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A recent study reported that Acinetobacter baumannii could induce autophagy, but the recognition and clearance mechanism of intracytosolic A. baumannii in the autophagic process and the molecular mechanism of autophagy induced by the pathogen remains unknown. In this study, we first demonstrated that invading A. baumannii induced a complete, ubiquitin-mediated autophagic response that is dependent upon septins SEPT2 and SEPT9 in mammalian cells. We also demonstrated that autophagy induced by A. baumannii was Beclin-1 dependent via the AMPK/ERK/mammalian target of rapamycin pathway. Of interest, we found that the isochorismatase mutant strain had significantly decreased siderophore-mediated ferric iron acquisition ability and had a reduced the ability to induce autophagy. We verified that isochorismatase was required for the recognition of intracytosolic A. baumannii mediated by septin cages, ubiquitinated proteins, and ubiquitin-binding adaptor proteins p62 and NDP52 in autophagic response. We also confirmed that isochorismatase was required for the clearance of invading A. baumannii by autophagy in vitro and in the mouse model of infection. Together, these findings provide insight into the distinctive recognition and clearance of intracytosolic A. baumannii by autophagy in host cells, and that isochorismatase plays a critical role in the A. baumannii-induced autophagic process.-Wang, Y., Zhang, K., Shi, X., Wang, C., Wang, F., Fan, J., Shen, F., Xu, J., Bao, W., Liu, M., Yu, L. Critical role of bacterial isochorismatase in the autophagic process induced by Acinetobacter baumannii in mammalian cells.
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Acinetobacter baumannii/enzimología , Autofagia/fisiología , Hidrolasas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Células Cultivadas , Proteínas del Citoesqueleto/metabolismo , Humanos , Masculino , Proteínas de la Membrana/metabolismo , Ratones Endogámicos BALB C , Ubiquitina/metabolismoRESUMEN
Babesiosis is an emerging tick-transmitted zoonosis prevalent in large parts of the world. This study was designed to determine the rates of Babesia microti infection among small rodents in Yunnan province, where human cases of babesiosis have been reported. Currently, distribution of Babesia in its endemic regions is largely unknown. In this study, we cataloged 1672 small wild rodents, comprising 4 orders, from nine areas in western Yunnan province between 2009 and 2011. Babesia microti DNA was detected by polymerase chain reaction in 4·3% (72/1672) of the rodents analyzed. The most frequently infected rodent species included Apodemus chevrieri and Niviventer fulvescens. Rodents from forests and shrublands had significantly higher Babesia infection rates. Genetic comparisons revealed that Babesia was most similar to the Kobe- and Otsu-type strains identified in Japan. A variety of rodent species might be involved in the enzootic maintenance and transmission of B. microti, supporting the need for further serological investigations in humans.
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Babesia microti/aislamiento & purificación , Babesiosis/epidemiología , Enfermedades de los Roedores/epidemiología , Animales , Babesia microti/genética , Babesiosis/diagnóstico , Babesiosis/parasitología , China/epidemiología , Filogenia , Reacción en Cadena de la Polimerasa/veterinaria , ARN Protozoario/genética , ARN Ribosómico 18S/genética , Enfermedades de los Roedores/parasitología , Análisis de Secuencia de ARN/veterinariaRESUMEN
The major histocompatibility complex is a diverse gene family that plays a crucial role in the adaptive immune system. In humans, the MHC class I genes consist of the classical loci of HLA-A, -B, and -C, and the nonclassical loci HLA-E, -F, and -G. In Platyrrhini species, few MHC class I genes have been described so far and were classified as MHC-E, MHC-F, and MHC-G, with MHC-G possibly representing a classical MHC class I locus while there were arguments about the existence of the MHC-B locus in Platyrrhini. In this study, MHC class I genes were identified in eight common marmosets (Callithrix jacchus) and two brown-headed spider monkeys (Ateles fusciceps). For common marmosets, 401 cDNA sequences were sequenced and 18 alleles were detected, including 14 Caja-G alleles and 4 Caja-B alleles. Five to eleven Caja-G alleles and one to three Caja-B alleles were detected in each animal. For brown-headed spider monkeys, 102 cDNA sequences were analyzed, and 9 new alleles were identified, including 5 Atfu-G and 4 Atfu-B alleles. Two or three Atfu-G and two Atfu-B alleles were obtained for each of animal. In phylogenetic analyses, the MHC-G and -B alleles from the two species and other Platyrrhini species show locus-specific clusters with bootstrap values of 86% and 50%. The results of pairwise sequence comparisons and an excess of non-synonymous nucleotide substitutions in the PBR region are consistent with the suggestion that Caja-G and Atfu-G may be classical MHC class I loci in the Platyrrhini species But it appears that MHC-B locus of the two Platyrrhini species shares features with both classical and nonclasical MHC class I loci. Our results are an important addition to the limited MHC immunogenetic information available for the Platyrrhini species.
