Particle-Induced Chronic Inflammation is Dependent upon Lysosomal Function and Autophagy

Chronic inflammation drives the development of many debilitating pulmonary diseases. NLRP3 Inflammasome activation following lysosomal membrane permeablization (LMP) has been identified as a necessary event in the maturation of IL-1β, a critical cytokine in the development of chronic inflammation. LMP and NLRP3 activation are known to occur rapidly following particle uptake by alveolar macrophages, however the longevity of these responses has not been defined. These studies are the first to describe the persistence of the NLRP3 Inflammasome response in relation to autophagy, the primary degradation pathway for NLRP3 components, and lysosomal integrity. Alveolar macrophages were isolated from C57Bl/6 mice 7 days following silica or vehicle exposure and assessed for impaired autophagy. LC3-II and p62 were elevated in cell lysates from alveolar macrophages isolated from silica-exposed mice, as well as Inflammasome protein components NLRP3 and ASC. IL-1β levels were below detection limits in the whole lung lavage fluid at 7 days and alveolar macrophages isolated from silica-treated mice after 7 days secreted negligible amounts of IL-1β after 24 hours of ex vivo culture, suggesting suppressed Inflammasome activity. Addition of low levels of endotoxin to isolated alveolar macrophages was sufficient to cause Inflammasome reactivation, resulting in renewed IL-1β production. Inflammasome reactivation could be suppressed with the Cathepsin B inhibitor Ca-074-Me, linking Inflammasome reactivation by endotoxin with impaired lysosome function. Multi-walled Carbon Nanotubes induced responses relative to silica, indicating that these mechanisms drive most particle-induced chronic inflammation. These studies demonstrate that lysosomal function and autophagy are integral to the development of chronic inflammation, and targeting of these two integrated pathways should be considered in therapeutic approaches to preventing chronic inflammatory disease

MECHANISMS AND CONSIQENCES OF LYSOSOMAL MEMBRANE PERMEABILIZATION FOLLOWING EXPOSURE TO BIOACTIVE PARTICLES

Exposure to bioactive environmental particles and engineered nanoparticles are a significant public health concern. Inhalation of bioactive particles can result in chronic inflammation, which drives tissue remodeling and fibrosis. Furthermore, chronic inflammation can increase individual susceptibility to other diseases including cancer and autoimmune diseases. The macrophage is the critical cell in particle clearance following exposure, and is central to the inflammatory responses and tissue remodeling. Phagocytosed bioactive particles within macrophages cause cytotoxicity and activation of the NLRP3 inflammasome, outcomes that are both essential to inflammation and disease development. However, mechanisms that regulate NLRP3 inflammasome activity and cytotoxicity have not fully been elucidated. The objective of this body of work was to further define common yet critical mechanisms that cause and/or mediate NLRP3 inflammasome activity following exposure to bioactive particles. In these studies we demonstrate that bioactive particles including silica and engineered nanomaterials cause lysosome membrane permeabilization (LMP) and the release of lysosomal proteases, which precedes and facilitates NLRP3 inflammasome activation. Bioactive particles cause LMP through a mechanism that requires phagolysosome acidification. LMP and the activation of the NLRP3 inflammasome are required for secretion of pro-inflammatory cytokines and the alarmin High Mobility Group Box 1 (HMGB1). Once secreted, HMGB1 can further drive NLRP3 inflammasome activity through sterile priming, similar to the nonsterile mechanism utilized by endotoxin. A second critical pathway for regulation of the NLRP3 inflammasome was autophagy. Mice with macrophages deficient in autophagy had greater inflammation and chronic disease following silica exposure, supporting a protective anti-inflammatory role for autophagic activity. Together, these data reveal novel critical mechanisms in the regulation of NLRP3 inflammasome activity following bioactive particle exposure, and provide multiple potential therapeutic targets for the suppression of inflammation and disease

Urinary Levoglucosan as a Biomarker of Wood Smoke Exposure: Observations in a Mouse Model and in Children

