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Soot particles (SP) are ubiquitous components of atmospheric particulate matter and have been shown to cause various adverse health effects. In the atmosphere, freshly emitted SP can be coated by condensed low-volatility secondary organic and inorganic species. In addition, gas-phase oxidants may react with the surface of SP. Due to the chemical and physical resemblance of SP carbon backbone with polyaromatic hydrocarbon species and their potent oxidation products, we investigated the biological responses of BEAS-2B lung epithelial cells following exposure to fresh- and photochemically aged-SP at the air–liquid interface. A comprehensive physical and chemical aerosol characterization was performed to depict the atmospheric transformations of SP, showing that photochemical aging increased the organic carbon fraction and the oxidation state of the SP. RNA-sequencing and qPCR analysis showed varying gene expression profiles for fresh- and aged-SP. Exposure to aged-SP increased DNA damage, oxidative damage, and upregulation of NRF2-mediated oxidative stress response genes compared to fresh-SP. Furthermore, aged-SP augmented inflammatory cytokine secretion and activated AhR-response, as evidenced by increased expression of AhR-responsive genes. These results indicate that oxidative stress, inflammation, and DNA damage play a key role in the cytotoxicity of SP in BEAS-2B cells, where aging leads to higher toxic responses. Collectively, our results suggest that photochemical aging may increase SP toxicity through surface modifications that lead to an increased toxic response by activating different molecular pathways.

The most commonly used construction material nowadays is steel-reinforced concrete which underlies corrosion and thus buildings are susceptible for structural collapses. Recently, a new construction material resistant to oxidation and with a higher tensile strength called carbon concrete composite (C3) was developed. The new material allows resource-saving constructions using carbon fiber instead of steel reinforcement materials embedded in a concrete matrix. C3 reinforcements consist of carbon fibers coated with an organic polymer matrix. In this study, abrasive dust from a dry cutting process of two C3 reinforcement materials, as well as a C3 material were investigated with respect to the occurrence of toxic fibers or harmful organic compounds in the inhalable particulate matter (PM) fractions PM2.5 and PM10. It could be shown that the ratio between elemental and organic carbon in PM10 is dependent on the shape of the C3 reinforcement material due to different mechanisms of PM formation. This could have an impact on the toxicity of different C3 reinforcement materials. Harmful fibers according to the World Health Organization (WHO) definition like they are found in asbestos concrete were not found. However, bisphenol A (BPA) as well as the PAHs phenanthrene, anthracene, fluoranthene and pyrene were found due to pyrolysis of the organic matrix material at the edge of the sawblade differentiating the Carcinogenic Equivalency (TEQ) of investigated materials and their PM fractions. Furthermore, derivatives of BPA occurred in abrasive dust from C3 reinforcement materials potentially leading to genotoxicity and reproductive toxicity.

BACKGROUND: Secondary organic aerosols (SOAs) formed from anthropogenic or biogenic gaseous precursors in the atmosphere substantially contribute to the ambient fine particulate matter [PM ≤2.5μm in aerodynamic diameter (PM2.5)] burden, which has been associated with adverse human health effects. However, there is only limited evidence on their differential toxicological impact. OBJECTIVES: We aimed to discriminate toxicological effects of aerosols generated by atmospheric aging on combustion soot particles (SPs) of gaseous biogenic (β-pinene) or anthropogenic (naphthalene) precursors in two different lung cell models exposed at the air-liquid interface (ALI). METHODS: Mono- or cocultures of lung epithelial cells (A549) and endothelial cells (EA.hy926) were exposed at the ALI for 4 h to different aerosol concentrations of a photochemically aged mixture of primary combustion SP and β-pinene (SOAβPIN-SP) or naphthalene (SOANAP-SP). The internally mixed soot/SOA particles were comprehensively characterized in terms of their physical and chemical properties. We conducted toxicity tests to determine cytotoxicity, intracellular oxidative stress, primary and secondary genotoxicity, as well as inflammatory and angiogenic effects. RESULTS: We observed considerable toxicity-related outcomes in cells treated with either SOA type. Greater adverse effects were measured for SOANAP-SP compared with SOAβPIN-SP in both cell models, whereas the nano-sized soot cores alone showed only minor effects. At the functional level, we found that SOANAP-SP augmented the secretion of malondialdehyde and interleukin-8 and may have induced the activation of endothelial cells in the coculture system. This activation was confirmed by comet assay, suggesting secondary genotoxicity and greater angiogenic potential. Chemical characterization of PM revealed distinct qualitative differences in the composition of the two secondary aerosol types. DISCUSSION: In this study using A549 and EA.hy926 cells exposed at ALI, SOA compounds had greater toxicity than primary SPs. Photochemical aging of naphthalene was associated with the formation of more oxidized, more aromatic SOAs with a higher oxidative potential and toxicity compared with β-pinene. Thus, we conclude that the influence of atmospheric chemistry on the chemical PM composition plays a crucial role for the adverse health outcome of emissions. https://doi.org/10.1289/EHP9413.

