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Chemical modifications of proteins induced by ambient ozone (O3) and nitrogen oxides (NOx) are of public health concerns due to their potential to trigger respiratory diseases. The laboratory and environmental exposure systems have been widely used to investigate their relevant mechanism in the atmosphere. Using bovine serum albumin (BSA) as a model protein, we evaluated the two systems and aimed to reduce the uncertainties of both the reactants and products in the corresponding kinetic study. In the laboratory simulation system, the generated gaseous pollutants showed negligible losses. Ten layers of BSA were coated on the flow tube with protein extraction recovery of 87.4%. For environmental exposure experiment, quartz fiber filter was selected as the upper filter with low gaseous O3 (8.0%) and NO2 (1.7%) losses, and cellulose acetate filter was appropriate for the lower filter with protein extraction efficiency of 95.2%. The protein degradation process was observed without the exposure to atmospheric oxidants and contributed to the loss of protein monomer mass fractions, while environmental factors (e.g., molecular oxygen and ultraviolet) may cause greater protein monomer losses. Based on the evaluation, the study exemplarily applied the two systems to protein modification and both showed that O3 promotes the protein oligomerization and nitration, while increased temperature can accelerate the oligomerization and increased relative humidity can inhibit the nitration in the environmental exposure samples. The developed laboratory and environmental systems are suitable for studying protein modifications formed under different atmospheric conditions. A combination of the two will further reveal the actual mechanism of protein modifications.
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Poluentes Atmosféricos , Ozônio , Ozônio/química , Poluentes Atmosféricos/análise , Soroalbumina Bovina/química , Exposição Ambiental , Óxidos de Nitrogênio/análise , Proteínas/químicaRESUMO
Proteins in atmospheric aerosol can react with atmospheric pollutants such as ozone (O3) and nitrogen dioxide (NO2) in the atmosphere via the reactions of oxidation, nitration, and cross-linking etc. Currently, the reactions have been more thoroughly studied in the laboratory but rarely investigated in the ambient environment. In this study, we used bovine serum albumin (BSA) as the model protein to conduct the exposure experiment in the ambient environment in southern China, an area with increasing oxidative capacity, to investigate the reactions of proteins in the atmosphere. We observed the occurrence of oligomerization, nitration and degradation of BSA upon exposure. The mass fraction of BSA monomer decreased by 5.86 ± 1.61% after exposure and those of dimers, trimers and higher oligomers increased by 1.04 ± 0.49%, 1.37 ± 0.74% and 3.40 ± 1.06%, respectively. Simultaneously, the nitration degrees of monomers, dimers, trimers and higher oligomers increased by 0.42 ± 0.15%, 0.53 ± 0.15%, 0.55 ± 0.28% and 2.15 ± 1.01%, respectively. The results show that oligomerization was significantly affected by O3 and temperature and nitration was jointly affected by O3, temperature and relative humidity, indicating the important role of atmospheric oxidants in the atmospheric reactions of protein. Atmospheric degradation of BSA was observed with the release of free amino acids (FAAs) such as glycine, alanine, serine and methionine. Glycine was the dominant FAA with a molar yield ranging from â¼8% to 33% for BSA. The estimated stoichiometric coefficient (α) of glycine is 10-7-10-6 for the degradation of BSA upon O3. Our observation suggests the occurrence of protein reactions in the oxidative ambient environment, leading to the production of nitrated products, oligomers and low molecular weight products such as peptides and FAAs. This study may deepen the current understanding of the atmospheric reaction mechanisms and reveal the influence of environmental factors in the atmosphere.
