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Perfluoroalkyl and polyfluoroalkyl substances (PFAS) are a class of synthetic compounds that have widespread use in consumer and industrial applications. PFAS are considered environmental pollutants that have various toxic properties, including effects on the immune system. Recent human studies indicate that prenatal exposure to PFAS leads to suppressed immune responses in early childhood. In this study, data from the Norwegian BraMat cohort was used to investigate transcriptomics profiles in neonatal cord blood and their association with maternal PFAS exposure, anti-rubella antibody levels at 3 years of age and the number of common cold episodes until 3 years. Genes associated with PFAS exposure showed enrichment for immunological and developmental functions. The analyses identified a toxicogenomics profile of 52 PFAS exposure-associated genes that were in common with genes associated with rubella titers and/or common cold episodes. This gene set contains several immunomodulatory genes (CYTL1, IL27) as well as other immune-associated genes (e.g.EMR4P, SHC4, ADORA2A). In addition, this study identifiedPPARDas a PFAS toxicogenomics marker. These markers can serve as the basis for further mechanistic or epidemiological studies. This study provides a transcriptomics connection between prenatal PFAS exposure and impaired immune function in early childhood and supports current views on PPAR- and NF-κB-mediated modes of action. The findings add to the available evidence that PFAS exposure is immunotoxic in humans and support regulatory policies to phase out these substances. [ABSTRACT FROM AUTHOR]

In vitrogene profiling studies have associated the molecular pathways of Nrf2-Keap1 and Toll-like receptor (TLR) signaling with skin sensitization. In this study, the role of these pathways in the regulation of protein biomarkers for skin sensitization was further elucidated using transient gene knock-down of key components of the signaling cascades in HaCaT cells after exposure to dinitrochlorobenzene (DNCB). The effect of targeting these pathways was established through evaluation of heme oxygenase1 (HMOX1) and interleukin (IL)-8 production. These experiments showed that Nrf2 is not involved in regulating HMOX1 after exposure to DNCB, but that activation of TLR signaling moderates the expression of HMOX1. The regulation of IL-8 depended on Nrf2, but also on the Toll/interleukin-1 receptor (TIR)-domain-containing adapter-inducing interferon-β(TRIF) adaptor protein in TLR signaling. This study provides new insights into the regulation of HMOX1 and IL-8, but the exact regulating mechanisms remain to be further elucidated. [ABSTRACT FROM PUBLISHER]

Signaling networks are essential elements that are involved in diverse cellular processes. One group of fundamental components in various signaling pathways concerns protein tyrosine kinases (PTK). Various toxicants have been demonstrated to exert their toxicity via modulation of tyrosine kinase activity. The present study aimed to identify common cellular signaling pathways that are involved in chemical-induced direct immunotoxicity. To this end, an antibody array-based profiling approach was applied to assess effects of five immunotoxicants, two immunosuppressive drugs and two non-immunotoxic control chemicals on the phosphorylation of 28 receptor tyrosine kinases and 11 crucial signaling nodes in Jurkat T-cells. The phosphorylation of ribosomal protein S6 (RPS6) and of kinases Akt, Src and p44/42 were found to be commonly regulated by immunotoxicants and/or immunosuppressive drugs (at least three compounds), with the largest effect observed upon RPS6. Flow cytometry and Western blotting were used to further examine the effect of the model immunotoxicant TBTO on the components of the mTOR-p70S6K-RPS6 pathway. These analyses revealed that both TBTO and the mTOR inhibitor rapamycin inactivate RPS6, but via different mechanisms. Finally, a comparison of the protein phosphorylation data to previously obtained transcriptome data of TBTO-treated Jurkat cells resulted in a good correlation at the pathway level and indicated that TBTO affects ribosome biogenesis and leukocyte migration. The effect of TBTO on the latter process was confirmed using a CXCL12 chemotaxis assay. [ABSTRACT FROM PUBLISHER]

