Chemical sensitization and allergotoxicology.

Chemical sensitization remains an important environmental and occupational health issue. A wide range of substances have been shown to possess the ability to induce skin sensitization or respiratory sensitization. As a consequence, there is a need to have appropriate methods to identify sensitizing agents. Although a considerable investment has been made in exploring opportunities to develop methods for hazard identification and characterization, there are, as yet, no validated nonanimal methods available. A state of the art of the different in vitro approaches to identify contact and respiratory capacity of chemicals is covered in this chapter.

Biomonitoring and biomarkers to unravel the risks from prenatal environmental exposures for later health outcomes.

An increasing number of studies have addressed the concern that environmental pollutants may contribute to the early origin of diseases. Epidemiologic studies suggest that prenatal exposure to air pollutants, several food contaminants, and chemicals present in consumer products are associated with nongenetically transmitted adverse health effects, which manifest after birth. Changes in neurobehavior, sexual development, the prevalence of asthma and allergy, and growth curves have been shown to be associated with pollutant exposure at early life stages. This review focuses on human molecular epidemiologic studies that contribute knowledge by introducing biomarker measurements to obtain a mechanistic understanding of the relation between early life exposures and health outcome. It has been hypothesized that subtle effects induced by pollutant exposure during development can lead to functional deficits and altered programming, which leads to increased disease or dysfunction risk later in life. Biomarker analysis may provide sensitive tools to trace these subtle changes and obtain mechanistic insight about the causal pathway between external exposure and health effects in human population studies. Biomarkers of exposure can be measured in mothers before conception, during pregnancy, or after birth. Different biological tissues-such as peripheral or cord blood samples, hair samples, meconium, and urine-provide specific information that reflects the actual dose during pregnancy or at birth. Biomarkers of effect may include changes in hormone concentrations, oxidative stress variables, changes in gene expression levels, and epigenetic changes.

Microarray analyses in dendritic cells reveal potential biomarkers for chemical-induced skin sensitization.

The assessment of the skin sensitising capacity of chemicals is up to now investigated using in vivo animal tests. However there has been an increasing public and governmental concern regarding the use of animals for chemical screening. This has raised the need for the development of validated in vitro alternatives. Langerhans cells are potent antigen-presenting cells that play a crucial role in the development of allergic contact dermatitis. We used CD34(+) progenitor-derived dendritic cells from cord blood as an in vitro alternative for Langerhans cells. The cells were exposed to four contact allergens (nickel sulphate, dinitrochlorobenzene, oxazolone and eugenol) and two irritants (sodium dodecyl sulphate and benzalkonium chloride) for 3, 6, 12 and 24h. Using microarray analyses we revealed a set of 25 genes with an altered gene expression pattern after exposure to allergens and not to irritants. Five out of these 25 genes were selected and their gene expression changes were confirmed with real-time reverse transcriptase polymerase chain reaction. The list of 25 genes represent valuable candidates to be further evaluated for their capacity to predict the sensitizing potential of different classes of chemicals in studies using a more extended set of (non) allergic substances.

Gene expression signatures in CD34+-progenitor-derived dendritic cells exposed to the chemical contact allergen nickel sulfate.

The detection of the sensitizing potential of chemicals is of great importance to industry. A promising in vitro alternative to the currently applied animal assays for sensitization testing makes use of dendritic cells (DCs) that have the capability to process and present antigens to naive T cells and induce their proliferation. Here, we studied changes in gene expression profiles after exposing DCs to the contact allergen nickel sulfate. CD34+-progenitor-derived DCs, initiated from 3 different donors, were exposed to 60 microM nickel sulfate, during 0.5, 1, 3, 6, 12 and 24 h. cDNA microarrays were used to assess the transcriptional activity of about 11,000 genes. Significant changes in the expression of 283 genes were observed; 178 genes were up-regulated and 93 down-regulated. These genes were involved in metabolism, cell structure, immune response, transcription, signal transduction, transport, and apoptosis. No functional information was found for 74 genes. Real-time RT-PCR was used to confirm the microarray results of 12 genes. In addition, 3 DC maturation markers not present on the microarrays (DEC205, DC LAMP and CCR7) were analyzed using real-time RT-PCR and found to be up-regulated at several time points. Our data indicate that a broad range of biological processes is influenced by nickel. Some processes are clearly linked to the immune response and DC maturation, others may indicate a toxic effect of nickel.

Capacity of CD34+ progenitor-derived dendritic cells to distinguish between sensitizers and irritants.

In an attempt to develop an in vitro test to identify contact sensitizers, mostly dendritic cells (DCs) derived from monocytes (Mo-DC) have been used. Less is known about the potency of DC derived from CD34+ progenitors (CD34-DC) for in vitro allergen testing. CD34+ progenitor derived DC were exposed to nine well-known allergens (one weak, three moderate and five strong allergens) and two irritants. Surface marker expression (CD86, CD83 and HLA-DR) and cytokine production (IL-6, IL-12 and TNF-alpha) were analyzed after 24 h exposure to these chemicals. All allergens tested induced a significant increase in at least one of the DC surface markers. In contrast, none of the irritants tested were able to significantly upregulate membrane marker expression in exposed DC. The level of upregulation of CD86, CD83 and HLA-DR was dependent on the nature and concentration of the chemical, but not on the classification of the allergen. Changes in cytokine production (IL-6, IL-12 and TNF-alpha) were not consistently related to exposure to an allergen. Based on these results, we conclude that the in vitro test using CD34-DC has the capacity to distinguish between allergens and irritants based on altered phenotypic characteristics.