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Atelinae/genética , Callithrix/genética , Genes MHC Clase I , Alelos , Secuencia de Aminoácidos , Animales , Evolución Molecular , Datos de Secuencia Molecular , Filogenia , Alineación de SecuenciaRESUMEN
With the increasing industrialization and urbanization, the ecological environment is suffering from severe deterioration in Liaohe coastal wetland, and petroleum spill is one of the pollution sources. Suaeda salsa (L.) Pall (S. salsa), one of the predominant plants in Liaohe coastal wetland, is facing the increasing degradation. Terpenes are a class of inherent compounds in plants, and play key role in maintain the growth of plants. However, the environmental stress on the terpene metabolism remained unclear in the plants. In the present study, the influence of petroleum spill on terpene metabolism in S. salsa was systematically investigated by analysis of concentrations, compositions and stable carbon isotope. Under the stress of petroleum spill, terpene concentrations showed the decreasing trend, indicating the inhibition effect of petroleum spill on terpene synthesis in S. salsa. The proportions of Sabinene and A-humulene showed the obviously increased with the influence of petroleum spill, implying that these congeners were more sensitive to petroleum spills. The significant changes in stable carbon isotope compositions were observed for Borneol, Dl-menthol, A-humulene and (-) -@-bisabolol, with the enrichment in heavier isotopes in residual fractions. This result indicated that the heavier 13C was preferentially fixed on terpene by S. salsa under the petroleum stress. The similar change trends along the incubation time was observed for A-humulene and (-) - trans caryophyllene, which might imply that A-humulene was one of the products of (-) - trans caryophyllene in S. salsa. Overall, the findings of present study verified the influence of petroleum spill on terpene metabolism in S. salsa, and were meaningful for protecting the plants in the petroleum-pollution wetlands.
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Chenopodiaceae , Petróleo , Sesquiterpenos Policíclicos , Humedales , Petróleo/toxicidad , Sesquiterpenos Monocíclicos , Isótopos de Carbono , CarbonoRESUMEN
BACKGROUND: To date, disease-modifying antirheumatic drugs (DMARDs) are widely used as the primary first-line treatment option for patients with rheumatoid arthritis (RA), and the curative effect of methotrexate (MTX) and leflunomide (LEF; MTX + LEF) is greater than that of single-agent MTX therapy, but the synergistic mechanism of MTX + LEF is unclear. METHODS: First, we explored the mechanism of action of MTX + LEF in RA through network pharmacology and molecular docking. Venn diagram analysis revealed 97 overlapping gene targets of MTX + LEF-RA and STRING, along with Cytoscape plug-in MOCDE and cytoHubba; and GO enrichment analysis revealed that the functions of 97 synergistic targets were related to 123 molecular functions (MF), 63 cell components (CC), and 1,068 biological processes (BP). The Cytoscape plug-in ClueGO demonstrated that these targets were enriched in KEGG pathways of 52 terms, whereas 9 pivotal genes were mainly involved in the signaling pathways of estrogen, Ras, Rap1, PI3K-Akt, relaxin, TNF, AMPK, FoxO, prolactin, IL-17, and adherens junction. Finally, CETSA and DARTS validated the direct binding of MTX or LEF to the selected target proteins EGFR, PPARG, MMP9, and SRC in RAW264.7 cells. RESULTS: We identified 292 MTX targets and 247 LEF targets from 7 databases. Furthermore, 2,814 potential targets of RA were identified by merging 1,925 targets from 7 databases and 999 differentially expressed genes (DEGs) between normal controls and patients with RA extracted from 5 GEO databases. Nine pivotal genes, ESR1, ALB, CASP3, EGFR, HSP90AA1, SRC, MMP9, PPARG, and IGF1, were identified. Molecular docking verified that both MTX and LEF strongly bind to most of the 9 pivotal proteins except ESR1 and IGF1. CONCLUSION: These results contribute to our understanding of the enhancement mechanism of MTX combined with LEF and provide a targeted basis for the clinical treatment of RA.
