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Abstract: Exposure to allergens is pivotal in determining sensitization and allergic symptoms in individuals. Pollen grain counts in ambient air have traditionally been assessed to estimate airborne allergen exposure. However, the exact allergen content of ambient air is unknown. We therefore monitored atmospheric concentrations of birch pollen grains and the matched major birch pollen allergen Bet v 1 simultaneously across Europe within the EU-funded project HIALINE (Health Impacts of Airborne Allergen Information Network). Pollen count was assessed with Hirst type pollen traps at 10 l min−1 at sites in France, United Kingdom, Germany, Italy and Finland. Allergen concentrations in ambient air were sampled at 800 l min−1 with a Chemvol® high-volume cascade impactor equipped with stages PM > 10 μm, 10 μm > PM > 2.5 μm, and in Germany also 2.5 μm > PM > 0.12 μm. The major birch pollen allergen Bet v 1 was determined with an allergen specific ELISA. Bet v 1 isoform patterns were analyzed by 2D-SDS-PAGE blots and mass spectrometric identification. Basophil activation was tested in an FcɛR1-humanized rat basophil cell line passively sensitized with serum of a birch pollen symptomatic patient. Compared to 10 previous years, 2009 was a representative birch pollen season for all stations. About 90% of the allergen was found in the PM > 10 μm fraction at all stations. Bet v 1 isoforms pattern did not vary substantially neither during ripening of pollen nor between different geographical locations. The average European allergen release from birch pollen was 3.2 pg Bet v 1/pollen and did not vary much between the European countries. However, in all countries a >10-fold difference in daily allergen release per pollen was measured which could be explained by long-range transport of pollen with a deviating allergen release. Basophil activation by ambient air extracts correlated better with airborne allergen than with pollen concentration. Although Bet v 1 is a mixture of different isoforms, its fingerprint is constant across Europe. Bet v 1 was also exclusively linked to pollen. Pollen from different days varied >10-fold in allergen release. Thus exposure to allergen is inaccurately monitored by only monitoring birch pollen grains. Indeed, a humanized basophil activation test correlated much better with allergen concentrations in ambient air than with pollen count. Monitoring the allergens themselves together with pollen in ambient air might be an improvement in allergen exposure assessment. [Copyright &y& Elsevier]

Reveals that cytochrome P450 CYP1B1 determines susceptibility to 7,12-dimethylbenz[a]anthracene-induced (DMBA) lymphomas. Non-detection of CYP1B1 in cultured embryonic fibroblast or in different tissues; Metabolization of DMBA; Role of extrahepatic P450 in determining susceptibility to chemical carcinogens.

Abstract: Background: Netherton syndrome (NS, MIM 256500) is a potential live threatening autosomal-recessive skin disorder clinically characterized by the trias of congenital erythroderma, hair shaft anomalies and atopic diathesis. It is caused by mutations in the gene SPINK5 resulting in a deficiency of its processed protein named lympho-epithelial Kazal-type related inhibitor (LEKTI). LEKTI controls the activity of several serine proteases in the skin that are involved in terminal differentiation. Loss of LEKTI results in protease hyperactivity, increased degradation of intercellular junctions, reduced stratum corneum adhesion and impaired skin barrier function. Today NS can only be treated symptomatically. Objective: Does gene transfer offer a therapeutic option for NS in the future? Methods: A recombinant adeno-associated virus type 2 vector was constructed containing the full length cDNA (rAAV2/C-SPINK5) of functional human LEKTI. Infectious virus particles were used for transfection of LEKTI-deficient-keratinocytes of NS patients in vitro. Results: Gene transfer of SPINK5 in NS-keratinocytes led to a five-fold increase in mRNA expression of SPINK5 reaching almost 75% of normal value. The functionality of the expressed LEKTI was proven in a hydrolytic activity assay demonstrating that the activity of LEKTI after gene transfer increased closely to the level seen in keratinocytes of healthy individuals. Conclusion: The results provide first evidence that gene transfer of SPINK5 results in increased LEKTI activity in NS-keratinocytes, thus offering a rational to further pursue such a gene therapy approach for NS. [Copyright &y& Elsevier]