The ubiquitous use of phthalates in various materials and the knowledge about their potential adverse effects is of great concern for human health. Several studies have uncovered their role in carcinogenic events and suggest various phthalate-associated adverse health effects that include pulmonary diseases. However, only limited information on pulmonary toxicity is available considering inhalation of phthalates as the route of exposure. While in vitro studies are often based on submerged exposures, this study aimed to expose A549 alveolar epithelial cells at the air-liquid interface (ALI) to unravel the genotoxic and oxidative stress-inducing potential of dibutyl phthalate (DBP) with concentrations relevant at occupational settings. Within this scope, a computer modelling approach calculating alveolar deposition of DBP particles in the human lung was used to define in vitro ALI exposure conditions comparable to potential occupational DBP exposures. The deposited mass of DBP ranged from 0.03 to 20 ng/cm2 , which was comparable to results of a human lung particle deposition model using an 8 hours workplace threshold limit value (TLV) of 580 μg/m3 proposed by the Scientific Committee on Occupational Exposure Limits (SCOEL) for the European Union. Comet and Micronucleus assay revealed that DBP induced genotoxicity at DNA and chromosome level in sub-cytotoxic conditions. Since genomic instability was accompanied by increased generation of the lipid peroxidation marker malondialdehyde (MDA), oxidative stress might play an important role in phthalate-induced genotoxicity. The results highlight the importance of adapting in vitro studies to exposure scenarios relevant at occupational settings and reconsidering occupational exposure limits for DBP.

Solid fuel usage in residential heating and cooking is one of the largest sources of ambient and indoor air particulate matter, which causes adverse effects on the health of millions of peoples worldwide. Emissions from solid fuel combustion, such as biomass or coal, are detrimental to health, but toxicological responses are largely unknown. In the present study, we compared the toxicological responses regarding cytotoxicity, inflammation and genotoxicity of spruce (SPR) and brown coal briquette (BCB) combustion aerosols on human alveolar epithelial cells (A549) as well as a coculture of A549 and differentiated human monocytic cells (THP-1) into macrophages exposed at the air-liquid interface (ALI). We included both the high emissions from the first hour and moderate emissions from the third hour of the batch combustion experiment in one ALI system, whereas, in the second ALI system, we exposed the cells during the whole 4-hour combustion experiment, including all combustion phases. Physico-chemical properties of the combustion aerosol were analysed both online and offline. Both SPR and BCB combustion aerosols caused mild cytotoxic but notable genotoxic effects in co-cultured A549 cells after one-hour exposure. Inflammatory response analysis revealed BCB combustion aerosols to cause a mild increase in CXCL1 and CXCL8 levels, but in the case of SPR combustion aerosol, a decrease compared to control was observed.

