/farmacología , Inhibidores de Fosfodiesterasa/farmacología , Hidrolasas Diéster Fosfóricas/metabolismo , Humanos , Papaverina/química , Inhibidores de Fosfodiesterasa/química , Hidrolasas Diéster Fosfóricas/químicaRESUMEN
Once the TCR-SAg-MHC II ternary complex is established, it triggers a variety of intracellular signal transduction pathways, which provoke extreme responses in the immune system. However, the signaling events that involved in SAg-induced immune activation are not well understood. In this study, we demonstrated that the Ca(2+)/calcineurin (CaN)/nuclear factor of activated T cells (NFAT) signaling pathway was involved in SEC2-induced immune activation, and selective blockade of CaN by its inhibitor cyclosporine A (CsA) can completely inhibited the SEC2-induced T-cell stimulating potency. In addition, we selected an engineered SEC2 mutant named SAM-1 based on a series of biological activity tests, and our further studies on it not only confirmed that the CaN activity and gene transcription of its key substrates were proportional to the SEC2/SAM-1-induced T-cell stimulating potency, but also suggested that intensified Ca(2+)/CaN/NFAT signaling transduction induced by SAM-1 resulted in enhanced T-cell stimulating potency, production of cytokines and cytotoxicity, which finally elicit the improved antitumor activity of SAM-1 in vivo.
Asunto(s)
Antineoplásicos/inmunología , Calcineurina/metabolismo , Enterotoxinas/inmunología , Superantígenos/inmunología , Linfocitos T/inmunología , Animales , Antineoplásicos/metabolismo , Enterotoxinas/metabolismo , Activación de Linfocitos , Ratones , Ratones Endogámicos BALB C , Superantígenos/metabolismoRESUMEN
Asthe most-used pesticides in the agricultural production process, herbicides are mainly applied to protect crops from weeds. However, with the increased global demand for food, the dosage of herbicides is rising annually, and the efficacy of herbicides is getting stronger, which can cause some environmental issues including the accumulation, migration and transformation, and toxic effects of herbicides in agricultural soils. According to the characteristics of herbicide contamination and regional agricultural production, developing green and low-carbon technologies to reduce the ecological risks of herbicides to the soil-crop systems is a current concern in the ecological environment field. In this paper, relevant studies in recent years on herbicide pollution management in agricultural soils were identified and reviewed, the research progress and application cases of remediation technologies for herbicide pollution was analyzed and demonstrated, and future research and development tendency regarding the remediation of herbicides pollution was also prospected. Current remediation technologies for herbicides mainly include bioremediation technologies (e.g., microbial remediation, enzyme remediation, and phytoremediation), adsorption, and immobilization technologies (e.g., biochar-based materials). The bioremediation technologieswere rather mature and had been applied to the herbicide-contaminated soil in fields. Additionally, many successful bioremediation cases have been reported. Moreover, in order to enhance the remediation effect on herbicide pollution in agriculture soils, remediation technologies have been gradually developed from a single model to a coupled model with physical,chemical, and biological technology, which can maximize the synergy of the multi-technology application.
Asunto(s)
Herbicidas , Contaminantes del Suelo , Suelo , Contaminantes del Suelo/análisis , Agricultura , Biodegradación Ambiental , TecnologíaRESUMEN
Micro/nanoplastics in the environment can be ingested by organisms and spread throughout the food chain, ultimately posing a threat to human health. However, the risk of continuous oral exposure in mammals remains unresolved. In this study, we utilized a continuous gavage mouse model to investigate the potential intestinal risks associated with oral exposure to polystyrene micro/nanoplastics (PS-MNPs) with environmentally relevant concentrations. The effects of PS-MNPs with different particle sizes on the gut microbiota, intestinal barrier, and intestinal immune function were evaluated. PS-MNPs can accumulate in the intestine after oral exposure and alter the composition of the gut microbiota. Exposure to PS-MNPs significantly reduced the ratio of Firmicutes to Bacteroidetes as well as the number of potentially beneficial bacteria in the gut, while the number of potentially harmful bacteria significantly increased. The short-chain fatty acids metabolized by gut microbiota were significantly changed by PS-MNPs. Exposure to PS-MNPs disrupts the function of the intestinal barrier and leads to inflammation in the intestines. The levels of secretory immunoglobulin A in the intestine and the differentiation of CD4+ and CD8+ T cells in mesenteric lymph nodes were significantly decreased by PS-MNPs. Moreover, the impact of PS-MNPs on mammalian intestinal health is influenced by the exposure duration and particle size, rather than the concentration. It also suggests that nanoplastics may pose more severe environmental risks.