BackgroundDuring the last 250 years, the level of exposure to combustion-derived particles raised dramatically in western countries, leading to increased particle loads in the ambient air. Among the environmental particles, diesel exhaust particulate matter (DEPM) plays a special role because of its omnipresence and reported effects on human health. During recent years, a possible link between air pollution and the progression of atherosclerosis is recognized. A central effect of DEPM is their impact on the endothelium, especially of the alveolar barrier. In the present study, a complex 3D tetraculture model of the alveolar barrier was used in a dose-controlled exposure scenario with realistic doses of DEPM to study the response of endothelial cells.ResultsTetracultures were exposed to different doses of DEPM (SRM2975) at the air-liquid-interface. DEPM exposure did not lead to the mRNA expression of relevant markers for endothelial inflammation such as ICAM-1 or E-selectin. In addition, we observed neither a significant change in the expression levels of the genes relevant for antioxidant defense, such as HMOX1 or SOD1, nor the release of pro-inflammatory second messengers, such as IL-6 or IL-8. However, DEPM exposure led to strong nuclear translocation of the transcription factor Nrf2 and significantly altered expression of CYP1A1 mRNA in the endothelial cells of the tetraculture.ConclusionIn the present study, we demonstrated the use of a complex 3D tetraculture system together with a state-of-the-art aerosol exposure equipment to study the effects of in vivo relevant doses of DEPM on endothelial cells in vitro. To the best of our knowledge, this study is the first that focuses on indirect effects of DEPM on endothelial cells of the alveolar barrier in vitro. Exposure to DEPM led to significant activation and nuclear translocation of the transcription factor Nrf2 in endothelial cells. The considerably low doses of DEPM had a low but measurable effect, which is in line with recent data from in vivo studies.Electronic supplementary materialThe online version of this article (doi:10.1186/s12989-017-0186-4) contains supplementary material, which is available to authorized users.

Quantitative data on sensitization potency of chemicals has long been derived from in vivo data obtained with the local lymph node assay (LLNA) in mice. The minimum concentration of a chemical that can induce a sensitization response, i.e. the chemical's sensitization potency, is an essential value in quantitative risk assessment (Mackay et al., 2013). In vitro methods intended to reduce or replace animal testing in this area need to be able to predict hazard and also to categorize the potency of sensitizers.We and others have shown sensitizer-induced CD86 and CD54 upregulation on monocytic THP-1 cells as a model for DC activation (Bocchietto et al., 2007;Goebel et al., 2014;Krutz et al., 2015;Tietze and Blömeke, 2008;Yoshida et al., 2003). However, the impact of the presence of keratinocytes on the activation of THP-1 cells has not yet been fully investigated.In this study, we used our HaCaT/THP-1 coculture setup (Hennen et al., 2011) to study the impact of keratinocytes on the expression of CD86 and CD54 on THP-1 after treatment with a set of 14 sensitizers and 10 non-sensitizers: We evaluated (1) the magnitude of CD86 and CD54 upregulation in the coculture model in comparison to THP-1 monoculture, (2) the sensitivity, specificity and accuracy of the HaCaT/THP-1 coculture model to IntroductionThe hallmarks of chemical sensitizers are their ability to form antigenic haptenated proteins, to induce inflammatory responses in keratinocytes, and to induce maturation of dendritic cells (DC) needed for efficient activation of naïve T cells (OECD, 2012).DC activation by sensitizers causes the upregulation of costimulatory molecules such as CD86 (Linsley et al., 1991), CD54 (Grakoui et al., 1999;Comrie et al., 2015), and other surface molecules (reviewed by Hubo et al., 2013) that are involved in antigen presentation. After interaction with their counterparts on T cells, these surface molecules generate a costimulatory signal (signal 2) that synergizes with the T cell receptor-mediated signal (signal 1) to promote an adaptive immune response.Keratinocytes exposed to sensitizers release proinflammatory or immunomodulatory cytokines (Gober and Gaspari, 2008;Pasparakis et al., 2014). These factors can trigger DC activation and/or DC mobilization (reviewed by Kaplan et al., 2012). This role of adjacent keratinocytes may therefore have a significant impact on the strength of the chemical-induced DC response. Accordingly, we postulated that keratinocytes impact on the quantitative response of DC to skin sensitizing chemicals. Research Article Keratinocytes Improve Prediction of Sensitization Potential and Potency of Chemicals with THP-1 Cells Jennifer Hennen and Brunhilde BlömekeDepartment of Environmental Toxicology, Trier University, Trier, Germany SummaryIn vitro approaches to address key steps of chemical-induced skin sensitization have been developed, but there is uncertainty how keratinocytes, which play a crucial role not only regarding xenobiotic metabolism but also skin inflammation, impact on a chemical's potential ...