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Tibetan macaques (Macaca thibetana), stump-tailed macaques (M. arctoides), Assamese macaques (M. assamensis), and northern pig-tailed macaques (M. leonina) are four major species of Macaca in China. In order to effectively use these species in biomedical research, thorough investigations of their MHC immunogenetics are required. In this study, we identified MHC class I sequences using cDNA cloning and sequencing on a cohort of six M. thibetana, three M. arctoides, three M. assamensis, and three M. leonina derived from Sichuan and Yunnan provinces of China. Eighty new alleles were identified, including 26 MHC-A alleles, 46 MHC-B alleles, and 8 MHC-I alleles. Among them, Math-A1*126:01, Math-B*190:01, Math-B*191:01, Math-B*192:01, Maar-A1*127:01, Maar-A1*129:01, and Maas-A1*128:01 represent lineages that had not been reported earlier in Macaca. Phylogenetic analyses show that no obvious separation of lineages among these species of Macaca. This study provides important information about the MHC immunogenetics for the four major species of Chinese macaques and adds value to these species as model organisms in biomedical research.
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Evolución Molecular , Genes MHC Clase I , Macaca/genética , Filogenia , Alelos , Animales , China , Genotipo , Macaca/inmunología , Especificidad de la EspecieRESUMEN
Ferroptosis is a newly discovered type of cell death and has recently been shown to be associated with asthma. However, the relationship between them at the genetic level has not been elucidated via informatics analysis. In this study, bioinformatics analyses are conducted using asthma and ferroptosis datasets to identify candidate ferroptosis-related genes using the R software. Weighted gene co-expression network analysis is performed to identify co-expressed genes. Protein-protein interaction networks, the Kyoto encyclopedia of genes and genomes, and gene ontology enrichment analysis are used to identify the potential functions of the candidate genes. We experimentally validate the results of our analysis using small interfering RNAs and plasmids to silence and upregulate the expression of the candidate gene in human bronchial epithelial cells (BEAS-2B). The ferroptosis signature levels are examined. Bioinformatics analysis of the asthma dataset GDS4896 shows that the level of the aldo-keto reductase family 1 member C3 (AKR1C3) gene in the peripheral blood of patients with severe therapy-resistant asthma and controlled persistent mild asthma (MA) is significantly upregulated. The AUC values for asthma diagnosis and MA are 0.823 and 0.915, respectively. The diagnostic value of AKR1C3 is verified using the GSE64913 dataset. The gene module of AKR1C3 is evident in MA and functions through redox reactions and metabolic processes. Ferroptosis indicators are downregulated by the overexpression of AKR1C3 and upregulated by silencing AKR1C3. The ferroptosis-related gene AKR1C3 can be used as a diagnostic biomarker for asthma, particularly for MA, and regulates ferroptosis in BEAS-2B cells.
