RESUMO
Gene expression noise (heterogeneity) leads to phenotypic diversity among isogenic individual cells. Our current understanding of gene expression noise is mostly limited to transcription, as separating translational noise from transcriptional noise has been challenging. It also remains unclear how translational heterogeneity originates. Using a transcription-normalized reporter system, we discovered that stop codon readthrough is heterogeneous among single cells, and individual cells with higher UGA readthrough grow faster from stationary phase. Our work also revealed that individual cells with lower protein synthesis levels exhibited higher UGA readthrough, which was confirmed with ribosome-targeting antibiotics (e.g., chloramphenicol). Further experiments and mathematical modeling suggest that varied competition between ternary complexes and release factors perturbs the UGA readthrough level. Our results indicate that fluctuations in the concentrations of translational components lead to UGA readthrough heterogeneity among single cells, which enhances phenotypic diversity of the genetically identical population and facilitates its adaptation to changing environments.
Assuntos
Códon de Terminação , Proteínas de Escherichia coli/biossíntese , Proteínas de Escherichia coli/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Genes Reporter , Microscopia de Fluorescência , Transferases de Grupo de Um Carbono , Proteínas de Bactérias/biossíntese , Proteínas de Bactérias/genética , Escherichia coli/crescimento & desenvolvimento , Regulação Bacteriana da Expressão Gênica , Aptidão Genética , Genótipo , Cinética , Proteínas Luminescentes/biossíntese , Proteínas Luminescentes/genética , Modelos Genéticos , Fenótipo , RNA Bacteriano/biossíntese , RNA Bacteriano/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Transcrição Gênica , Proteína Vermelha FluorescenteRESUMO
Acetylation is a global post-translational modification that regulates various cellular processes. Bacterial acetylomic studies have revealed extensive acetylation of ribosomal proteins. However, the role of acetylation in regulating ribosome function remains poorly understood. In this study, we systematically profiled ribosomal protein acetylation and identified a total of 289 acetylated lysine residues in 52 ribosomal proteins (r-proteins) from Salmonella Typhimurium. The majority of acetylated lysine residues of r-proteins were found to be regulated by both acetyltransferase Pat and metabolic intermediate acetyl phosphate. Our results show that acetylation plays a critical role in the assembly of the mature 70S ribosome complex by modulating r-proteins binding to rRNA. Moreover, appropriate acetylation is important for the interactions between elongation factors and polysomes, as well as regulating ribosome translation efficiency and fidelity. Dysregulation of acetylation could alter bacterial sensitivity to ribosome-targeting antibiotics. Collectively, our data suggest that the acetylation homeostasis of ribosomes is crucial for their assembly and function. Furthermore, this mechanism may represent a universal response to environmental signals across different cell types.
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Processamento de Proteína Pós-Traducional , Proteínas Ribossômicas , Salmonella typhimurium , Acetilação , Homeostase , Lisina/metabolismo , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Ribossomos/genética , Ribossomos/metabolismo , Salmonella typhimurium/metabolismoRESUMO
Obstructive sleep apnea (OSA) can lead to intestinal injury, endotoxemia, and disturbance of intestinal flora. Additionally, as a crucial component of the endocannabinoid system, some studies have demonstrated that cannabinoid 1 (CB1) receptors are closely linked to the multiple organ dysfunction triggered by OSA. However, the role of the CB1 receptor in alleviating OSA-induced colon injury remains unclear. Here, through the construction of the OSA classic model, we found that the colon tissue of chronic intermittent hypoxia (CIH)-induced mice exhibited an overexpression of the CB1 receptor. The results of hematoxylin-eosin staining and transmission electron microscopy revealed that inhibition of the CB1 receptor could decrease the gap between the mucosa and muscularis mucosae, alleviate mitochondrial swelling, reduce