Exploiting Mass Spectrometry to Unlock the Mechanism of Nanoparticle-Induced Inflammasome Activation

Nanoparticles (NPs) elicit sterile inflammation, but the underlying signaling pathways are poorly understood. Here, we report that human monocytes are particularly vulnerable to amorphous silica NPs, as evidenced by single-cell-based analysis of peripheral blood mononuclear cells using cytometry by time-of-flight (CyToF), while silane modification of the NPs mitigated their toxicity. Using human THP-1 cells as a model, we observed cellular internalization of silica NPs by nanoscale secondary ion mass spectrometry (nanoSIMS) and this was confirmed by transmission electron microscopy. Lipid droplet accumulation was also noted in the exposed cells. Furthermore, time-of-flight secondary ion mass spectrometry (ToF-SIMS) revealed specific changes in plasma membrane lipids, including phosphatidylcholine (PC) in silica NP-exposed cells, and subsequent studies suggested that lysophosphatidylcholine (LPC) acts as a cell autonomous signal for inflammasome activation in the absence of priming with a microbial ligand. Moreover, we found that silica NPs elicited NLRP3 inflammasome activation in monocytes, whereas cell death transpired through a non-apoptotic, lipid peroxidation-dependent mechanism. Together, these data further our understanding of the mechanism of sterile inflammation

Isolation, characterization, statistical optimization, and application of a novel broad-spectrum capsular depolymerase against Klebsiella pneumoniae from Bacillus siamensis SCVJ30

Background: Bacterial resistance is one of the most challenging and emerging public healthcare crisis in the modern era. Along with antibiotic degrading strategies, bacteria also evolved to produce extracellular polymers such as capsular polysaccharide (CPS) which not only provides immune protection but also act as a permeability barrier to antibiotics. The use of therapeutic enzymes alone and in combination with antibiotics has opened a new window for clinicians and researchers. Methods: In the present study, isolation of broad-spectrum capsular depolymerase bacterium was attempted from a number of environmental samples followed by 16SrRNA characterization. Optimization of capsular depolymerase production was performed by the one variable at time (OVAT) method and response surface methodology (RSM). Capsular depolymerase was partially purified using ammonium sulfate saturation method. Capsule stripping effects of depolymerase were analyzed using microscopic visualization of the capsule and antibiotic susceptibility test. Results: Thirty-two capsular depolymerase producing bacteria were isolated in this study and broad-spectrum depolymerase producing Isolate-30 was characterized as Bacillus siamensis SCVJ30 according to the 16srRNA sequencing. Depolymerase production was optimized using OVAT method and RSM. Relatively high yields (1.92 IU/ml) of capsular depolymerase were obtained in a medium containing 1 mg magnesium sulfate, 7 mg peptone and at 9 pH. A 115% increase in capsular depolymerase production was observed under optimal conditions than unoptimized conditions. Microscopic visualization of the capsule and antibiotic susceptibility testing postulates the positive effect of depolymerase on antibiotic effi cacy against Klebsiella pneumoniae. Conclusion: Further characterization of the enzyme will help in developing broad-spectrum depolymerase as a potent therapeutic agent against drug-resistant strains

Label-free detection of polystyrene nanoparticles in Daphnia magna using Raman confocal mapping

Micro- and nanoplastic pollution has emerged as a global environmental problem. Moreover, plastic particles are of increasing concern for human health. However, the detection of so-called nanoplastics in relevant biological compartments remains a challenge. Here we show that Raman confocal spectroscopy-microscopy can be deployed for the non-invasive detection of amine-functionalized and carboxy-functionalized polystyrene (PS) nanoparticles (NPs) in Daphnia magna. The presence of PS NPs in the gastrointestinal (GI) tract of D. magna was confirmed by using transmission electron microscopy. Furthermore, we investigated the ability of NH2-PS NPs and COOH-PS NPs to disrupt the epithelial barrier of the GI tract using the human colon adenocarcinoma cell line HT-29. To this end, the cells were differentiated for 21 days and then exposed to PS NPs followed by cytotoxicity assessment and transepithelial electrical resistance measurements. A minor disruption of barrier integrity was noted for COOH-PS NPs, but not for the NH2-PS NPs, while no overt cytotoxicity was observed for both NPs. This study provides evidence of the feasibility of applying label-free approaches, i.e., confocal Raman mapping, to study PS NPs in a biological system

Exploiting Mass Spectrometry to Unlock the Mechanism of Nanoparticle-Induced Inflammasome Activation