Airborne pollen is a recognized biological indicator and its monitoring has multiple uses such as providing a tool for allergy diagnosis and prevention. There is a knowledge gap related to the distribution of pollen traps needed to achieve representative biomonitoring in a region. The aim of this manuscript is to suggest a method for setting up a pollen network (monitoring method, monitoring conditions, number and location of samplers etc.). As a case study, we describe the distribution of pollen across Bavaria and the design of the Bavarian pollen monitoring network (ePIN), the first operational automatic pollen network worldwide. We established and ran a dense pollen monitoring network of 27 manual Hirst-type pollen traps across Bavaria, Germany, during 2015. Hierarchical cluster analysis of the data was then performed to select the locations for the sites of the final pollen monitoring network. According to our method, Bavaria can be clustered into three large pollen regions with eight zones. Within each zone, pollen diversity and distribution among different locations does not vary significantly. Based on the pollen zones, we opted to place one automatic monitoring station per zone resulting in the ePIN network, serving 13 million inhabitants. The described method defines stations representative for a homogeneous aeropalynologically region, which reduces redundancy within the network and subsequent costs (in the study case from 27 to 8 locations). Following this method, resources in pollen monitoring networks can be optimized and allergic citizens can then be informed in a timely and effective way, even in larger geographical areas. Unlabelled Image • The first automatic pollen monitoring network in the world was built in Bavaria, based on the pollen robot BAA500. • Collapsing a dense network by clustering analysis determined the number and position of monitoring stations. • Bavaria (Germany) can be clustered in 3 pollen zones and 8 sub-zones, based on airborne pollen concentrations. • In the studied network, the most abundant pollen types in Bavaria are: Pinus , Betula , Urticaceae and Poaceae. • In the studied network, the main pollination period in Bavaria ranges from February (Alnus) to October (Ambrosia). [ABSTRACT FROM AUTHOR]

Background The level of symptoms in pollen allergy sufferers and users of the Patient's Hayfever Diary (PHD), does not directly reflect the total amount of pollen in the air. It is necessary to explain the symptom load and thus the development of allergic symptoms and to determine which environmental factors, besides the pollen load, influence variables. It seems reasonable to suspect allergen content because the amount of allergen varies throughout seasons and regions and is not always correlated with the total pollen amount. Methods Data on the allergen content of ambient air (Bet v 1 and Phl p 5) from 2009 until 2011 was used to compare the respective pollen and symptom loads for study regions in Austria, Germany, France and Finland. Results Our findings suggest that allergen amount (Bet v 1/Phl p 5) has a strong but regionally dependent impact on the symptom load of pollen allergy sufferers. Peak symptom loads can be traced with peak allergen loads. The influence of other important aeroallergens should also be assessed during the pollen season. Conclusion Allergen concentrations have an impact on pollen allergy sufferers although not as clear as assumed previously. The pattern of pollen load and major allergen content distribution does not directly explain the symptom load pattern, although significant positive correlations were found. Thus, monitoring of symptoms via voluntary crowdsourcing should be considered for future pollen and symptom forecasts in order to support pollen allergy sufferers. [ABSTRACT FROM AUTHOR]

Less vegetated mountainous areas may provide better conditions for allergy sufferers. However, atmospheric transport can result in medically relevant pollen loads in such regions. The majority of investigations has focused on the pollen load, expressed as daily averages of pollen per cubic meter of air (pollen grains/m³); however, the severity of allergic symptoms is also determined by the actual allergen content of this pollen, its pollen potency, which may differ between high and low altitudes. We analysed airborne birch and grass pollen concentrations along with allergen content (birch: Bet v 1, grass: Phl p 5) at two different altitudes (734 and 2650 m a.s.l.) in the Zugspitze region (2009–2010). Back-trajectories were calculated for the high altitude site and for specific days with abrupt increases in pollen potency. We observed several days with medically relevant pollen concentrations at the highest site. In addition, a few days with pollen were not associated with allergens and vice versa . The calculated seasonal mean allergen release per pollen grain was 1.8–3.3 pg Bet v 1 and 5.7 pg Phl p 5 in the valley and 1.1–3.7 pg Bet v 1 and 0.7–1.5 pg Phl p 5 at the high altitude site. Back-trajectories revealed that high pollen potency at the higher site was generally associated with south-westerly to south-easterly (birch), or northerly (grass) wind directions. By investigating days with sudden increases in pollen potency, however, it was difficult to draw definitive conclusions on long- or short-range transport. Our findings suggest that people allergic to pollen might suffer less at higher altitudes and further indicate that a risk assessment relying on the actual concentration of airborne pollen does not necessarily reflect the actual allergy exposure of individuals. [ABSTRACT FROM AUTHOR]