microvilli shedding, and promote the recovery of tight junctions of CIH-induced mice. Furthermore, CB1 receptor inhibition reduced the levels of metabolic endotoxemia and inflammatory responses, exhibiting significant protective effects on the colon injury caused by CIH. At the molecular level, through western blotting and real-time polymerase chain reaction techniques, we found that inhibiting the CB1 receptor can significantly increase the expression of ZO-1 and Occludin proteins, which are closely related to the maintenance of intestinal mucosal barrier function. Through 16S rRNA high-throughput sequencing and short-chain fatty acid (SCFA) determination, we found that inhibition of the CB1 receptor increased the diversity of the microbial flora and controlled the makeup of intestinal flora. Moreover, butyric acid concentration and the amount of SCFA-producing bacteria, such as Ruminococcaceae and Lachnospiraceae, were both markedly elevated by CB1 receptor inhibition. The results of the spearman correlation study indicated that Lachnospiraceae showed a positive association with both ZO-1 and Occludin but was negatively correlated with the colon CB1 receptor, IL-1ß, and TNF-α. According to this study, we found that inhibiting CB1 receptor can improve CIH-induced colon injury by regulating gut microbiota, reducing mucosal damage and promoting tight junction recovery. KEY POINTS: â¢CIH leads to overexpression of CB1 receptor in colon tissue. â¢CIH causes intestinal flora disorder, intestinal mucosal damage, and disruption of tight junctions. â¢Inhibition of CB1 receptor can alleviate the colon injury caused by CIH through regulating the gut microbiota, reducing mucosal injury, and promoting tight junction recovery.
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Colo , Modelos Animais de Doenças , Mucosa Intestinal , Receptor CB1 de Canabinoide , Animais , Receptor CB1 de Canabinoide/metabolismo , Receptor CB1 de Canabinoide/genética , Camundongos , Colo/patologia , Colo/microbiologia , Colo/metabolismo , Masculino , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Hipóxia/metabolismo , Camundongos Endogâmicos C57BL , Proteína da Zônula de Oclusão-1/metabolismo , Ocludina/metabolismo , Ocludina/genética , Microbioma Gastrointestinal , Junções Íntimas/metabolismoRESUMO
OBJECTIVES: The current guidelines contain substantial inconsistency regarding the use of metformin concomitantly with contrast media. The objective of this study is to appraise the guidelines and summarize the agreements and differences among recommendations. METHODS: Our search focused on English language guidelines published between 2018 and 2021. Guidelines for the management of contrast media in patients with continuous metformin were included. Guidelines were assessed using the Appraisal of Guidelines for Research and Evaluation II instrument. RESULTS: Six guidelines out of 1134 fulfilled the inclusion criteria with an AGREE II score of 79.2% (IQR 72.7 to 85.1%). There was good overall quality of the guidelines, with six considered "strongly recommended." CPGs scored poorly in "Clarity of Presentation" and "Applicability," with scores of 75.9% and 76.4%, respectively. The intraclass correlation coefficients were excellent in each domain. There are some guidelines (33.3%) that recommend discontinuation of metformin in patients with an eGFR of < 30 mL/min/1.73 m2, while some guidelines (16.7%) suggest the threshold of renal function should be eGFR < 40 mL/min/1.73 m2. CONCLUSIONS: Most guidelines recommend withdrawing metformin before using contrast agents in diabetic patients with severely impaired kidney function but disagree on the renal function thresholds. Furthermore, the gaps regarding discontinuing metformin with moderate renal impairment (30 mL/min/1.73 m2 < eGFR < 60 mL/min/1.73 m2) must be considered in future studies. KEY POINTS: ⢠Guidelines involving metformin and contrast agents are reliable and optimal. ⢠Most guidelines advocate discontinuing metformin before using contrast agents in diabetic patients with advanced renal failure, but there are controversial suggestions regarding kidney function thresholds. ⢠The gaps regarding the time of discontinuation of the metformin with moderate renal impairment (30 mL/min/1.73 m2 < eGFR < 60 mL/min/1.73 m2) must be considered in the extensive RCT studies.