BACKGROUND: Biomass smoke is an important source of particulate matter (PM), and much remains to be discovered with respect to the human health effects associated with this specific PM source. Exposure to biomass smoke can occur in one of two main categories: short-term exposures consist of periodic, seasonal exposures typified by communities near forest fires or intentional agricultural burning, and long-term exposures are chronic and typified by the use of biomass materials for cooking or heating. Levoglucosan (LG), a sugar anhydride released by combustion of cellulose-containing materials, is an attractive candidate as a biomarker of wood smoke exposure. OBJECTIVES: In the present study, Balb/c mice and children were assessed for LG in urine to determine its feasibility as a biomarker. METHODS: We performed urinary detection of LG by gas chromatography/mass spectrometry after intranasal instillations of LG or concentrated PM (mice) or biomass exposure (mice or humans). RESULTS: After instillation, we recovered most of the LG within the first 4 hr. Experiments using glucose instillation proved the specificity of our system, and instillation of concentrated PM from wood smoke, ambient air, and diesel exhaust supported a connection between wood smoke and LG. In addition, LG was detected in the urine of mice exposed to wood smoke. Finally, a pilot human study proved our ability to detect LG in urine of children. CONCLUSIONS: These results demonstrate that LG in the lungs is detectable in the urine of both mice and humans and that it is a good candidate as a biomarker of exposure to biomass smoke

Measurement of the forward-backward asymmetry in the B→K(*) μ\u3csup\u3e+\u3c/sup\u3eμ\u3csup\u3e-\u3c/sup\u3e decay and first observation of the Bs0→μ\u3csup\u3e+\u3c/sup\u3eμ\u3csup\u3e-\u3c/sup\u3e decay

We reconstruct the rare decays B+→K+μ +μ-, B0→K*(892)0μ +μ-, and Bs0→(1020)μ+μ - in a data sample corresponding to 4.4fb-1 collected in pp̄ collisions at √s=1.96TeV by the CDF II detector at the Tevatron Collider. Using 121±16 B+→K+μ +μ- and 101±12 B0→K*0μ +μ- decays we report the branching ratios. In addition, we report the differential branching ratio and the muon forward-backward asymmetry in the B+ and B0 decay modes, and the K*0 longitudinal polarization fraction in the B0 decay mode with respect to the squared dimuon mass. These are consistent with the predictions, and most recent determinations from other experiments and of comparable accuracy. We also report the first observation of the Bs0→μ+μ- decay and measure its branching ratio BR(Bs0→μ+μ-)= [1.44±0.33±0.46]×10-6 using 27±6 signal events. This is currently the most rare Bs0 decay observed. © 2011 American Physical Society

Search for a new heavy gauge boson W′ with event signature electron+missing transverse energy in pp̅ collisions at √s=1.96 TeV

We present a search for a new heavy charged vector boson W′ decaying to an electron-neutrino pair in pp̅ collisions at a center-of-mass energy of 1.96 TeV. The data were collected with the CDF II detector and correspond to an integrated luminosity of 5.3  fb-1. No significant excess above the standard model expectation is observed and we set upper limits on σ·B(W′→eν). Assuming standard model couplings to fermions and the neutrino from the W′ boson decay to be light, we exclude a W′ boson with mass less than 1.12  TeV/c2 at the 95% confidence level.We thank the Fermilab staff and the technical staffs of the participating institutions for their vital contributions. This work was supported by the U.S. Department of Energy and National Science Foundation; the Italian Istituto Nazionale di Fisica Nucleare; the Ministry of Education, Culture, Sports, Science and Technology of Japan; the Natural Sciences and Engineering Research Council of Canada; the National Science Council of the Republic of China; the Swiss National Science Foundation; the A. P. Sloan Foundation; the Bundesministerium für Balduin Una Forschung, Germany; the World Class University Program, the National Research Foundation of Korea; the Science and Technology Facilities Council and the Royal Society, United Kingdom; the Institut National de Physique Nucleaire et Physique des Particules/CNRS and Universite Pierre et Marie Curie; the Russian Foundation for Basic Research; the Ministerio de Ciencia e Innovación, and Programa Consolider-Ingenio 2010, Spain; the Slovak R&D Agency; and the Academy of Finland