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Asma , Ferroptosis , Humanos , Miembro C3 de la Familia 1 de las Aldo-Ceto Reductasas/metabolismo , Hidroxiprostaglandina Deshidrogenasas/análisis , Hidroxiprostaglandina Deshidrogenasas/genética , Hidroxiprostaglandina Deshidrogenasas/metabolismo , Ferroptosis/genética , Biomarcadores , Asma/genética , Biología ComputacionalRESUMEN
Introduction: Group 2 innate lymphoid cells (ILC2s) play a crucial role in house dust mite (HDM)-induced allergic inflammation, and allergen immunotherapy (AIT) holds promise for treating the disease by reducing the frequency of ILC2s. Despite significant progress in AIT for allergic diseases, there remains a need to improve the control of allergic symptoms. Methods: We investigated the synergistic effect of the Notch signaling pathway and subcutaneous immunotherapy (SCIT) in treating allergic airway inflammation in mice and their impact on the ratio of ILC2s in lung tissues. This was achieved by establishing the HDM-induced airway allergic disorders (HAAD) model and SCIT model. Additionally, we conducted in vitro investigations into the effect of the Notch signaling pathway on the secretory function of activated ILC2s using fluorescence-activated cell sorting. Furthermore, we explored the coactivation of the Notch signaling pathway with SCIT in vitro by sorting ILC2s from the lung tissues of mice after SCIT modeling. Results: Previously, our group demonstrated that Notch signaling pathway inhibitors can reduce allergic airway inflammation in mice. Notch signaling induces lineage plasticity of mature ILC2s. In this study, we showed that AIT alleviates allergic airway inflammation and suppresses the frequency of ILC2s induced by HDM. Interestingly, AIT combined with a γ-secretase inhibitor (GSI), an inhibitor of the Notch signaling pathway, significantly inhibited the frequency of ILC2s, reduced airway inflammation, and suppressed Th2-type responses in a mouse model. Furthermore, lung ILC2s from HDM-challenged mice with or without AIT were treated with GSI in vitro, and we found that GSI dramatically reduced the secretion of type 2 inflammatory factors in ILC2s. Discussion: These findings suggest that Notch signaling pathway inhibitors can be used as adjuvant therapy for AIT and may hold potential treatment value in the cooperative control of allergic airway inflammation during early AIT.
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Hipersensibilidad , Pyroglyphidae , Ratones , Animales , Inmunidad Innata , Linfocitos , Pulmón , Hipersensibilidad/terapia , Desensibilización Inmunológica , Inflamación/terapiaRESUMEN
Diabetes mellitus (DM) complicated with Staphylococcus aureus (S. aureus) infection lacks effective treatment strategies. In this study, we found that insulin combined with linezolid has potential to deal with the thorny problem. In vitro, our drug sensitivity assay, bacterial growth curve and hemolytic tests showed that a combination of insulin and linezolid exerted good antibacterial and anti-α-hemolysin activity, CCK8 experiment, glucose content and glycogen content determination showed that the combination of insulin and linezolid increased murine macrophage survival rate and reduced the extracellular glucose level of high glucose-treated MH-S cells and intracellular glycogen level, and Western blot showed that the combination inhibited TLR2/MAPKs/NLRP3-related inflammatory pathways in MH-S cells. The results of in vivo experiments showed that the combination therapy stabilized glucose level, remained body weight, ameliorated lung injury including improving pulmonary edema and decreasing lung wet/dry weight ratio, reduced the CFUs and inflammation in the lung tissue in a mouse model of diabetes with S. aureus pneumonia, and inhibited the expression of TLR2, MAPKs and NLRP3 inflammatory pathway. Overall, the combination of insulin and linezolid as autolytic inhibitor exhibited the effects of significant antibacterial and improving glucose level in vitro and in vivo, and also has an anti-inflammation activity via the TLR2/MAPKs/NLRP3 pathway, this paves the way for new treatments for diabetes mellitus complicated with S. aureus infection.