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Diabetes Mellitus , Metformina , Insuficiência Renal , Humanos , Metformina/uso terapêutico , Meios de Contraste , ConsensoRESUMO
Accurate protein synthesis is a tightly controlled biological process with multiple quality control steps safeguarded by aminoacyl-transfer RNA (tRNA) synthetases and the ribosome. Reduced translational accuracy leads to various physiological changes in both prokaryotes and eukaryotes. Termination of translation is signaled by stop codons and catalyzed by release factors. Occasionally, stop codons can be suppressed by near-cognate aminoacyl-tRNAs, resulting in protein variants with extended C termini. We have recently shown that stop-codon readthrough is heterogeneous among single bacterial cells. However, little is known about how environmental factors affect the level and heterogeneity of stop-codon readthrough. In this study, we have combined dual-fluorescence reporters, mass spectrometry, mathematical modeling, and single-cell approaches to demonstrate that a metabolic stress caused by excess carbon substantially increases both the level and heterogeneity of stop-codon readthrough. Excess carbon leads to accumulation of acid metabolites, which lower the pH and the activity of release factors to promote readthrough. Furthermore, our time-lapse microscopy experiments show that single cells with high readthrough levels are more adapted to severe acid stress conditions and are more sensitive to an aminoglycoside antibiotic. Our work thus reveals a metabolic stress that promotes translational heterogeneity and phenotypic diversity.
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Códon de Terminação , Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Glucose/farmacologia , Concentração de Íons de Hidrogênio , MutaçãoRESUMO
Electrobiocorrosion, the process in which microbes extract electrons from metallic iron (Fe0 ) through direct Fe0 -microbe electrical connections, is thought to contribute to the costly corrosion of iron-containing metals that impacts many industries. However, electrobiocorrosion mechanisms are poorly understood. We report here that electrically conductive pili (e-pili) and the conductive mineral magnetite play an important role in the electron transfer between Fe0 and Geobacter sulfurreducens, the first microbe in which electrobiocorrosion has been rigorously documented. Genetic modification to express poorly conductive pili substantially diminished corrosive pitting and rates of Fe0 -to-microbe electron flux. Magnetite reduced resistance to electron transfer, increasing corrosion currents and intensifying pitting. Studies with mutants suggested that the magnetite promoted electron transfer in a manner similar to the outer-surface c-type cytochrome OmcS. These findings, and the fact that magnetite is a common product of iron corrosion, suggest a potential positive feedback loop of magnetite produced during corrosion further accelerating electrobiocorrosion. The interactions of e-pili, cytochromes, and magnetite demonstrate mechanistic complexities of electrobiocorrosion, but also provide insights into detecting and possibly mitigating this economically damaging process.
Assuntos
Óxido Ferroso-Férrico , Geobacter , Oxirredução , Elétrons , Corrosão , Transporte de Elétrons , Citocromos/metabolismo , Ferro , Geobacter/genética , Geobacter/metabolismoRESUMO
PURPOSE: We aimed to develop a simple risk score for patients with HFpEF and assessed the efficacy of spironolactone across baseline risk. METHODS: We developed risk stratification scheme for cardiovascular death in placebo arm of the Treatment of Preserved Cardiac Function Heart Failure with an Aldosterone Antagonist trial (TOPCAT). We screened candidate risk indicators and determined strong risk predictors using COX regression. The absolute risk reduction (ARR) in cardiovascular death with spironolactone was evaluated across baseline risk groups. COX regressions were performed to assess the hazard ratios (HRs) of spironolactone therapy for cardiovascular death and drug discontinuation in each risk category. RESULTS: A simple risk score scheme was constructed based on five risk indicators weighted by estimates from the model, including age, diastolic blood pressure, renal dysfunction, white blood cell, and left ventricular ejection fraction. The risk score scheme showed good discrimination in placebo cohort (C index=0.70). ARR with spironolactone therapy was observed only in patients at very high risk (7.9%). Spironolactone therapy significantly reduced the risk of cardiovascular death in the very high-risk group (HR: 0.57; 95%CI, 0.39-0.84; P =0.005 and P for interaction 0.03) but showed similar risk of drug discontinuation across risk categories (P for interaction=0.928). CONCLUSION: This simple risk score stratifies patients with HFpEF by their baseline risk of cardiovascular death. Patients at very high risk derive great benefits from spironolactone therapy. This easy-to-use risk score provides a practical tool that can facilitate risk stratification and tailoring therapy for those who benefit most from spironolactone. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT00094302.