Nanoparticles (NPs) elicit sterile inflammation, but the underlying signaling pathways are poorly understood. Here, we report that human monocytes are particularly vulnerable to amorphous silica NPs, as evidenced by single-cell-based analysis of peripheral blood mononuclear cells using cytometry by time-of-flight (CyToF), while silane modification of the NPs mitigated their toxicity. Using human THP-1 cells as a model, we observed cellular internalization of silica NPs by nanoscale secondary ion mass spectrometry (nanoSIMS) and this was confirmed by transmission electron microscopy. Lipid droplet accumulation was also noted in the exposed cells. Furthermore, time-of-flight secondary ion mass spectrometry (ToF-SIMS) revealed specific changes in plasma membrane lipids, including phosphatidylcholine (PC) in silica NP-exposed cells, and subsequent studies suggested that lysophosphatidylcholine (LPC) acts as a cell autonomous signal for inflammasome activation in the absence of priming with a microbial ligand. Moreover, we found that silica NPs elicited NLRP3 inflammasome activation in monocytes, whereas cell death transpired through a non-apoptotic, lipid peroxidation-dependent mechanism. Together, these data further our understanding of the mechanism of sterile inflammation

Hazard assessment of hexagonal boron nitride and hexagonal boron nitride reinforced thermoplastic polyurethane composites using human skin and lung cells

Hexagonal boron nitride (hBN) is an emerging two-dimensional material attracting considerable attention in the industrial sector given its innovative physicochemical properties. Potential risks are associated mainly with occupational exposure where inhalation and skin contact are the most relevant exposure routes for workers. Here we aimed at characterizing the effects induced by composites of thermoplastic polyurethane (TPU) and hBN, using immortalized HaCaT skin keratinocytes and BEAS-2B bronchial epithelial cells. The composite was abraded using a Taber® rotary abraser and abraded TPU and TPU-hBN were also subjected to photo-Fenton-mediated degradation mimicking potential weathering across the product life cycle. Cells were exposed to the materials for 24 h (acute exposure) or twice per week for 4 weeks (chronic exposure) and evaluated with respect to material internalization, cytotoxicity, and proinflammatory cytokine secretion. Additionally, comprehensive mass spectrometry-based proteomics and metabolomics (secretomics) analyses were performed. Overall, despite evidence of cellular uptake of the material, no significant cellular and/or protein expression profiles alterations were observed after acute or chronic exposure of HaCaT or BEAS-2B cells, identifying only few pro-inflammatory proteins. Similar results were obtained for the degraded materials. These results support the determination of hazard profiles associated with cutaneous and pulmonary hBN-reinforced polymer composites exposure.ISSN:0304-3894ISSN:1873-333

Early goal-directed therapy in severe sepsis and septic shock: insights and comparisons to ProCESS, ProMISe, and ARISE

Prior to 2001 there was no standard for early management of severe sepsis and septic shock in the emergency department. In the presence of standard or usual care, the prevailing mortality was over 40-50 %. In response, a systems-based approach, similar to that in acute myocardial infarction, stroke and trauma, called early goal-directed therapy was compared to standard care and this clinical trial resulted in a significant mortality reduction. Since the publication of that trial, similar outcome benefits have been reported in over 70 observational and randomized controlled studies comprising over 70,000 patients. As a result, early goal-directed therapy was largely incorporated into the first 6 hours of sepsis management (resuscitation bundle) adopted by the Surviving Sepsis Campaign and disseminated internationally as the standard of care for early sepsis management. Recently a trio of trials (ProCESS, ARISE, and ProMISe), while reporting an all-time low sepsis mortality, question the continued need for all of the elements of early goal-directed therapy or the need for protocolized care for patients with severe and septic shock. A review of the early hemodynamic pathogenesis, historical development, and definition of early goal-directed therapy, comparing trial conduction methodology and the changing landscape of sepsis mortality, are essential for an appropriate interpretation of these trials and their conclusions. Electronic supplementary material The online version of this article (doi:10.1186/s13054-016-1288-3) contains supplementary material, which is available to authorized users

Environmental and Health Impacts of Graphene and other Two-Dimensional Materials: A Graphene Flagship Perspective

Two-dimensional (2D) materials have attracted tremendous interest ever since the isolation of atomically thin sheets of graphene in 2004 due to the unique and versatile properties of these materials. However, the increasing production and use of 2D materials necessitate a thorough evaluation of the potential impact on human health and the environment. Furthermore, harmonized test protocols are needed with which to assess the safety of 2D materials. The Graphene Flagship project (2013-2023), funded by the European Commission, addressed the identification of the possible hazard of graphene-based materials as well as emerging 2D materials including transition metal dichalcogenides, hexagonal boron nitride, and others. Additionally, so-called green chemistry approaches were explored to achieve the goal of a safe and sustainable production and use of this fascinating family of nanomaterials. The present review provides a compact survey of the findings and the lessons learned in the Graphene Flagship