The aim of this study was to explore the impact of three different standard reference particulate matter (ERM-CZ100, SRM-1649, and SRM-2975) on epigenetic DNA modifications including cytosine methylation, cytosine hydroxymethylation, and adenine methylation. For the determination of low levels of adenine methylation, we developed and applied a novel DNA nucleobase chemical derivatization and combined it with liquid chromatography tandem mass spectrometry. The developed method was applied for the analysis of epigenetic modifications in monocytic THP-1 cells exposed to the three different reference particulate matter for 24 h and 48 h. The mass fraction of epigenetic active elements As, Cd, and Cr was analyzed by inductively coupled plasma mass spectrometry. The exposure to fine dust ERM-CZ100 and urban dust SRM-1649 decreased cytosine methylation after 24 h exposure, whereas all 3 p.m. increased cytosine hydoxymethylation following 24 h exposure, and the epigenetic effects induced by SRM-1649 and diesel SRM-2975 were persistent up to 48 h exposure. The road tunnel dust ERM-CZ100 significantly increased adenine methylation following the shorter exposure time. Two-dimensional scatters analysis between different epigenetic DNA modifications were used to depict a significantly negative correlation between cytosine methylation and cytosine hydroxymethylation supporting their possible functional relationship. Metals and polycyclic aromatic hydrocarbons differently shapes epigenetic DNA modifications.

Cell exposure experiments at the air-liquid interface (ALI) are used increasingly as indicators for health effects and for the impact of aerosols on the lung. Thereby the aerosol particles are kept airborne and can deposit on a cell surface area similar to the human respiratory tract (RT). However, geometry and air flow rates of an ALI system deviate considerably from the RT. As the tissue-delivered particle dose to the lungs (TD) can hardly be measured, computer models of particle deposition are used here to mimic both the particle deposition at ALI and in the RT. An ALI exposure setup (VitroCell GmbH) for an airflow rate of 100 cm(3) min(-1) is selected, where the particle deposition model has been verified experimentally. For the RT we use the hygroscopic lung deposition model of Ferron et al. (2013). Model runs are performed for the particle deposition and for the deposited particles per surface area in both the ALI and the RT. The results show that the ALI-deposited mass is 1-2 orders of magnitude higher than in the alveolar region, because the surface area of the lung region is substantially larger. A particle size range from 40 to 450 nm is identified, where the ratio of both the deposition in a lung region and the deposition at the ALI varies by a factor less than two. Mean values for this ratio are 31 and 101 for the tracheo-bronchial and the alveolar region, respectively. The same size range is found for the ratio of the deposited particles per surface area in a lung region and at the ALI. For this range the mean surface deposition at the ALI is 23- and 1575-times larger than in the tracheo-bronchial and the alveolar lung region, respectively. The effect is partly compensated by different flow rate and cell size.

Tariquidar derivatives have been described as potent and selective ABCG2 inhibitors. However, their susceptibility to hydrolysis limits their applicability. The current study comprises the synthesis and characterization of novel tariquidar-related inhibitors, obtained by bioisosteric replacement of the labile moieties in our previous tariquidar analog UR-ME22-1 (9). CuAAC (“click” reaction) gave convenient access to a triazole core as a substitute for the labile amide group and the labile ester moiety was replaced by different acyl groups in a Sugasawa reaction. A stability assay proved the enhancement of the stability in blood plasma. Compounds UR-MB108 (57) and UR-MB136 (59) inhibited ABCG2 in a Hoechst 33342 transport assay with an IC50 value of about 80 nM and belong to the most potent ABCG2 inhibitors described so far. Compound 57 was highly selective, whereas its PEGylated analog 59 showed some potency at ABCB1. Both 57 and 59 produced an ABCG2 ATPase-depressing effect which is in agreement with our precedent cryo-EM study identifying 59 as an ATPase inhibitor that exerts its effect via locking the inward-facing conformation. Thermostabilization of ABCG2 by 57 and 59 can be taken as a hint to comparable binding to ABCG2. As reference substances, compounds 57 and 59 allow additional mechanistic studies on ABCG2 inhibition. Due to their stability in blood plasma, they are also applicable in vivo. The highly specific inhibitor 57 is suited for PET labeling, helping to further elucidate the (patho)physiological role of ABCG2, e.g. at the BBB., European Journal of Medicinal Chemistry, 191, ISSN:0223-5234, ISSN:1768-3254