Search for New Dielectron Resonances and Randall-Sundrum Gravitons at the Collider Detector at Fermilab

A search for new dielectron-mass resonances using data recorded by the CDF II detector and corresponding to an integrated luminosity of 5.7fb-1 is presented. No significant excess over the expected standard model prediction is observed. In this data set, an event with the highest dielectron mass ever observed (960GeV/c2) was recorded. The results are interpreted in the Randall-Sundrum (RS) model. Combined with the 5.4fb-1 diphoton analysis, the RS-graviton lower-mass limit for the coupling k/M ̄Pl=0.1 is 1058GeV/c2, making it the strongest limit to date. © 2011 American Physical Society

Extracellular HMGB1 regulates multi-walled carbon nanotube-induced inflammation <i>in vivo</i>

<p>Endotoxin is often used to activate NF-κB <i>in vitro</i> when assessing NLRP3 inflammasome activation by various exogenous particles including nanoparticles. However, the endogenous source of this signal 1 is unknown. High-mobility group box 1 (HMGB1) is known to play a critical role in acute lung injury, however the potential contribution of the alarmin HMGB1 to NLRP3 Inflammasome activation has not been determined in response to nanoparticles <i>in vivo</i>. In this study, the ability of multi-walled carbon nanotubes (MWCNT) to cause release of HMGB1 <i>in vitro</i> and <i>in vivo</i>, as well as the potential of HMGB1 to function as signal 1 <i>in vitro</i> and <i>in vivo</i>, was determined. HMGB1 activity <i>in vivo</i> was assessed by administration of HMGB1 neutralization antibodies following MWCNT exposure. <i>Caspase-1</i>^{<i>−/−</i>} mice were utilized to elucidate the dependence of HMGB1 secretion on NLRP3 inflammasome activity. MWCNT exposure increased extracellular HMGB1 levels in primary alveolar macrophages from C57Bl/6 mice and C10 mouse epithelial cell culture supernatants, and in C57Bl/6 mouse lung lavage fluid. MWCNT-induced HMGB1 secretion was dependent upon caspase-1. HMGB1 increased MWCNT-induced IL-1β release from macrophages <i>in vitro</i>, and neutralization of extracellular HMGB1 reduced MWCNT-induced IL-1β secretion <i>in vivo</i>. HMGB1 neutralization was accompanied with overall decreased inflammation. In summary, this study suggests extracellular HMGB1 participates in NLRP3 inflammasome activity and regulates IL-1β associated sterile inflammation induced by MWCNT.</p

Route of Francisella tularensis infection informs spatiotemporal metabolic reprogramming and inflammation in mice.

Route of exposure to pathogens can inform divergent disease pathogenesis and mortality rates. However, the features that contribute to these differences are not well established. Host metabolism has emerged as a critical element governing susceptibility and the metabolism of tissue exposure sites are unique. Therefore, specific metabolic niches may contribute to the course and outcome of infection depending on route of infection. In the current study, we utilized a combination of imaging and systems metabolomics to map the spatiotemporal dynamics of the host response to intranasal (i.n.) or intradermal (i.d.) infection of mice using the bacterium Francisella tularensis subsp tularensis (FTT). FTT causes lethal disease through these infection routes with similar inoculation doses and replication kinetics, which allowed for isolation of host outcomes independent of bacterial burden. We observed metabolic modifications that were both route dependent and independent. Specifically, i.d. infection resulted in early metabolic reprogramming at the site of infection and draining lymph nodes, whereas the lungs and associated draining lymph nodes were refractory to metabolic reprogramming following i.n. infection. Irrespective of exposure route, FTT promoted metabolic changes in systemic organs prior to colonization, and caused massive dysregulation of host metabolism in these tissues prior to onset of morbidity. Preconditioning infection sites towards a more glycolytic and pro-inflammatory state prior to infection exacerbated FTT replication within the lungs but not intradermal tissue. This enhancement of replication in the lungs was associated with the ability of FTT to limit redox imbalance and alter the pentose phosphate pathway. Together, these studies identify central metabolic features of the lung and dermal compartments that contribute to disease progression and identify potential tissue specific targets that may be exploited for novel therapeutic approaches