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Diabetes Mellitus , Neumonía Estafilocócica , Animales , Ratones , Linezolid/farmacología , Linezolid/metabolismo , Linezolid/uso terapéutico , Neumonía Estafilocócica/tratamiento farmacológico , Staphylococcus aureus , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Receptor Toll-Like 2/metabolismo , Insulina/metabolismo , Antibacterianos/uso terapéutico , Diabetes Mellitus/tratamiento farmacológicoRESUMEN
Interleukin-17A (IL-17A) levels are elevated in patients with asthma. Ferroptosis has been identified as the non-apoptotic cell death type associated with asthma. Data regarding the relation of ferroptosis with asthma and the effect of IL-17A on modulating ferroptosis in asthma remain largely unclear. The present work focused on investigating the role of IL-17A in allergic asthma-related ferroptosis and its associated molecular mechanisms using public datasets, clinical samples, human bronchial epithelial cells, and an allergic asthma mouse model. We found that IL-17A was significantly upregulated within serum in asthma cases. Adding IL-17A significantly increased ferroptosis within human bronchial epithelial cells (BEAS-2B). In ovalbumin (OVA)-induced allergic asthmatic mice, IL-17A regulated and activated lipid peroxidation induced ferroptosis, whereas IL-17A knockdown effectively inhibited ferroptosis in vivo by protection of airway epithelial cells via the xCT-GSH-GPX4 antioxidant system and reduced airway inflammation. Mouse mRNA sequencing results indicated that the tumor necrosis factor (TNF) pathway was the differential KEGG pathway in the OVA group compared to healthy controls and the OVA group compared to the IL-17A knockout OVA group. We further used N-acetylcysteine (TNF inhibitor) to inhibit the TNF signaling pathway, which was found to protect BEAS-2B cells from IL-17A induced lipid peroxidation and ferroptosis damage. Our findings reveal a novel mechanism for the suppression of ferroptosis in airway epithelial cells, which may represent a new strategy for the use of IL-17A inhibitors against allergic asthma.
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Asma , Ferroptosis , Animales , Humanos , Ratones , Asma/patología , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Inflamación/genética , Interleucina-17/genética , Interleucina-17/efectos adversos , Interleucina-17/metabolismo , Pulmón/metabolismo , Ratones Endogámicos BALB C , Ovalbúmina/efectos adversosRESUMEN
Candida albicans is the most common fungal pathogen in humans. The emergence of resistance to azole antifungals has raised the issue of using such antifungals in combination to optimise therapeutic outcome. The objective of this study was to evaluate in vitro synergy of pseudolaric acid B (PAB) and fluconazole (FLC) against clinical isolates of C. albicans. The in vitro antifungal activity of PAB, a diterpene acid from Pseudolarix kaempferi Gordon, was evaluated alone and in combination with FLC against 22 FLC-resistant (FLC-R) and 12 FLC-susceptible (FLC-S) C. albicans using the chequerboard microdilution method and time-killing test assays. Synergism was observed in all 22 (100%) FLC-R strains tested as determined by both fractional inhibitory concentration index (FICI) with values ranging from 0.02 to 0.13 and bliss independence (BI) models. Synergism was observed in two of 12 (17%) FLC-S strains as determined by FICI model with values ranging from 0.25 to 0.5 and in three of 12 (18%) FLC-S strains as determined by BI model. For FLC-R strains, the drug concentrations of FLC and PAB, where synergistic interactions were found, ranged from 0.06 to 4 µg ml(-1) and 0.5 to 4 µg ml(-1) respectively. For FLC-S strains, the drug concentrations of FLC and PAB were 1-8 µg ml(-1) and 0.5-4 µg ml(-1) respectively. The BI model gave results consistent with FICI, but no antagonistic activity was observed in any of the strains tested. These interactions between PAB and FLC were confirmed using the time-killing test for the selected strains. Fluconazole and PAB exhibited a good synergism against azole-R isolates of C. albicans.
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Antifúngicos/farmacología , Candida albicans/efectos de los fármacos , Diterpenos/farmacología , Fluconazol/farmacología , Candida albicans/aislamiento & purificación , Candidiasis/microbiología , Sinergismo Farmacológico , Humanos , Pruebas de Sensibilidad Microbiana , Viabilidad Microbiana/efectos de los fármacos , Factores de TiempoRESUMEN
p-Anisaldehyde (4-methoxybenzaldehyde), an extract from Pimpinella anisum L. seeds, is a potential novel preservative. To reveal the possible action mechanism of p-anisaldehyde against microorganisms, yeast-based commercial oligonucleotide microarrays were used to analyze the genome-wide transcriptional changes in response to p-anisaldehyde. Quantitative real-time RT-PCR was performed for selected genes to verify the microarray results. We interpreted our microarray data with the clustering tool, T-profiler. Analysis of microarray data revealed that p-anisaldehyde induced the expression of genes related to sulphur assimilation, aromatic aldehydes metabolism, and secondary metabolism, which demonstrated that the addition of p-anisaldehyde may influence the normal metabolism of aromatic aldehydes. This genome-wide transcriptomics approach revealed first insights into the response of Saccharomyces cerevisiae (S. cerevisiae) to p-anisaldehyde challenge.