Cytokine transcript profiling in CD34+-progenitor derived dendritic cells exposed to contact allergens and irritants.

We here investigated wether genes encoding the interleukins IL-1beta, IL-6 and IL-8, and the chemokines CCL2, CCL3, CCL3L1 and CCL4 are useful markers for sensitization testing in CD34+-progenitor derived dendritic cells (CD34-DC). CD34-DC from at least three donors were exposed during 0.5 up to 24h to the chemical sensitizers nickel sulphate, oxazolone, 2,4-dinitrochlorobenzene (DNCB) and eugenol, and to the irritants sodium dodecyl sulphate (SDS) and benzalkonium sulphate (BC). mRNA expression was evaluated using real-time RT-PCR. We observed a large inter-individual variation in mRNA expression in CD34-DC exposed to the chemicals. No or limited effects on expression were observed for the irritant BC and the weak sensitizer eugenol. All other chemicals modulated the transcript levels of most cytokines that were investigated. Most of the time, no clear-cut distinctions could be made between the sensitizers and SDS. After 24 h, consistent upregulatory effects of all sensitizing compounds on transcript expression of CCL2, CCL3 and CCL4 were observed, whereas SDS (and BC) had no effect. Our findings suggest that the CCL2, CCL3 and CCL4 genes may be selective end-point markers in the CD34-DC model to discern chemical sensitizers from irritants.

Immunologic biomarkers in relation to exposure markers of PCBs and dioxins in Flemish adolescents (Belgium).

In this study, we investigated 17- to 18-year-old boys and girls to determine whether changes in humoral or cellular immunity or respiratory complaints were related to blood serum levels of polychlorinated biphenyls (PCBs) and dioxin-like compounds after lifetime exposure in Flanders (Belgium). We obtained blood samples from and administered questionnaires to 200 adolescents recruited from a rural area and two urban suburbs. Physicians recorded medical history and respiratory diseases. We measured immunologic biomarkers such as differential blood cell counts, lymphocyte phenotypes, and serum immunoglobulins. As biomarkers of exposure, we determined the serum concentrations of PCBs (PCB 138, PCB 153, and PCB 180) and dioxin-like compounds [chemical-activated luciferase expression (CALUX) bioassay]. The percentages of eosinophils and natural killer cells in blood were negatively correlated with CALUX toxic equivalents (TEQs) in serum (p = 0.009 and p = 0.05, respectively). Increased serum CALUX TEQs resulted in an increase in serum IgA levels (p = 0.05). Furthermore, levels of specific IgEs (measured by radioallergosorbent tests) of cat dander, house dust mite, and grass pollen were also significantly and negatively associated with the CALUX TEQ, with odds ratios (ORs) equal to 0.63 [95% confidence interval (CI), 0.42-0.96], 0.68 (0.5-0.93), and 0.70 (0.52-0.95), respectively. In addition, reported allergies of the upper airways and past use of antiallergic drugs were negatively associated with CALUX TEQs, with ORs equal to 0.66 (0.47-0.93) and 0.58 (0.39-0.85), respectively. We found a negative association between IgGs and marker PCBs in serum (p = 0.009). This study shows that immunologic measurements and respiratory complaints in adolescents were associated with environmental exposure to polyhalogenated aromatic hydrocarbons (PHAHs). The negative correlation between PHAHs and allergic responses in adolescents suggested that exposure may entail alterations in the immune status.

Phenotypic alterations and IL-1beta production in CD34(+) progenitor- and monocyte-derived dendritic cells after exposure to allergens: a comparative analysis.

Dendritic cells (DC) have been shown to capture and process antigens and play an initiating role in contact sensitization. Cells with dendritic morphology can be generated in vitro either from CD34(+) cord blood cells or from CD14(+) peripheral monocytes. The aim of this study was to determine the state of maturation/activation of both populations after exposure to several concentrations of four well-established model allergens (nickel sulfate, eugenol, alpha-hexylcinnamaldehyde and 2,4,6-trinitrobenzene sulfonic acid) or the irritant sodium dodecyl sulfate. We analyzed the surface expression of CD86, CD83 and HLA-DR and the production of IL-1beta. DC from the two sources were generated separately in two laboratories, but challenged using identical test protocols. Using both DC populations it was possible to detect the allergens under investigation, though minor differences regarding effective concentrations were noted. The non-responsiveness of CD34-DC to CIN was probably due to non-optimal concentrations. Ni(2+), known as a moderate allergen in vivo, showed the most prominent effect in both cell systems. CD86 expression was the most reliable phenotypic marker for the in vitro identification of allergens. Due to substantial individual variations it was difficult to draw any definite conclusions as to the relevance of IL-1beta production as an activation endpoint. We conclude that both test systems are able to respond to allergens, but CD34-DC must be exposed to higher concentrations to demonstrate significant phenotypic changes. On the other hand, Mo-DC from only some of the donors reacted to allergens, in contrast to CD34-DC, which responded to allergens irrespective of the donor, thus necessitating the use of Mo-DC cultures from several blood donors.