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Poluentes Atmosféricos , Ozônio , Soroalbumina Bovina/química , Peptídeos , Aminoácidos , Poluentes Atmosféricos/química , Glicina , Ozônio/químicaRESUMO
Exposure to ambient fine particulate matter (PM2.5) is associated with millions of premature deaths annually. Oxidative stress through overproduction of reactive oxygen species (ROS) is a possible mechanism for PM2.5-induced health effects. Organic aerosol (OA) is a dominant component of PM2.5 worldwide, yet its role in PM2.5 toxicity is poorly understood due to its chemical complexity. Here, through integrated cellular ROS measurements and detailed multi-instrument chemical characterization of PM in urban southeastern United States, we show that oxygenated OA (OOA), especially more-oxidized OOA, is the main OA type associated with cellular ROS production. We further reveal that highly unsaturated species containing carbon-oxygen double bonds and aromatic rings in OOA are major contributors to cellular ROS production. These results highlight the key chemical features of ambient OA driving its toxicity. As more-oxidized OOA is ubiquitous and abundant in the atmosphere, this emphasizes the need to understand its sources and chemical processing when formulating effective strategies to mitigate PM2.5 health impacts.
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Estresse Oxidativo , Oxigênio , Espécies Reativas de Oxigênio , Aerossóis , Sudeste dos Estados UnidosRESUMO
Amorphous zero-valent iron (AZVI) has attracted wide attention due to its high-efficiency reduction ability. However, the effect of different EDA/Fe(II) molar ratios on the physicochemical properties of the synthesized AZVI requires further investigation. Herein, series of AZVI samples were prepared by changing the molar ratio of EDA/Fe(II) to 1/1 (AZVI@1), 2/1 (AZVI@2), 3/1 (AZVI@3), and 4/1 (AZVI@4). When the EDA/Fe(II) ratio increased from 0/1 to 3/1, the Fe0 proportion on the AZVI surface increased from 26.0 to 35.2% and the reducing ability was enhanced. As for AZVI@4, the surface was severely oxidized to form a large amount of Fe3O4, and the Fe0 content was only 74.0%. Moreover, the removal ability of Cr(VI) was in the order AZVI@3 > AZVI@2 > AZVI@1 > AZVI@4. The isothermal titration calorimetry results revealed that the increase of the molar ratio of EDA/Fe(II) would lead to the stronger complexation of EDA with Fe(II), which resulted in the gradual decrease of the yield of AZVI@1 to AZVI@4 and the gradual deterioration of water pollution after the synthesis. Therefore, based on the evaluation of all indicators, AZVI@2 was the optimal material, not only because its yield was as high as 88.7% and the secondary water pollution level was low, but most importantly, the removal efficiency of Cr(VI) by AZVI@2 was excellent. Furthermore, the actual Cr(VI) wastewater with the concentration of 14.80 mg/L was treated with AZVI@2, and the removal rate of 97.0% was achieved after only a 30 min reaction. This work clarified the effect of different ratios of EDA/Fe(II) on the physicochemical properties of AZVI, which provided insights for guiding the reasonable synthesis of AZVI and is also conducive to investigating the reaction mechanism of AZVI in Cr(VI) remediation.
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Microplastics (MPs) pollution is becoming one of the most pressing environmental issues globally. MPs in the marine, freshwater and terrestrial environments have been fairly well investigated. However, knowledge of the atmospheric-mediated deposition of MPs within rural environments is limited. Here, we present the results of bulk (dry and wet) atmospheric MPs deposition in a rural region of Quzhou County in the North China Plain (NCP). Samples of MPs in the atmospheric bulk deposition were collected for individual rainfall events over a 12-month period from August 2020 to August 2021. The number and size of MPs from 35 rainfall samples were measured by fluorescence microscopy, while the chemical composition of MPs was identified using micro-Fourier transform infrared spectroscopy (µ-FTIR). The results showed that the atmospheric MPs deposition rate in summer (892-75,421 particles/m2/day) was highest compared to 735-9428, 280-4244 and 86-1347 particles/m2/day in spring, autumn, and winter, respectively. Furthermore, the MPs deposition rates in our study were 1-2 orders of magnitude higher than those in other regions, indicating a higher rate of MPs deposition in the rural region of the NCP. MPs with a diameter of 3-50 µm accounted for 75.6 %, 78.4 %, 73.4 % and 66.1 % of total MPs deposition in spring, summer, autumn, and winter, respectively, showing that the majority of MPs in the current study were small in size. Rayon fibers accounted for the largest proportion (32 %) of all MPs, followed by polyethylene terephthalate (12 %) and polyethylene (8 %). This study also found that a significant positive correlation between rainfall volume and MPs deposition rate. In addition, HYSPLIT back-trajectory modelling showed that the farthest source of deposition MPs may have come from Russia.