Ambient particulate matter (PM) is a leading global environmental health risk. Current air quality regulations are based on airborne mass concentration. However, PM from different sources have distinct chemical compositions and varied toxicity. Connections between emission control measures, air quality, PM composition, and toxicity remain insufficiently elucidated. The current study assessed the composition and toxicity of PM collected in Nanjing, China before, during, and after an air quality intervention for the 2014 Youth Olympic Games. A co-culture model that mimics the alveolar epithelium with the associated macrophages was created using A549 and THP-1 cells. These cells were exposed to size-segregated inhalable PM samples. The composition and toxicity of the PM samples were influenced by several factors including seasonal variation, emission sources, and the air quality intervention. For example, we observed a size-dependent shift in particle mass concentrations during the air quality intervention with an emphasized proportion of smaller particles (PM2.5) present in the air. The roles of industrial and fuel combustion and traffic emissions were magnified during the emission control period. Our analyses revealed that the PM samples demonstrated differential cytotoxic potencies at equal mass concentrations between sampling periods, locations, and time of day, influenced by variations in the predominant emission sources. Coal combustion and industrial emissions were the most important sources affecting the toxicological responses and displayed the least variation in emission contributions between the sampling periods. In conclusion, emission control mitigated cytotoxicity and oxidative stress for particles larger than 0.2 μm, but there was inadequate evidence to determine if it was the key factor reducing the harmful effects of PM0.2.

Recently reported compounds such as UR-COP78 (6) are among the most potent and selective ABCG2 modulators known so far but are prone to rapid enzymatic cleavage at the central benzanilide moiety. In search for more stable analogues, according to a bioisosteric approach, a series of N-(biphenyl-3-yl)quinoline carboxamides was prepared by solid phase and solution phase synthesis. The biphenyl moiety was constructed by Suzuki coupling. Inhibition of ABCB1 and ABCG2 was determined in a calcein-AM and a Hoechst 33342 microplate assay, respectively. Most synthesized compounds selectively inhibited the ABCG2 transporter at submicromolar concentrations with a maximal inhibitory effect (I max) over 90% (e.g., UR-COP228 (22a), IC50 591 nM, I max 109%; UR-COP258 (31), IC50 544 nM, I max 112%), though with lower potency and selectivity than 6. The biphenyl analogues are considerably more stable and demonstrate that the benzanilide core is not a crucial structural feature of quinoline carboxamide-type ABCG2 modulators.