Background: Pollen are monitored in Europe by a network of about 400 pollen traps, all operated manually. To date, automated pollen monitoring has only been feasible in areas with limited variability in pollen species. There is a need for rapid reporting of airborne pollen as well as for alleviating the workload of manual operation. We report our experience with a fully automated, image recognition-based pollen monitoring system, BAA500. Methods: The BAA500 sampled ambient air intermittently with a 3-stage virtual impactor at 60 m3/h in Munich, Germany. Pollen is deposited on a sticky surface that was regularly moved to a microscope equipped with a CCD camera. Images of the pollen were constructed and compared with a library of known samples. A Hirst-type pollen trap was operated simultaneously. Results: Over 480,000 particles sampled with the BAA500 were both manually and automatically identified, of which about 46,000 were pollen. Of the automatically reported pollen, 93.3% were correctly recognized. However, compared with manual identification, 27.8% of the captured pollen were missing in the automatic report, with most reported as unknown pollen. Salix pollen grains were not identified satisfactorily. The daily pollen concentrations reported by a Hirst-type pollen trap and the BAA500 were highly correlated (r = 0.98). Conclusions: The BAA500 is a functional automated pollen counter. Its software can be upgraded, and so we expected its performance to improve upon training. Automated pollen counting has great potential for workload reduction and rapid online pollen reporting. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Abstract: In the present study V79 Chinese hamster cells were genetically engineered for stable expression of the cytochromes P450 1A1, 1A2, 1B1, and 2E1 from man and mouse to investigate species-specific differences in the regioselective metabolism and toxicity of phenanthrene (Phe), the simplest polycyclic aromatic hydrocarbon (PAH) forming a bay-region. Phe is present in various environmental samples and serves as a model substrate for PAH exposure in human biomonitoring studies. For this reason we explored metabolite profiles and metabolite-dependent cytotoxic activities in vitro. The total turnover of CYP-mediated transformation of Phe was as follows: human CYP1B1>CYP1A1>CYP1A2≫CYP2E1, and for mouse CYP1A2≫CYP2E1>CYP1A1. Striking species differences were seen as mouse CYP1B1 did not activate Phe at all, but human CYP1B1 exhibited a significant metabolic turnover comparable to CYP1A1 and CYP1A2. In vivo studies monitoring the whole blood Phe elimination in CYP1A2 knockout and wild-type mice after oral administration confirmed involvement of CYP1A2 in the bioactivation of Phe, but other processes must contribute also. Our data suggest that in humans not only CYP1A2 expressed solely in the liver plays a crucial role in Phe metabolism, but also constitutively expressed extrahepatic CYP1B1 in tissues such as lung, kidney or intestine. This finding will substantially improve the validity of human biomonitoring studies using individual Phe metabolites for the assessment of PAH exposure. [Copyright &y& Elsevier]

Abstract: Cytochrome P4501B1 (CYP1B1), a polycyclic aromatic hydrocarbon (PAH) metabolizing CYP, is genetically polymorphic in humans and may be involved in the individual susceptibility to chemical-induced cancer. In the present study, genotype and haplotype frequencies of four single nucleotide polymorphisms (SNPs) in CYP1B1 that cause amino acid changes (Arg-Gly at codon 48, Ala-Ser at codon 119, Leu-Val at codon 432 and Asn-Ser at codon 453) were studied in 150 cases suffering from head and neck squamous cell carcinoma (HNSCC) and in an equal number of controls. A significant difference was observed for the distribution of variant genotypes of Arg48Gly (CYP1B1*2) and Ala119Ser (CYP1B1*2) polymorphisms of CYP1B1 in cases versus controls. No significant differences were observed for the distribution of variant genotypes-Leu432Val (CYP1B1*3) and Asn453Ser (CYP1B1*4), respectively. When the four SNPs were analyzed using a haplotype approach, SNPs at codon 48 (Arg48Gly) and codon 119 (Ala119Ser) exhibited complete linkage disequilibrium (LD) in all the cases and controls. Significant differences in the distribution of the two haplotypes (G-T-C-A and G-T-G-A) were observed both in the cases and in controls. Furthermore, our data indicates a several fold increase in risk in the cases who use tobacco (cigarette smoking or tobacco chewing) or alcohol with the variant genotypes of CYP1B1 (CYP1B1*2 and CYP1B1*3) suggesting the role of gene–environment interaction in the susceptibility to HNSCC. [Copyright &y& Elsevier]