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The family of atmospheric oxides of nitrogen, NOy (e.g., nitrogen oxides (NOx) + nitric acid (HNO3) + nitrous acid (HONO) + peroxyacetyl nitrate (PAN) + particulate nitrate (pNO3-) + other), have an influential role in atmospheric chemistry, climate, and the environment. The nitrogen (δ15N) and oxygen (δ18O and Δ17O) stable isotopes of NOy are novel tools for potentially tracking emission sources and quantifying oxidation chemistry. However, there is a lack of well-established methods, particularly for speciated gas-phase components of NOy, to accurately quantify δ15N, δ18O, and Δ17O. This work presents controlled laboratory experiments and complex chamber α-pinene/NOx oxidation experiments of a sampling apparatus constructed for the simultaneous capture of multiple NOy species for isotope analysis using a series of coated denuders, with a focus on nitrogen dioxide (NO2â¢). The laboratory tests indicate complete NO2⢠capture for the targeted concentration of 15 ppbv for at least 24 h collections at 10 liters per minute, with δ15N and δ18O precisions of ±1.3 and 1.0, respectively, and minimal (2.2% ± 0.1%) NO2⢠collection on upstream denuders utilized for the capture of HNO3 and other acidic gases. The multispecies NOy collection system showed excellent concentration correlations with online instrumentation for both HNO3 and NO2⢠and isotope reproducibility of ±1.7, ±1.8, and ±0.7 for δ15N, δ18O, and Δ17O, respectively, for replicate experiments and highly time-resolved collections. This work demonstrates a new method that can enable the simultaneous collection of HNO3 and NO2⢠for accurate quantification of concentration and isotopic composition.
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Oxidative stress is a possible mechanism by which ambient fine particulate matter (PM) exerts adverse biological effects. While multiple biological effects and reactive oxygen species (ROS) production have been observed upon PM exposure, whether the biological effects are ROS-mediated remains unclear. Secondary organic aerosols (SOA) constitute a major fraction of fine PM and can contribute substantially to its toxicity. In this work, we measured three types of cell responses (mitochondrial membrane potential (MMP), caspase 3/7 activity, and ROS) and investigated their associations upon exposure to SOA formed from anthropogenic (naphthalene) and biogenic (α-pinene) precursors. MMP and caspase 3/7 activity (an early indicator of apoptosis) are key indicators of cell health, and changes of them could occur downstream of ROS-mediated pathways. We observed a significant increase in caspase 3/7 activity after SOA exposure, suggesting that apoptosis is an important pathway of cell death induced by SOA. We further found strong associations between a decrease in MMP and increase in caspase 3/7 activity with an increase in cellular ROS level. These results suggest that cell health is largely dependent on the cellular ROS level, highlighting oxidative stress as a key mechanism for biological effects from SOA exposure. Linear regression analyses reveal greater changes of the three cellular responses with increasing carbon oxidation state (OSc) of SOA, suggesting that SOA are more toxic when they are more oxidized. Overall, our work provides critical insights into the associations between cell health and ROS level upon SOA exposure and proposes that OSc could be a suitable proxy to assess the overall SOA toxicity.
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Poluentes Atmosféricos , Espécies Reativas de Oxigênio/análise , Poluentes Atmosféricos/toxicidade , Poluentes Atmosféricos/análise , Caspase 3/análise , Material Particulado/análise , Aerossóis/análiseRESUMO
Cloth masks are a tool for controlling community transmission during pandemics, as well as during other outbreak situations. However, cloth masks vary in their designs, and the consequences of this variability for their effectiveness as source control have received little attention, particularly in terms of user discomfort and problematic mask-wearing behaviors. In the present studies, common design parameters of cloth masks were systematically varied to ascertain their effect(s) on the subjective discomfort and frequency of problematic mask-wearing behaviors, which detract from the effectiveness of cloth masks as source control. The type of fabric comprising a mask (flannel or twill made of 100% cotton) and the attachment-style of a mask (i.e., ear loops or fabric ties) were varied in adults (18 to 65 years) and children (ages 6 to 11 years). For adults, ear loops were less comfortable than ties (p = .035) and were associated with greater face- (p = .005) and mask-touching (p = .001). Children, however, found flannel masks to be more breathable than twill masks (p = .007) but touched their masks more frequently when wearing a mask made of flannel than twill (p = .033). Common design parameters of cloth masks not only affect user discomfort and behavior but do so differently in adults and children. To improve the effectiveness of cloth masks as source control, the present studies highlight the importance of measuring the effect(s) of design decisions on user discomfort and behavior in different populations.