In 2012, the first cases of infection with the Middle East respiratory syndrome coronavirus (MERS-CoV) were identified. Since then, more than 1,000 cases of MERS-CoV infection have been confirmed; infection is typically associated with considerable morbidity and, in approximately 30% of cases, mortality. Currently, there is no protective vaccine available. Replication-competent recombinant measles virus (MV) expressing foreign antigens constitutes a promising tool to induce protective immunity against corresponding pathogens. Therefore, we generated MVs expressing the spike glycoprotein of MERS-CoV in its full-length (MERS-S) or a truncated, soluble variant of MERS-S (MERS-solS). The genes encoding MERS-S and MERS-solS were cloned into the vaccine strain MV vac2 genome, and the respective viruses were rescued (MV vac2 -CoV-S and MV vac2 -CoV-solS). These recombinant MVs were amplified and characterized at passages 3 and 10. The replication of MV vac2 -CoV-S in Vero cells turned out to be comparable to that of the control virus MV vac2 -GFP (encoding green fluorescent protein), while titers of MV vac2 -CoV-solS were impaired approximately 3-fold. The genomic stability and expression of the inserted antigens were confirmed via sequencing of viral cDNA and immunoblot analysis. In vivo , immunization of type I interferon receptor-deficient (IFNAR −/− )-CD46Ge mice with 2 × 10 5 50% tissue culture infective doses of MV vac2 -CoV-S(H) or MV vac2 -CoV-solS(H) in a prime-boost regimen induced robust levels of both MV- and MERS-CoV-neutralizing antibodies. Additionally, induction of specific T cells was demonstrated by T cell proliferation, antigen-specific T cell cytotoxicity, and gamma interferon secretion after stimulation of splenocytes with MERS-CoV-S presented by murine dendritic cells. MERS-CoV challenge experiments indicated the protective capacity of these immune responses in vaccinated mice. IMPORTANCE Although MERS-CoV has not yet acquired extensive distribution, being mainly confined to the Arabic and Korean peninsulas, it could adapt to spread more readily among humans and thereby become pandemic. Therefore, the development of a vaccine is mandatory. The integration of antigen-coding genes into recombinant MV resulting in coexpression of MV and foreign antigens can efficiently be achieved. Thus, in combination with the excellent safety profile of the MV vaccine, recombinant MV seems to constitute an ideal vaccine platform. The present study shows that a recombinant MV expressing MERS-S is genetically stable and induces strong humoral and cellular immunity against MERS-CoV in vaccinated mice. Subsequent challenge experiments indicated protection of vaccinated animals, illustrating the potential of MV as a vaccine platform with the potential to target emerging infections, such as MERS-CoV.

Background: Cold-sensitive mutations in the reactive thiol region of myosin interfere with motor function. Results: Structural and functional data indicate that the mutations induce the preferential population of a state that resembles the ADP-bound state. Conclusion: Gly-680 mutations lead to uncoupling of the reactive thiol region from the surrounding structural elements. Significance: Our results explain how moderate increases in temperature suppress the mutant defects., The cold-sensitive single-residue mutation of glycine 680 in the reactive thiol region of Dictyostelium discoideum myosin-2 or the corresponding conserved glycine in other myosin isoforms has been reported to interfere with motor function. Here we present the x-ray structures of myosin motor domain mutants G680A in the absence and presence of nucleotide as well as the apo structure of mutant G680V. Our results show that the Gly-680 mutations lead to uncoupling of the reactive thiol region from the surrounding structural elements. Structural and functional data indicate that the mutations induce the preferential population of a state that resembles the ADP-bound state. Moreover, the Gly-680 mutants display greatly reduced dynamic properties, which appear to be related to the recovery of myosin motor function at elevated temperatures.

Type I interferon (IFN-α/β) induction upon viral infection contributes to the early antiviral host defense and ensures survival until the onset of adaptive immunity. Many viral infections lead to an acute, transient IFN expression which peaks a few hours after infection and reverts to initial levels after 24 to 36 h. Robust IFN expression often is conferred by specialized plasmacytoid dendritic cells (pDC) and may depend on positive-feedback amplification via the type I IFN receptor (IFNAR). Here, we show that mice infected with Thogoto virus (THOV), which is an influenza virus-like orthomyxovirus transmitted by ticks, mounted sustained IFN responses that persisted up to 72 h after infection. For this purpose, we used a variant of THOV lacking its IFN-antagonistic protein ML, an elongated version of the matrix (M) protein [THOV(ΔML)]. Of note, large amounts of type I IFN were also found in the serum of mice lacking the IFNAR. Early IFN-α expression seemed to depend on Toll-like receptor (TLR) signaling, whereas prolonged IFN-α responses strictly depended on RIG-I-like helicase (RLH) signaling. Unexpectedly, THOV(ΔML)-infected bone marrow-derived pDC (BM-pDC) produced only moderate IFN levels, whereas myeloid DC (BM-mDC) showed massive IFN induction that was IPS-1-dependent, suggesting that BM-mDC are involved in the massive, sustained IFN production in THOV(ΔML)-infected animals. Thus, our data are compatible with the model that THOV(ΔML) infection is sensed in the acute phase via TLR and RLH systems, whereas at later time points only RLH signaling is responsible for the induction of sustained IFN responses.