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Antifúngicos/toxicidad , Benzaldehídos/toxicidad , Perfilación de la Expresión Génica , Regulación Fúngica de la Expresión Génica , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/genética , Biología Computacional/métodos , Genes Fúngicos , Redes y Vías Metabólicas , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Estadística como Asunto/métodosRESUMEN
Interferon-induced transmembrane proteins (IFITMs) are transmembrane proteins induced by interferon that can provide broad-spectrum antiviral activities. However, there are few reports on the antiviral activity of monkey-derived IFITMs. In this study, the IFITM1 and IFITM3 genes of African green monkey (AGM) were cloned and overexpressed in Vero cells, followed by infection with mouse norovirus (MNV) and severe fever with thrombocytopenia syndrome virus (SFTSV). The results showed that monkey IFITM1 and IFITM3 can be stably overexpressed in Vero cells. Both IFITM1 and IFITM3 from AGM could effectively restrict infection by SFTSV, and the viral inhibition rate of IFITM3 was more obvious compared with IFITM1. However, both monkey IFITM1 and IFITM3 had no significant effect on the replication of MNV. These results indicate that different IFITMs have different functions, which may be related to the structure of the host IFITMs and the types of pathogens.
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Proteínas de la Membrana/genética , Norovirus/fisiología , Phlebovirus/fisiología , Replicación Viral/genética , Animales , Chlorocebus aethiops , Clonación Molecular , Proteínas de la Membrana/clasificación , Ratones , Células VeroRESUMEN
Dictamnine, a natural plant product, has been reported to have antimicrobial activity against bacteria and fungi; however, the dictamnine response mechanisms of microorganisms are still poorly understood. We have shown that dictamnine has antimicrobial activities against the model fungus Saccharomyces cerevisiae, with a minimum inhibitory concentration (MIC) value of 64 microg/ml. Commercial oligonucleotide microarrays were used to determine the global transcriptional response of S. cerevisiae triggered by treatment with dictamnine. We interpreted our microarray data using the hierarchical clustering tool, T-profiler. Several major transcriptional responses were induced by dictamnine. The first was the induced environmental stress response, mainly under the control of the Msn2p and Msn4p transcription factors, and the repressed environmental stress response in genes containing the PAC (RNA polymerase A and C box) and rRPE (ribosomal RNA processing element) motifs. The second was the Upc2p-mediated response involved in lipid biosynthesis. The third comprised the PDR3- and RPN4-mediated responses involved in multidrug resistance (MDR). Finally, the TBP-mediated response was induced with dictamnine treatment. TBP is an essential general transcription factor involved in directing the transcription of genes. Quantitative real-time RT-PCR was performed on selected genes to verify the microarray results. Furthermore, morphological transitions during dictamnine exposure to S. cerevisiae L1190 (MATa/alpha) were examined, using confocal laser microscopy.