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COVID-19 , Adulto , Criança , Humanos , COVID-19/prevenção & controle , SARS-CoV-2 , Têxteis , Pandemias/prevenção & controle , TatoRESUMO
Health risks associated with exposure to ambient particulate matter (PM) are a major concern around the world. Adverse PM health effects have been proposed to be linked to oxidative stress through the generation of reactive oxygen species (ROS). In vitro cellular assays can provide insights into components or characteristics of PM that best account for its toxicity at a cellular level. However, most current assays report cell population averages and are mostly time endpoint measurements and thus provide no temporal information. This poses limitations on our understanding of PM health effects. In this study, we developed a microfluidic assay that can measure cellular ROS responses at the single-cell level and evaluate temporal dynamic behavior of single cells. We first established a protocol that enables culturing cells in our microfluidic platform and that can provide reproducible ROS readouts. We further examined the heterogeneous ROS responses of cell populations and tracked the dynamics of individual cellular responses upon exposure to different concentrations of PM extracts. Our results show that in an alveolar macrophage cell line, cellular ROS responses are highly heterogeneous. ROS responses from different cells can vary over an order of magnitude, and large coefficients of variation at each timepoint measurement indicate a high variability. The dynamic behavior of single-cell responses is strongly dependent on PM concentrations. Our work serves as a proof-of-principle demonstration of the capability of our microfluidic technology to study time-resolved single-cell responses upon PM exposure. We envision applying this high-resolution, high-content assay to investigate a wide array of single-cell responses (beyond ROS) upon exposure to different types of PM in the future.
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Poluentes Atmosféricos , Material Particulado , Poluentes Atmosféricos/análise , Macrófagos Alveolares , Estresse Oxidativo , Material Particulado/análise , Material Particulado/toxicidade , Espécies Reativas de Oxigênio/farmacologiaRESUMO
Highly oxygenated molecules (HOMs) play an important role in the formation and evolution of secondary organic aerosols (SOA). However, the abundance of HOMs in different environments and their relation to the oxidative potential of fine particulate matter (PM) are largely unknown. Here, we investigated the relative HOM abundance and radical yield of laboratory-generated SOA and fine PM in ambient air ranging from remote forest areas to highly polluted megacities. By electron paramagnetic resonance and mass spectrometric investigations, we found that the relative abundance of HOMs, especially the dimeric and low-volatility types, in ambient fine PM was positively correlated with the formation of radicals in aqueous PM extracts. SOA from photooxidation of isoprene, ozonolysis of α- and ß-pinene, and fine PM from tropical (central Amazon) and boreal (Hyytiälä, Finland) forests exhibited a higher HOM abundance and radical yield than SOA from photooxidation of naphthalene and fine PM from urban sites (Beijing, Guangzhou, Mainz, Shanghai, and Xi'an), confirming that HOMs are important constituents of biogenic SOA to generate radicals. Our study provides new insights into the chemical relationship of HOM abundance, composition, and sources with the yield of radicals by laboratory and ambient aerosols, enabling better quantification of the component-specific contribution of source- or site-specific fine PM to its climate and health effects.