Under simulated hair dye use conditions, a significantly lower degree of cross-elicitation to ME-PPD (30%) was observed than previously reported for PPD (32 of 38, 84%). Additionally, a decreased cross-elicitation strength was observed across all three patch test grades, likely reflecting the reduced skin-sensitization properties of ME-PPD. Consequently, careful dermatological evaluation is required to assess cross-reactivity to ME-PPD in patients allergic to hair dyes.

Respiratory sensitization as a consequence of exposure to chemical products has increased over the last decades, leading to an increase of morbidity. The increased use of synthetic compounds resulted in an exponential growth of substances to which we are potentially exposed on a daily basis. Some of them are known to induce respiratory sensitization, meaning that they can trigger the development of allergies. In the past, animal studies provided useful results for the understanding of mechanisms involved in the development of respiratory allergies. However, the mechanistic understanding of the involved cellular effects is still limited. Currently, no in vitro or in vivo models are validated to identify chemical respiratory sensitizers. Nonetheless, chemical respiratory sensitizers elicit a positive response in validated assays for skin sensitization. In this review, we will discuss how these assays could be used for respiratory sensitization and if necessary, what can be learnt from these assays to develop a model to assess the respiratory sensitizing potential of chemicals. In the last decades, much work has been done to study the respiratory toxicity of inhaled compounds especially in developing in vitro assays grown at the air-liquid interface. We will discuss how possibly the tests currently used to investigate general particle toxicity could be transformed to investigate respiratory sensitization. In the present review, we describe the most known mechanism involved in the sensitization process and the experimental in vivo and alternative in vitro models, which are currently available and how to adapt and improve existing models to study respiratory sensitization.

represents the most prevalent occupational disease of the lungs in developed countries and occupational exposure to dusts, gases, fumes, vapors, and chemicals is responsible for 16% of asthma in adults (Torén and Blanc, 2009). Around a hundred chemicals were described to act as respiratory sensitizers (Bloemen et al., 2009), among which different chemical classes such as acid anhydrides, diisocyanates, and chloroplatinate salts were identified.Protein allergens (or high molecular weight; HMW compounds), which are also a potential cause of occupational asthma, are the primary cause for the development of respiratory allergy in the general population. Certain proteins from environmental sources such as pollen, animal dander and house dust mites (HDM) are common causes of asthma. Depending on the geographical location, 50 to 85% of people with asthma are allergic to certain HDM proteins (Gregory and Lloyd, 2011).An early identification of compounds with the potential to act as respiratory sensitizers is still difficult. This is due to an incomplete understanding of the systemic mechanisms involved in the development of respiratory sensitization and to the absence of fit-for-purpose, validated, or even widely accepted in vivo models or in vitro assays to identify respiratory sensitizers. Currently,

The monoterpene ascaridole, a fairly stable endoperoxide found in essential oils such as tea tree oil can provoke allergic contact dermatitis which has been evidenced under patch test conditions. However, concomitantly we observed irritative skin reactions that demand further data underlining the sensitization potential of ascaridole. Here, we studied the effects of ascaridole on dendritic cell (DC) activation and protein reactivity, 2 key steps of chemical-induced skin sensitization. Treatment of human monocyte-derived DC with ascaridole found support for full DC maturation, a capability of sensitizers but not irritants. It induced significant upregulation of the expression of the costimulatory molecules CD86, CD80, CD40, and the adhesion molecule CD54 in a time-dependent manner. Maturation was accompanied by release of proinflammatory cytokines interleukin (IL)-1ß, tumor necrosis factor-α, IL-6, and IL-8. Similar to other chemical skin sensitizers including hydroperoxides, we observed a certain reactivity of ascaridole toward cysteine- but not lysine-containing peptides. During recent years, evidence accumulated for a radical mechanism as trigger for protein reactivity of peroxides. Treatment of the fairly stable endoperoxide ascaridole with iron as radical inducer ("activated ascaridole") resulted in cysteine peptide reactivity exceeding by far that of ascaridole itself. Furthermore, activated ascaridole showed increased potential for induction of the Nrf2 target gene heme oxygenase 1 and upregulation of CD86 and CD54 on THP-1 cells, an established DC surrogate. These results indicate that radical formation could be involved in the steps leading to skin sensitization induced by the endoperoxide ascaridole.

Background: Positive patch test reactions to mixtures of oxidized terpenes con-

Regulation requires that skin sensitization hazard and potency of chemicals is assessed by a combination of in vitro and in chemico studies and that in vivo testing is conducted only as a last resort (EU, 2016).Given the complexity of the biological mechanisms underlying skin sensitization, it is generally recognized that a combination of mechanistically-based test methods is needed for both skin sensitization hazard assessment and potency prediction (Jowsey et al., 2006;OECD, 2014). Considerable progress has been made in the last decade regarding the development, validation, and adoption of alternative methods for skin sensitization hazard identification. In particular, a number of OECD Test Guidelines (TG) have been adopted that address the three major key events of the adverse outcome pathway (AOP) leading to skin sensitization. These are i) the covalent binding of electrophilic substances to nucleophilic centers in skin proteins as the molecular initiating event or first key event (OECD, 2019),