RESUMO
BACKGROUND: The ability of antimicrobial agents to affect microbial adherence to eukaryotic cell surfaces is a promising antivirulence strategy for combating the global threat of antimicrobial resistance. Inadequate use of antimicrobials has led to widespread instances of suboptimal antibiotic concentrations around infection sites. Therefore, we aimed to examine the varying effect of an antimicrobial peptidase lysostaphin (APLss) on staphylococcal adherence to host cells, biofilm biomass formation, and toxin production as a probable method for mitigating staphylococcal virulence. RESULTS: Initially, soluble expression in E. coli and subsequent purification by immobilized-Ni2+ affinity chromatography (IMAC) enabled us to successfully produce a large quantity of highly pure ~ 28-kDa His-tagged mature APLss. The purified protein exhibited potent inhibitory effects against both methicillin-sensitive and methicillin-resistant staphylococcal strains, with minimal inhibitory concentrations (MICs) ranging from 1 to 2 µg/mL, and ultrastructural analysis revealed that APLss-induced concentration-specific changes in the morphological architecture of staphylococcal surface membranes. Furthermore, spectrophotometric and fluorescence microscopy revealed that incubating staphylococcal strains with sub-MIC and MIC of APLss significantly inhibited staphylococcal adherence to human vaginal epithelial cells and biofilm biomass formation. Ultimately, transcriptional investigations revealed that APLss inhibited the expression of agrA (quorum sensing effector) and other virulence genes related to toxin synthesis. CONCLUSIONS: Overall, APLss dose-dependently inhibited adhesion to host cell surfaces and staphylococcal-associated virulence factors, warranting further investigation as a potential anti-staphylococcal agent with an antiadhesive mechanism of action using in vivo models of staphylococcal toxic shock syndrome.
Assuntos
Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Humanos , Lisostafina/farmacologia , Lisostafina/metabolismo , Escherichia coli/genética , Antibacterianos/farmacologia , Antibacterianos/metabolismo , Staphylococcus , Biofilmes , Testes de Sensibilidade MicrobianaRESUMO
BACKGROUND: In ipecac (Carapichea ipecacuanha (Brot.) L. Andersson), adventitious shoots can be induced simply by placing internodal segments on phytohormone-free culture medium. The shoots form locally on the epidermis of the apical region of the segments, but not the basal region. Levels of endogenous auxin and cytokinin transiently increase in the segments after 1 week of culture. RESULTS: Here, we conducted RNA-seq analysis to compare gene expression patterns in apical and basal regions of segments before culture and after 1 week of culture for adventitious shoot formation. The results revealed 8987 differentially expressed genes in a de novo assembly of 76,684 genes. Among them, 276 genes were upregulated in the apical region after 1 week of culture relative to before culture and the basal region after 1 week of culture. These genes include 18 phytohormone-response genes and shoot-formation-related genes. Validation of the gene expression by quantitative real-time PCR assay confirmed that the expression patterns were similar to those of the RNA-seq data. CONCLUSIONS: The transcriptome data show that expression of cytokinin biosynthesis genes is induced along with the acquisition of cellular pluripotency and the initiation of cell division by wounding in the apical region of internodal segments, that trigger adventitious shoot formation without callusing.
Assuntos
Ácidos Indolacéticos , Ipeca , Citocininas/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Ipeca/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Brotos de Planta/genética , Brotos de Planta/metabolismoRESUMO
To investigate the changes in myosin heavy chain (MyHC) isoforms of rat masseter muscle fibres caused by chronic sleep deprivation and a possible link with the pathogenesis of disorders of the temporomandibular joint (TMJ). A total of 180 male rats were randomly divided into three groups (n=60 in each): cage controls, large platform controls, and chronic sleep deprivation group. Each group was further divided into three subgroups with different observation periods (7, 14, and 21 days). We investigated he expression of MyHC isoforms in masseter muscle fibres by real-time quantitative polymerase chain reaction (PCR), Western blotting, and immunohistochemical staining. In rats with chronic sleep deprivation there was increased MyHC-I expression in layers of both shallow and deep muscles at 7 and 21 days compared with the control groups, whereas sleep deprivation was associated with significantly decreased MyHC-II expression. At 21 days, there were no differences in MyHC-I or MyHC-II expression between the groups and there were no differences between the two control groups at any time point. These findings suggest that chronic sleep deprivation alters the expression of MyHC isoforms, which may contribute to the pathogenesis of disorders of the TMJ.