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Poluentes Atmosféricos , Material Particulado , Aerossóis , Pequim , China , FinlândiaRESUMO
Toll-like receptor 4 (TLR4) plays a crucial role in the recognition of invading pathogens. Upon activation by lipopolysaccharides (LPS), TLR4 is recruited into specific membrane domains and dimerizes. In addition to LPS, TLR4 can be stimulated by wheat amylase-trypsin inhibitors (ATI). ATI are proteins associated with gluten containing grains, whose ingestion promotes intestinal and extraintestinal inflammation. However, the effect of ATI vs. LPS on the membrane distribution of TLR4 at the nanoscale has not been analyzed. In this study, we investigated the effect of LPS and ATI stimulation on the membrane distribution of TLR4 in primary human macrophages using single molecule localization microscopy (SMLM). We found that in unstimulated macrophages the majority of TLR4 molecules are located in clusters, but with donor-dependent variations from â¼51% to â¼75%. Depending on pre-clustering, we found pronounced variations in the fraction of clustered molecules and density of clusters on the membrane upon LPS and ATI stimulation. Although clustering differed greatly among the human donors, we found an almost constant cluster diameter of â¼44 nm for all donors, independent of treatment. Together, our results show donor-dependent but comparable effects between ATI and LPS stimulation on the membrane distribution of TLR4. This may indicate a general mechanism of TLR4 activation in primary human macrophages. Furthermore, our methodology visualizes TLR4 receptor clustering and underlines its functional role as a signaling platform.
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Lipopolissacarídeos/farmacologia , Receptor 4 Toll-Like/metabolismo , Inibidores da Tripsina/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células Cultivadas , Humanos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Microscopia de FluorescênciaRESUMO
Cerium oxide nanoparticles (CeNPs) have been shown to exhibit antioxidant capabilities, but their efficiency in scavenging reactive oxygen species (ROS) and the underlying mechanisms are not yet well understood. In this study, cerium dioxide nanoparticles (CeNPs) and nanorods (CeNRs) were found to exhibit much stronger scavenging activity than ·OH generation in phosphate buffered saline (PBS) and surrogate lung fluid (SLF). The larger surface area and higher defect density of CeNRs may lead to higher ·OH scavenging activity than for CeNPs. These insights are important to understand the redox activity of cerium nanomaterials and provide clues to the role of CeNPs in biological and environmental processes.
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Amylase trypsin inhibitors (ATI) can be found in all gluten containing cereals and are, therefore, ingredient of basic foods like bread or pasta. In the gut ATI can mediate innate immunity via activation of the Toll-like receptor 4 (TLR4) on immune cells residing in the lamina propria, promoting intestinal, as well as extra-intestinal, inflammation. Inflammatory conditions can induce formation of peroxynitrite (ONOO-) and, thereby, endogenous protein nitration in the body. Moreover, air pollutants like ozone (O3) and nitrogen dioxide (NO2) can cause exogenous protein nitration in the environment. Both reaction pathways may lead to the nitration of ATI. To investigate if and how nitration modulates the immunostimulatory properties of ATI, they were chemically modified by three different methods simulating endogenous and exogenous protein nitration and tested in vitro. Here we show that ATI nitration was achieved by all three methods and lead to increased immune reactions. We found that ATI nitrated by tetranitromethane (TNM) or ONOO- lead to a significantly enhanced TLR4 activation. Furthermore, in human primary immune cells, TNM nitrated ATI induced a significantly higher T cell proliferation and release of Th1 and Th2 cytokines compared to unmodified ATI. Our findings implicate a causative chain between nitration, enhanced TLR4 stimulation, and adaptive immune responses, providing major implications for public health, as nitrated ATI may strongly promote inhalative wheat allergies (baker's asthma), non-celiac wheat sensitivity (NCWS), other allergies, and autoimmune diseases. This underlines the importance of future work analyzing the relationship between endo- and exogenous protein nitration, and the rise in incidence of ATI-related and other food hypersensitivities.