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Insuficiência Cardíaca , Espironolactona , Insuficiência Cardíaca/diagnóstico , Insuficiência Cardíaca/tratamento farmacológico , Hospitalização , Humanos , Antagonistas de Receptores de Mineralocorticoides/efeitos adversos , Medição de Risco , Espironolactona/efeitos adversos , Volume Sistólico/fisiologia , Resultado do Tratamento , Função Ventricular EsquerdaRESUMO
BACKGROUND: In most situations, many patients undergoing coronary artery bypass graft (CABG) are on dual antiplatelet therapy (DAPT), which is also required after CABG. The adjustment of antiplatelet strategy remains controversial. In this study, we systematically review current guidelines, seeking consensus and controversies to facilitate clinical practice. METHODS AND RESULTS: Guidelines are searched in PubMed, Embase, ECRI Guidelines Trust and websites of guidelines organizations and professional society. Guidelines with recommendations of DAPT for patients undergo CABG are included. Two reviewers appraised guidelines with the Appraisal of Guidelines for Research and Evaluation II (AGREE II). Relevant recommendations are extracted and summarized. A total of 14 guidelines meeting inclusion criteria are selected, with average AGREE II scores from 44% to 86%. Most guidelines score high in domains other than 'applicability'. Many guidelines are not detailed enough in reporting considerations behind recommendations. Current guidelines are consistent on the management of antiplatelet strategy before elective CABG and using DAPT after surgery for preventing graft vessel occlusion. Evidence is still lacking in urgent CABG and resumption of the previous DAPT after surgery. CONCLUSIONS: Current guidelines on DAPT in CABG are generally satisfying. Suspending P2Y12 inhibitors while aspirin continued before elective CABG is recommended, as well as 12 months of DAPT following CABG. More evidence is needed to guide antiplatelet therapy in urgent CABG and to prove the benefits of resuming previous DAPT.
Assuntos
Ponte de Artéria Coronária/métodos , Doença da Artéria Coronariana/cirurgia , Terapia Antiplaquetária Dupla/métodos , Guias de Prática Clínica como Assunto , Aspirina/uso terapêutico , Desprescrições , Duração da Terapia , Procedimentos Cirúrgicos Eletivos , Emergências , Humanos , Inibidores da Agregação Plaquetária/uso terapêutico , Hemorragia Pós-Operatória/prevenção & controle , Antagonistas do Receptor Purinérgico P2Y/uso terapêuticoRESUMO
BACKGROUND: To examine the association of low educational attainment with incident heart failure (HF) and explore potential behavioral mediators of the causal pathway.MethodsâandâResults:A total of 12,109 participants in the Atherosclerosis Risk in Communities Study (ARIC) were included. Educational attainment was measured at baseline, and the risk of HF across educational attainment groups was assessed by Cox proportional hazards models. Using mediation analysis, we evaluated the mediating role of behavioral factors in the causal pathway between educational attainment and HF. During a median follow-up of 25.1 years, 2,407 cases (19.9%) of HF occurred. Educational attainment showed an inverse association with HF risk (hazard ratio (HR), 1.41; 95% confidence interval (CI), 1,26-1.57 for low educational attainment; HR, 1.13; 95% CI, 1.02-1.25 for medium educational attainment). In the mediation analysis, the association between educational attainment and HF was partially mediated by income, waist-to-hip ratio, current smoking, body mass index, current drinking, sports and physical activity, which explained 24.3%, 20.2%, 13.8%, 10.1%, 7.7%, 7.3% and 4.5%, respectively, of the relationship. In total, all mediators contributed 56.3% of the total effect. CONCLUSIONS: Low educational attainment was associated with increased risk for HF. Income, obesity and current smoking mediated a great proportion of the total effect of educational attainment on HF. Our results provide underlying insights for the development of targeted public health interventions to reduce educational disparities on HF incidence.