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Quinolinas/farmacología , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/metabolismo , Transcripción Genética/efectos de los fármacos , Perfilación de la Expresión Génica/métodos , Concentración 50 Inhibidora , Pruebas de Sensibilidad Microbiana , Microscopía Confocal , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN de Hongos/química , ARN de Hongos/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Saccharomyces cerevisiae/genéticaRESUMEN
In this study, we show that berberine chloride (BBR) has antimicrobial activities against all 43 tested strains of Staphylococcus aureus, an important human and animal pathogen. However, the response mechanisms of S. aureus to BBR are still poorly understood. Affymetrix GeneChips were used to determine the global transcription of S. aureus triggered by treatment with subinhibitory concentrations of BBR. 468 genes were up-regulated and 262 genes were down-regulated upon exposure to BBR. There was elevated transcription of various transporter genes, including genes involved in multidrug resistance, members of the multidrug and toxin extrusion family, the ferrous iron transporter, the amino acid transporter, the Na(+)/H(+) antiporter, and the potassium cation transporter. Measurements of active transport were used to demonstrate a phenotypic correlation between efflux transporter overexpression and inhibition of BBR uptake. Furthermore, BBR induced the expression of urease genes, sortase enzyme, and iron-regulated surface determinant genes, but repressed transcription of a gene encoding arylamine N-acetyltransferase activity (N315-SA2490). To our knowledge, this is the first analysis of a genome-wide transcription profile of S. aureus cells in response to BBR treatment. These results will pave the way to exploring the mechanisms of BBR against S. aureus.
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Antibacterianos/farmacología , Berberina/farmacología , Perfilación de la Expresión Génica , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genética , Berberina/metabolismo , Transporte Biológico Activo , Genes Bacterianos , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Bacteriano/biosíntesis , ARN Bacteriano/genética , Staphylococcus aureus/metabolismoRESUMEN
Staphylococcus aureus (S. aureus) is one of the most important pathogens in humans and animals, multiply resistant strains are increasingly widespread, new agents are needed for the treatment of S. aureus. Rhein (RH), a natural plant product, has been reported to have potential antimicrobial activity against S. aureus, but the response mechanisms of S. aureus to RH are still poorly understood. RH showed good in vitro antibacterial activity against all 21 tested S. aureus strains in this experiment. We performed commercial Affymetrix GeneChips to determine the overall transcriptional response of S. aureus ATCC25923 triggered by the treatment of subinhibitory concentrations of RH at one time point 45 min. A total of 88 genes were identified to be differentially regulated by RH. Of these, 28 transporter genes were differentially regulated by RH; RH stress elevated the transcription of genes (srtB and isdABCDEFGI) encoding iron-regulated surface determinants system and genes (nrdIEF and nrdDG) involved in ribonucleotide reductase systems; but RH repressed genes (pflAB, nirBDR, narGH, ldh1, COL-SA0660, COL-SA2363 and COL-SA2386) responsible for anaerobic respiration and fermentation. To our knowledge, this genome-wide transcriptomics approach revealed first insights into the response of S. aureus to RH challenge.
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Antraquinonas/farmacología , Productos Biológicos/farmacología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genética , Transcripción Genética/efectos de los fármacos , Anaerobiosis/efectos de los fármacos , Fermentación/efectos de los fármacos , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Genes Bacterianos , Hierro/farmacología , Análisis de Secuencia por Matrices de Oligonucleótidos , Consumo de Oxígeno/efectos de los fármacos , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ribonucleótido Reductasas/metabolismo , Staphylococcus aureus/enzimología , Staphylococcus aureus/crecimiento & desarrollo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Some antibacterials have been reported to regulate the host immune and inflammatory responses both in vitro and in vivo. Florfenicol is an antibiotics used in treatment of infection. We investigated the effects of florfenicol on cytokine production by lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages in vitro, and the results showed that florfenicol reduced tumor necrosis factor (TNF) and interleukin-6 (IL-6) production but had little effect on interleukin-1beta (IL-1beta) and interleukin IL-10 (IL-10) secretion. This inspired us to further study the effects of florfenicol in vivo. Florfenicol significantly attenuated TNF and IL-6 production in serum from mice challenged with LPS, and in consistent with the results in vitro. In murine model of endotoxemia, mice were prophylactically or therapeutically treated with florfenicol prior to or after LPS challenge. The results showed that florfenicol significantly increased mouse survival. Further studies revealed that florfenicol prevented the LPS-induced nuclear factor-kappaB (NF-kappaB) translocation from cytoplasm into nuclear in RAW 264.7 macrophages. These observations indicate that florfenicol modulates early cytokine responses by blocking NF-kappaB pathway, and thus, increases mouse survival. This effect of the drug may be of potential usefulness in treatment of bacterial shock.