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Imunidade Adaptativa/efeitos dos fármacos , Amilases/farmacologia , Imunidade Inata/efeitos dos fármacos , Triticum/metabolismo , Inibidores da Tripsina/farmacologia , Amilases/química , Biomarcadores , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Imunofenotipagem , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Proteínas de Plantas/metabolismo , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Receptor 4 Toll-Like/metabolismo , Inibidores da Tripsina/químicaRESUMO
The allergenic potential of airborne proteins may be enhanced via post-translational modification induced by air pollutants like ozone (O3) and nitrogen dioxide (NO2). The molecular mechanisms and kinetics of the chemical modifications that enhance the allergenicity of proteins, however, are still not fully understood. Here, protein tyrosine nitration and oligomerization upon simultaneous exposure of O3 and NO2 were studied in coated-wall flow-tube and bulk solution experiments under varying atmospherically relevant conditions (5-200 ppb O3, 5-200 ppb NO2, 45-96% RH), using bovine serum albumin as a model protein. Generally, more tyrosine residues were found to react via the nitration pathway than via the oligomerization pathway. Depending on reaction conditions, oligomer mass fractions and nitration degrees were in the ranges of 2.5-25% and 0.5-7%, respectively. The experimental results were well reproduced by the kinetic multilayer model of aerosol surface and bulk chemistry (KM-SUB). The extent of nitration and oligomerization strongly depends on relative humidity (RH) due to moisture-induced phase transition of proteins, highlighting the importance of cloud processing conditions for accelerated protein chemistry. Dimeric and nitrated species were major products in the liquid phase, while protein oligomerization was observed to a greater extent for the solid and semi-solid phase states of proteins. Our results show that the rate of both processes was sensitive towards ambient ozone concentration, but rather insensitive towards different NO2 levels. An increase of tropospheric ozone concentrations in the Anthropocene may thus promote pro-allergic protein modifications and contribute to the observed increase of allergies over the past decades.
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Poluentes Atmosféricos/química , Atmosfera/química , Dióxido de Nitrogênio/química , Ozônio/química , Proteínas/química , Poluentes Atmosféricos/metabolismo , Dióxido de Nitrogênio/metabolismo , Ozônio/metabolismo , Proteínas/metabolismoRESUMO
Water-soluble proteinaceous matter including proteins and free amino acids (FAAs) as well as some other chemical components was analyzed in fine particulate matter (PM2.5) samples collected over a period of one year in rural Guangzhou. Annual averaged protein and total FAAs concentrations were 0.79 ± 0.47 µg m-3 and 0.13 ± 0.05 µg m-3, accounting for 1.9 ± 0.7% and 0.3 ± 0.1% of PM2.5, respectively. Among FAAs, glycine was the most abundant species (19.9%), followed by valine (18.5%), methionine (16.1%), and phenylalanine (13.5%). Both proteins and FAAs exhibited distinct seasonal variations with higher concentrations in autumn and winter than those in spring and summer. Correlation analysis suggests that aerosol proteinaceous matter was mainly derived from intensive agricultural activities, biomass burning, and fugitive dust/soil resuspension. Significant correlations between proteins/FAAs and atmospheric oxidant (O3) indicate that proteins/FAAs may be involved in O3 related atmospheric processes. Our observation confirms that ambient FAAs could be degraded from proteins under the influence of O3, and the stoichiometric coefficients of the reactions were estimated for FAAs and glycine. This finding provides a possible pathway for the production of aerosol FAAs in the atmosphere, which will improve the current understanding on atmospheric processes of proteinaceous matter.
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Aminoácidos/análise , Monitoramento Ambiental , Material Particulado , Proteínas/análise , Poluentes Atmosféricos , ChinaRESUMO
Air pollution and climate change are potential drivers for the increasing burden of allergic diseases. The molecular mechanisms by which air pollutants and climate parameters may influence allergic diseases, however, are complex and elusive. This article provides an overview of physical, chemical and biological interactions between air pollution, climate change, allergens, adjuvants and the immune system, addressing how these interactions may promote the development of allergies. We reviewed and synthesized key findings from atmospheric, climate, and biomedical research. The current state of knowledge, open questions, and future research perspectives are outlined and discussed. The Anthropocene, as the present era of globally pervasive anthropogenic influence on planet Earth and, thus, on the human environment, is characterized by a strong increase of carbon dioxide, ozone, nitrogen oxides, and combustion- or traffic-related particulate matter in the atmosphere. These environmental factors can enhance the abundance and induce chemical modifications of allergens, increase oxidative stress in the human body, and skew the immune system toward allergic reactions. In particular, air pollutants can act as adjuvants and alter the immunogenicity of allergenic proteins, while climate change affects the atmospheric abundance and human exposure to bioaerosols and aeroallergens. To fully understand and effectively mitigate the adverse effects of air pollution and climate change on allergic diseases, several challenges remain to be resolved. Among these are the identification and quantification of immunochemical reaction pathways involving allergens and adjuvants under relevant environmental and physiological conditions.