Assuntos
Insuficiência Cardíaca , Análise de Mediação , Índice de Massa Corporal , Exercício Físico , Insuficiência Cardíaca/complicações , Humanos , Obesidade/complicações , Obesidade/epidemiologiaRESUMO
Translational fidelity is required for accurate flow of genetic information, but is frequently altered by genetic changes and environmental stresses. To date, little is known about how translational fidelity affects the virulence and host interactions of bacterial pathogens. Here we show that surprisingly, either decreasing or increasing translational fidelity impairs the interactions of the enteric pathogen Salmonella Typhimurium with host cells and its fitness in zebrafish. Host interactions are mediated by Salmonella pathogenicity island 1 (SPI-1). Our RNA sequencing and quantitative RT-PCR results demonstrate that SPI-1 genes are among the most down-regulated when translational fidelity is either increased or decreased. Further, this down-regulation of SPI-1 genes depends on the master regulator HilD, and altering translational fidelity destabilizes HilD protein via enhanced degradation by Lon protease. Our work thus reveals that optimal translational fidelity is pivotal for adaptation of Salmonella to the host environment, and provides important mechanistic insights into this process.
Assuntos
Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Ilhas Genômicas , Interações entre Hospedeiro e Microrganismos , Protease La/metabolismo , Salmonella typhimurium/patogenicidade , Fatores de Transcrição/metabolismo , Animais , Linhagem Celular , Regulação para Baixo , Genoma Bacteriano , Humanos , Macrófagos/metabolismo , Camundongos , Salmonella typhimurium/genética , Análise de Sequência de RNA , Virulência , Fatores de Virulência/genética , Peixe-ZebraRESUMO
The effect of renin-angiotensin-aldosterone system (RAAS) blockers [angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers] on Contrast-induced nephropathy (CIN) is unclear in patients with renal insufficiency. Thus, we conduct a meta-analysis to evaluate the association between the administration of RAAS blockers and CIN in patients with renal insufficiency. We searched PubMed, EMBASE, and Cochrane Library for relevant studies published before September 2019. The primary outcome was the incidence of CIN, and the secondary outcome was the changes in serum creatinine (SCr) from baseline to postprocedure (ΔSCr). Pooled odds ratio (OR) or weighted mean difference (WMD) with their 95% confidence interval (CIs) for the CIN incidence, ΔSCr were used to calculate original data. A total of 8 studies were included in the meta-analysis. Compared with controls, ACEI/angiotensin receptor blocker increased the risk of CIN (OR = 1.61, 95% CI 1.14-2.28, I = 30%; P = 0.007), whereas this association was not significant in Chinese patients (OR = 1.07, 95% CI 0.65-1.77, I = 19%, P = 0.79). The total weighted mean differences of the ΔSCr were 0.06 mg/dL (95% CI: 0.01-0.11, I = 82%; P = 0.03). Administration of RAAS blockers in patients with renal insufficiency was associated with a significantly higher incidence of CIN, whereas it did not show a significant effect on Chinese patients.
Assuntos
Injúria Renal Aguda/prevenção & controle , Antagonistas de Receptores de Angiotensina/uso terapêutico , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Meios de Contraste/efeitos adversos , Rim/efeitos dos fármacos , Insuficiência Renal/complicações , Sistema Renina-Angiotensina/efeitos dos fármacos , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/diagnóstico , Injúria Renal Aguda/fisiopatologia , Antagonistas de Receptores de Angiotensina/efeitos adversos , Inibidores da Enzima Conversora de Angiotensina/efeitos adversos , Humanos , Rim/patologia , Rim/fisiopatologia , Insuficiência Renal/diagnóstico , Insuficiência Renal/fisiopatologia , Medição de Risco , Fatores de Risco , Resultado do TratamentoRESUMO
BACKGROUND: The protein synthesis machinery uses 22 natural amino acids as building blocks that faithfully decode the genetic information. Such fidelity is controlled at multiple steps and can be compromised in nature and in the laboratory to rewire protein synthesis with natural and synthetic amino acids. SCOPE OF REVIEW: This review summarizes the major quality control mechanisms during protein synthesis, including aminoacyl-tRNA synthetases, elongation factors, and the ribosome. We will discuss evolution and engineering of such components that allow incorporation of natural and synthetic amino acids at positions that deviate from the standard genetic code. MAJOR CONCLUSIONS: The protein synthesis machinery is highly selective, yet not fixed, for the correct amino acids that match the mRNA codons. Ambiguous translation of a codon with multiple amino acids or complete reassignment of a codon with a synthetic amino acid diversifies the proteome. GENERAL SIGNIFICANCE: Expanding the genetic code with synthetic amino acids through rewiring protein synthesis has broad applications in synthetic biology and chemical biology. Biochemical, structural, and genetic studies of the translational quality control mechanisms are not only crucial to understand the physiological role of translational fidelity and evolution of the genetic code, but also enable us to better design biological parts to expand the proteomes of synthetic organisms. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue.