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Antibacterianos/uso terapéutico , Citocinas/biosíntesis , Endotoxemia/tratamiento farmacológico , Tianfenicol/análogos & derivados , Animales , Células Cultivadas , Endotoxemia/inmunología , Endotoxemia/mortalidad , Femenino , Interleucina-6/antagonistas & inhibidores , Interleucina-6/biosíntesis , Ratones , Ratones Endogámicos C57BL , Tianfenicol/uso terapéutico , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
CRISPR/Cas9-mediated genome editing can inhibit virus infection by targeting the conserved regions of the viral genomic DNA. Unexpectedly, we found previously that pseudorabies virus (PRV) could escape from CRISPR/Cas9-mediated inhibition. In order to elucidate whether the escape of PRV from Cas9-mediated inhibition was due to cell deficiencies, such as genetic instability of sgRNA or Cas9 protein, the positive cells were passaged ten times, and PRV infection in the sgRNA-expressing cells was evaluated in the present study. The results showed that subculturing cells has no effect on Cas9-mediated cleavage of PRV. Different passages of PX459-PRV cells can stably express sgRNA to facilitate Cas9/sgRNA cleavage on the UL30 gene of PRV, resulting in a pronounced inhibition of PRV infection. Studies to elucidate the mechanism of PRV escape are currently in progress.
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This study was aimed to prepare the polyclonal antibody against the soluble proliferation-inducing ligand (sAPRIL) antigen and to investigate its effects in suppressing sAPRIL mediated lymphocyte proliferation. Mutated recombinant sAPRIL protein, which lacks biological activity but maintains immunogenicity, was used as antigen to immunize humanized SCID mice. Sera were obtained at 6 weeks after immunization. Indirect ELISA and Western blot were used to detect the antibody titer and specificity. The inhibition of polyclonal antibodies on Raji and Jurkat cell proliferation stimulated by sAPRIL was assessed by the MTT assay. The results showed that the mutant of sAPRIL could induce the production of polyclonal antibodies against human sAPRIL. Western blot and indirect ELISA analyses indicated that the anti-serum had higher specificity with a titer of 1:640. Functional analysis revealed that these polyclonal antibodies significantly inhibited the proliferation of Raji and Jurkat cell stimulated by sAPRIL (p < 0.05). It is concluded the polyclonal antibody against human sAPRIL is successfully prepared, which can inhibit the proliferation of Raji and Jurkat cells stimulated by sAPRIL in vitro.
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Anticuerpos/genética , Anticuerpos/farmacología , Miembro 13 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/genética , Animales , Anticuerpos/inmunología , Especificidad de Anticuerpos/inmunología , Proliferación Celular/efectos de los fármacos , Clonación Molecular , Humanos , Sueros Inmunes/análisis , Sueros Inmunes/inmunología , Células Jurkat , Ratones , Miembro 13 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/inmunologíaRESUMEN
Reports have shown that oleanolic acid (OA), a triterpenoid, exists widely in food, medicinal herbs and other plants, and that it has antimycobacterial activity against the Mycobacterium tuberculosis strain H37Rv (ATCC 27294). In this study it was found that OA had antimycobacterial properties against eight clinical isolates of M. tuberculosis and that the MICs of OA against drug-sensitive and drug-resistant isolates were 50-100 and 100-200 microg ml(-1), respectively. The combination of OA with isoniazid (INH), rifampicin (RMP) or ethambutol (EMB) showed favourable synergistic antimycobacterial effects against six drug-resistant strains, with fractional inhibitory concentration indices of 0.121-0.347, 0.113-0.168 and 0.093-0.266, respectively. The combination treatments of OA/INH, OA/RMP and OA/EMB displayed either a synergistic interaction or did not show any interaction against two drug-sensitive strains. No antagonism resulting from the OA/INH, OA/RMP or OA/EMB combination was observed for any of the strains tested. OA exhibited a relatively low cytotoxicity in Vero cells. These results indicate that OA may serve as a promising lead compound for future antimycobacterial drug development.