Abstract: Objectives: In previous studies it could be demonstrated that methacrylic acid (MA) is an intermediate in the metabolism of unpolymerized dental comonomers, released from dental restorative materials. This study was performed to identify the possible dental material intermediate 2,3-epoxymethacrylic acid (2,3-EMA) from MA in human liver microsomes. Most epoxy compounds are regarded as highly toxic substances. Methods: The formation and hydrolysis were studied in defined systems containing only MA and human liver microsomes at 37°C. Hydrolysis was inhibited by cyclohexene oxide, a competitive inhibitor of epoxide hydrolase. The reaction product 2,3-EMA was analyzed by the headspace gas chromatography–mass spectrometry. After 5, 30, and 60min samples were taken and analyzed. Results: For the reaction of MA to 2,3-EMA the average conversion rate was about 5% within 1h. It was found that without cyclohexene oxide the rate constant of enzymatic hydrolysis at pH 7.4 was about 10 times higher than the rate constant of the formation from MA in combination with cyclohexene oxide (k =8.3 versus 0.83μmol/lmin), indicating an instability and thus a high reactivity of 2,3-EMA. The formation of the MA intermediate 2,3-EMA was not observed when heat-inactivated liver microsomes were used (controls). Significance: It could be clearly demonstrated that 2,3-EMA is a product of dental material metabolisms in biological systems. Therefore, increased toxicity might occur on dental restorative materials which are able to release (co)monomers which can be metabolized to MA. [Copyright &y& Elsevier]

The daily pollen forecast provides crucial information for allergic patients to avoid exposure to specific pollen. Pollen counts are typically measured with air samplers and analyzed with microscopy by trained experts. In contrast, this study evaluated the effectiveness of identifying the component pollens using the metabolites extracted from an air-sampled pollen mixture. Ambient air-sampled pollen from Munich in 2016 and 2017 was visually identified from reference pollens and extracts were prepared. The extracts were lyophilized, rehydrated in optimal NMR buffers, and filtered to remove large proteins. NMR spectra were analyzed for pollen associated metabolites. Regression and decision-tree based algorithms using the concentration of metabolites calculated from the NMR spectra outperformed algorithms using the NMR spectra themselves as input data for pollen identification. Categorical prediction algorithms trained for low, medium, high, and very high pollen count groups had accuracies of 74% for the tree, 82% for the grass, and 93% for the weed pollen count. Deep learning models using convolutional neural networks performed better than regression models using NMR spectral input, and were the overall best method in terms of relative error and classification accuracy (86% for tree, 89% for grass, and 93% for weed pollen count). This study demonstrates that NMR spectra of air-sampled pollen extracts can be used in an automated fashion to provide taxa and type-specific measures of the daily pollen count. • This study identified the component pollens from metabolites extracted from air-sampled pollen. • Deep learning methods trained on NMR spectra directly were the best performing algorithms. • Automated analyses of air-sampled pollen extracts using NMR can provide the daily pollen count. [ABSTRACT FROM AUTHOR]

There is high demand for online, real-time and high-quality pollen data. To the moment pollen monitoring has been done manually by highly specialized experts. Here we evaluate the electronic Pollen Information Network (ePIN) comprising 8 automatic BAA500 pollen monitors in Bavaria, Germany. Automatic BAA500 and manual Hirst-type pollen traps were run simultaneously at the same locations for one pollen season. Classifications by BAA500 were checked by experts in pollen identification, which is traditionally considered to be the "gold standard" for pollen monitoring. BAA500 had a multiclass accuracy of over 90%. Correct identification of any individual pollen taxa was always >85%, except for Populus (73%) and Alnus (64%). The BAA500 was more precise than the manual method, with less discrepancies between determinations by pairs of automatic pollen monitors than between pairs of humans. The BAA500 was online for 97% of the time. There was a significant correlation of 0.84 between airborne pollen concentrations from the BAA500 and Hirst-type pollen traps. Due to the lack of calibration samples it is unknown which instrument gives the true concentration. The automatic BAA500 network delivered pollen data rapidly (3 h delay with real-time), reliably and online. We consider the ability to retrospectively check the accuracy of the reported classification essential for any automatic system. • A fully automatic pollen Network was built in Bavaria, based on the system BAA500. • BAA500 had a multiclass accuracy of over 90%. • There was a significant correlation of 0.84 between BAA500 and Hirst data. [ABSTRACT FROM AUTHOR]