Assuntos
Aminoácidos , Códon , Biossíntese de Proteínas/fisiologia , Engenharia de Proteínas/métodos , Biologia Sintética/métodos , Aminoácidos/síntese química , Aminoácidos/metabolismo , Animais , Clonagem Molecular/métodos , Códon/síntese química , Códon/química , Códon/metabolismo , Código Genético/fisiologia , Humanos , Modelos MolecularesRESUMO
Accurate flow of genetic information from DNA to protein requires faithful translation. An increased level of translational errors (mistranslation) has therefore been widely considered harmful to cells. Here we demonstrate that surprisingly, moderate levels of mistranslation indeed increase tolerance to oxidative stress in Escherichia coli. Our RNA sequencing analyses revealed that two antioxidant genes katE and osmC, both controlled by the general stress response activator RpoS, were upregulated by a ribosomal error-prone mutation. Mistranslation-induced tolerance to hydrogen peroxide required rpoS, katE and osmC. We further show that both translational and post-translational regulation of RpoS contribute to peroxide tolerance in the error-prone strain, and a small RNA DsrA, which controls translation of RpoS, is critical for the improved tolerance to oxidative stress through mistranslation. Our work thus challenges the prevailing view that mistranslation is always detrimental, and provides a mechanism by which mistranslation benefits bacteria under stress conditions.
Assuntos
Escherichia coli/metabolismo , Estresse Oxidativo , Biossíntese de Proteínas , Escherichia coli/genética , Peróxido de Hidrogênio/metabolismo , Mutação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ribossomos/metabolismoRESUMO
Aminoacyl-tRNA synthetases maintain the fidelity during protein synthesis by selective activation of cognate amino acids at the aminoacylation site and hydrolysis of misformed aminoacyl-tRNAs at the editing site. Threonyl-tRNA synthetase (ThrRS) misactivates serine and utilizes an editing site cysteine (C182 in Escherichia coli) to hydrolyze Ser-tRNA(Thr). Hydrogen peroxide oxidizes C182, leading to Ser-tRNA(Thr) production and mistranslation of threonine codons as serine. The mechanism of C182 oxidation remains unclear. Here we used a chemical probe to demonstrate that C182 was oxidized to sulfenic acid by air, hydrogen peroxide and hypochlorite. Aminoacylation experiments in vitro showed that air oxidation increased the Ser-tRNA(Thr) level in the presence of elongation factor Tu. C182 forms a putative metal binding site with three conserved histidine residues (H73, H77 and H186). We showed that H73 and H186, but not H77, were critical for activating C182 for oxidation. Addition of zinc or nickel ions inhibited C182 oxidation by hydrogen peroxide. These results led us to propose a model for C182 oxidation, which could serve as a paradigm for the poorly understood activation mechanisms of protein cysteine residues. Our work also suggests that bacteria may use ThrRS editing to sense the oxidant levels in the environment.