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Alérgenos/imunologia , Mudança Climática , Poluentes Atmosféricos , Poluição do Ar , Humanos , HipersensibilidadeRESUMO
Chemical modifications such as nitration and cross-linking may enhance the allergenic potential of proteins. The kinetics and mechanisms of the underlying chemical processes, however, are not yet well understood. Here, we present a size-exclusion chromatography/spectrophotometry method (SEC-HPLC-DAD) that allows a simultaneous detection of mono-, di-, tri-, and higher protein oligomers, as well as their individual nitration degrees (NDs). The ND results of proteins from this new method agree well with the results from an alternative well-established method, for the analysis of tetranitromethane (TNM)- and nitrogen dioxide and ozone (NO2/O3)-nitrated protein samples. Importantly, the NDs for individual oligomer fractions can be obtained from the new method, and also, we provide a proof of principle for the calculation of the concentrations for individual protein oligomer fractions by their determined NDs, which will facilitate the investigation of the kinetics and mechanism for protein tyrosine nitration and cross-linking.
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Nitratos/química , Proteínas/química , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Dióxido de Nitrogênio/química , Ozônio/química , Proteínas/análise , Espectrofotometria Ultravioleta , Tetranitrometano/químicaRESUMO
Hydroxyl radical-induced oxidation of proteins and peptides can lead to the cleavage of the peptide, leading to a release of fragments. Here, we used high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) and pre-column online ortho-phthalaldehyde (OPA) derivatization-based amino acid analysis by HPLC with diode array detection and fluorescence detection to identify and quantify free amino acids released upon oxidation of proteins and peptides by hydroxyl radicals. Bovine serum albumin (BSA), ovalbumin (OVA) as model proteins, and synthetic tripeptides (comprised of varying compositions of the amino acids Gly, Ala, Ser, and Met) were used for reactions with hydroxyl radicals, which were generated by the Fenton reaction of iron ions and hydrogen peroxide. The molar yields of free glycine, aspartic acid, asparagine, and alanine per peptide or protein varied between 4 and 55%. For protein oxidation reactions, the molar yields of Gly (â¼32-55% for BSA, â¼10-21% for OVA) were substantially higher than those for the other identified amino acids (â¼5-12% for BSA, â¼4-6% for OVA). Upon oxidation of tripeptides with Gly in C-terminal, mid-chain, or N-terminal positions, Gly was preferentially released when it was located at the C-terminal site. Overall, we observe evidence for a site-selective formation of free amino acids in the OH radical-induced oxidation of peptides and proteins, which may be due to a reaction pathway involving nitrogen-centered radicals.
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Aminoácidos/química , Radical Hidroxila/química , Peptídeos/química , Proteínas/química , Oxirredução , Espécies Reativas de Oxigênio/químicaRESUMO
Metaproteomic analysis of air particulate matter provides information about the abundance and properties of bioaerosols in the atmosphere and their influence on climate and public health. We developed and applied efficient methods for the extraction and analysis of proteins from glass fiber filter samples of total, coarse, and fine particulate matter. Size exclusion chromatography was applied to remove matrix components, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied for protein fractionation according to molecular size, followed by in-gel digestion and LC-MS/MS analysis of peptides using a hybrid Quadrupole-Orbitrap MS. Maxquant software and the Swiss-Prot database were used for protein identification. In samples collected at a suburban location in central Europe, we found proteins that originated mainly from plants, fungi, and bacteria, which constitute a major fraction of primary biological aerosol particles (PBAP) in the atmosphere. Allergenic proteins were found in coarse and fine particle samples, and indications for atmospheric degradation of proteins were observed. Graphical abstract Workflow for the metaproteomic analysis of atmospheric aerosol samples.