Assuntos
Proteínas de Escherichia coli/química , Oxidantes/farmacologia , Treonina-tRNA Ligase/química , Aminoacilação de RNA de Transferência , Cisteína/química , Proteínas de Escherichia coli/metabolismo , Histidina/química , Peróxido de Hidrogênio/farmacologia , Estresse Oxidativo , Fator Tu de Elongação de Peptídeos/metabolismo , RNA de Transferência de Treonina/metabolismo , Serina/metabolismo , Ácidos Sulfênicos/química , Treonina-tRNA Ligase/metabolismoRESUMO
Accurate quantification of bacterial burden within macrophages, termed bacterial burden quantification (BBQ), is crucial for understanding host-pathogen interactions. Various methods have been employed, each with strengths and weaknesses. This article addresses limitations in existing techniques and introduces two novel, automated methods for BBQ within macrophages based on confocal microscopy data analysis. The first method refines total fluorescence quantification by incorporating filtering steps to exclude uninfected cells, while the second method calculates total bacterial volume per cell to mitigate potential biases in fluorescence-based readouts. These workflows utilize PyImageJ and Cellpose software, providing reliable, unbiased, and rapid quantification of bacterial load. The proposed workflows were validated using Salmonella enterica serovar Typhimurium and Mycobacterium tuberculosis models, demonstrating their effectiveness in accurately assessing bacterial burden. These automated workflows offer valuable tools for studying bacterial interactions within host cells and provide insights for various research applications.
Assuntos
Macrófagos , Salmonella typhimurium , Fluxo de Trabalho , Interações Hospedeiro-PatógenoRESUMO
BACKGROUND: Studies have revealed the beneficial effects of sodium-glucose co-transporter 2 inhibitors (SGLT2i) for the treatment of heart failure (HF) regardless of the presence of diabetes. Besides, SGLT2i can decrease the incidence of atrial fibrillation (AF) in a broad population. However, the effects of SGLT2i on AF recurrence following catheter ablation (CA) remain uncertain. Therefore, this meta-analysis was undertaken to elucidate the effects of SGLT2i on AF recurrence after CA in AF patients. METHODS: A comprehensive search of PubMed, Embase, and Cochrane library was conducted for relevant studies, encompassing data from inception until March 20, 2024. The data were pooled using a fixed-effects model if the I2 value was <50%; otherwise, a random-effects model was adopted. RESULTS: One randomized controlled trial (RCT) and five observational studies involving 5623 patients with AF who underwent CA were included. SGLT2i treatment was associated with a significantly lower rate of AF recurrence (odds ratio [OR] = 0.45, 95% confidence interval [CI]: 0.31-0.66). Subgroup analysis demonstrated that patients treated with SGLT2i exhibited a lower incidence of AF recurrence compared to those treated with dipeptidyl peptidase-4 inhibitors (DPP4i). The favorable effects of SGLT2i on AF recurrence were more pronounced in male patients and patients with persistent AF. CONCLUSIONS: This meta-analysis provided evidence supporting the effectiveness of SGLT2i in reducing the risk of AF recurrence after CA in AF patients. SGLT2i may serve as an additional therapy option in this population.
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Oceanic facilities and equipment corrosion present considerable economic and safety concerns, predominantly due to microbial corrosion. Early detection of corrosive microbes is pivotal for effective monitoring and prevention. Yet, traditional detection methods often lack specificity, require extensive processing time, and yield inaccurate results. Hence, the need for an efficient real-time corrosive microbe monitoring technology is evident. Pseudomonas aeruginosa, a widely distributed microorganism in aquatic environments, utilizes its production of quinone-like compounds, specifically pyocyanin (PYO), to corrode metals. Here, we report a novel fiber optic surface plasmon resonance (SPR) sensor modified by the C-terminal of BrlR protein (BrlR-C), which is a specific receptor of PYO molecule, to detect P. aeruginosa in aquatic environments. The results showed that the sensor had a good ability to recognize PYO in the concentration range of 0-1 µg/mL, and showed excellent sensing performance in real-time monitoring the growth status of P. aeruginosa. With a strong selectivity of PYO, the sensor could clearly detect P. aeruginosa against other bacteria in seawater environment, and exhibited excellent anti-interference ability against variations in pH, temperature and pressure and other interfering substances. This study provides a useful tool for monitoring corrosive P. aeruginosa biofilm in aquatic environments, which is a first of its kind example that serves as a laboratory model for the application of fiber optic technology in real-world scenarios to monitoring biofilms in microbial corrosion and biofouling.
Assuntos
Biofilmes , Técnicas Biossensoriais , Tecnologia de Fibra Óptica , Pseudomonas aeruginosa , Piocianina , Ressonância de Plasmônio de Superfície , Pseudomonas aeruginosa/isolamento & purificação , Ressonância de Plasmônio de Superfície/métodos , Piocianina/análise , Piocianina/química , Técnicas Biossensoriais/métodos , Corrosão , Fibras Ópticas , Água do Mar/microbiologia , Água do Mar/química , Desenho de EquipamentoRESUMO
BACKGROUND: Gap junction remodeling is an important cause of ventricular arrhythmia in heart failure. However, it remains unclear whether renal denervation (RDN) regulates gap junction remodeling in heart failure. To explore the effect of RDN on gap junction remodeling in dogs with high-pacing-induced heart failure. METHODS: Fifteen dogs were randomly divided into control (n = 5), heart failure (HF) (n = 5), and RDN+HF (n = 5) group. A high-pacing-induced-heart failure model was established using rapid right ventricular pacing for 4 weeks. The RDN+HF group underwent surgical and chemical ablation of both renal arteries before 4 weeks rapid right ventricular pacing. After 4 weeks, echocardiography, High-Performance Liquid Chromatography-Mass Spectrometry test for norepinephrine and epinephrine, and pathological analysis were performed in the above 3 groups. Further, immunohistochemical staining was used to detect tyrosine hydroxylase, ChaT, connexin 43 (Cx43), and connexin 40 (Cx40). Connexin 43 and Cx40 expression was detected by western blotting. Transmission electron microscopy was used to observe the gap junction. RESULTS: Compared to the control group, myocardial fibrosis and sympathetic hyperactivity were observed in the HF group. Immunohistochemical staining and western blotting showed that Cx40 expression and Cx43 expression was significantly reduced in the HF group. Compared with the HF group, the RDN+HF group showed reduced sympathetic hyperactivity, Cx40 expression, Cx40/Cx43 ratio, and increased Cx43 expression. CONCLUSION: Renal denervation alleviates gap junction remodeling in high-pacing-induced heart failure dogs.
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Surface coatings are designed to mitigate pervasive biofouling herald, a new era of surface protection in complex biological environments. However, existing strategies are plagued by persistent and recurrent biofilm attachment, despite the use of bactericidal agents. Herein, a chiral metal-organic framework (MOF)-based coating with conformal microstructures to enable a new anti-biofouling mode that involves spontaneous biofilm disassembly followed by bacterial eradication is developed. A facile and universal metal-polyphenol network (MPN) is designed to robustly anchor the MOF nanoarmor of biocidal Cu2+ ions and anti-biofilm d-amino acid ligands to a variety of substrates across different material categories and surface topologies. Incorporating a diverse array of chiral amino acids endows the resultant coatings with widespread signals for biofilm dispersal, facilitating copper-catalyzed chemodynamic reactions and inherent mechano-bactericidal activities. This synergistic mechanism yields unprecedented anti-biofouling efficacy elucidated by RNA-sequencing transcriptomics analysis, enhancing broad-spectrum antibacterial activities, preventing biofilm formation, and destroying mature biofilms. Additionally, the chelation-directed amorphous/crystalline coatings can activate photoluminescent properties to inhibit the settlement of microalgae biofilms. This study provides a distinctive perspective on chirality-enhanced antimicrobial behaviors and pioneers a rational pathway toward developing next-generation anti-biofouling coatings for diverse applications.
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Microbially influenced corrosion (MIC) is a formidable challenge in the marine industry, resulting from intricate interactions among various biochemical reactions and microbial species. Many preventions used to mitigate biocorrosion fail due to ignorance of the MIC mechanisms. This review provides a summary of the current research on microbial corrosion in marine environments, including corrosive microbes and biocorrosion mechanisms. We also summarized current strategies for inhibiting MIC and proposed future research directions for MIC mechanisms and prevention. This review aims to comprehensively understand marine microbial corrosion and contribute to novel strategy developments for biocorrosion control in marine environments.