Deoxynivalenol (DON) is one of the most abundant mycotoxins worldwide and mostly detected in cereals and grains. As such, DON poses a risk for many adverse health effects to human and animals. In particular, immune cells are very sensitive to DON, with the initiating step leading to toxicity being a binding to the eukaryotic 60S ribosomal subunit and induction of ribotoxic stress. The present study aimed to: (1) extend insight into the mechanism of action (MOA) of DON in immune cells; and (2) understand why immune cells are more sensitive to DON than most other cell types. Previously published microarray studies have described the effects of DON on immune cells. To build upon these findings, here, immunocytological and biochemical studies were performed using human T-lymphocyte Jurkat cells that were exposed for 3 h to 0.5 µM DON. Induction of ER stress by DON was confirmed by immunocytology demonstrating increased protein expression of two major ER stress markers ATF3 and DDIT3. T-cell activation was confirmed by induction of phosphorylation of protein kinases JNK and AKT, activation of NF- κB (p65), and increased expression of NFAT target gene NUR77; each of these are known inducers of the T-cell activation response. Induction of an oxidative stress response was also confirmed by monitoring the nuclear translocation of major oxidative stress markers NRF2 and KEAP1, as well as by changes (i.e. decreases) in cell levels of reduced glutathione. Lastly, this study showed that DON induced cleavage of caspase-3, an event known to mediate apoptosis. Taken together, these results allowed us to formulate a potential mechanism of action of DON in immune cells, i.e. binding to eukaryotic 60S ribosomal subunit → ribotoxic stress → ER stress → calcium release from the ER into cytoplasm → T-cell activation and oxidative stress → apoptosis. It is proposed that immune cells are more sensitive to DON than other cell types due to the induction of a T-cell activation response by increased intracellular calcium levels. [ABSTRACT FROM AUTHOR]

The objective of this work was to investigate whether proteomic analysis of thymoma cells treated with the trichothecene deoxynivalenol (DON) as compared to non-treated (control) cells would reveal differential protein expression, and thus would contribute to a better understanding of the mechanisms of its toxicity. For that purpose the mouse thymoma cell line EL4 was exposed to 0.5 μM DON for 6 hr. A total of 30 proteins were affected after exposure of EL4 cells to DON. Most of these proteins were up-regulated and included key metabolic enzymes (e.g., fatty acid synthase, aldose reductase, carbamoyl phosphate synthetase, glucose-6-phosphate isomerase), chaperones (e.g., HSP9AB1 and HSP70), enzymes implicated in protein folding (PDI and ERO1-lα), and proteins involved in protein degradation (ubiquitin-conjugating enzyme (E1) and proteasome subunit alpha type-1). In addition, an IgE-binding protein with a molecular weight of 60 kDa and My-binding protein 1a (MYBBP1A), a transcription factor, were found to be up-regulated by DON. The observed up-regulation of MYBBP1A, a known repressor of a number of transcription factors such as PGC-1α, C-myb, and p65 of the NF-κB family, suggests that this protein might play a role in the mechanism of DON toxicity. [ABSTRACT FROM AUTHOR]

Here, we report the results of proteomic analysis of the mouse thymoma EL4 cell line exposed to bis(tri-n-butylin)oxide (TBTO), an immunotoxic organotin compound. The objective of the work was to examine whether TBTO affects the expression of proteins in this cell line and to compare the differentially expressed proteins with the corresponding mRNA expression data. The identified proteins were quantified using a label-free quantitative method based on counting the observed peptides as an index of protein abundance. The calculation of the ratio of peptides obtained from exposed and control samples allowed us to evaluate the effect of TBTO on protein expression and to compare these results to those obtained in gene expression profiling studies. Correlation of some of the differentially expressed proteins and their corresponding mRNAs was observed. The analysis of the protein ratios revealed that 12 proteins were significantly affected. These proteins included cytoskeleton proteins myosin-9, spectrin beta 2 and plectin 8. The first two proteins were down-regulated 3-fold, whereas the third was up-regulated 2-fold. Ras-related Rab1, a GTP binding protein and T-complex protein-1 subunit alpha, a chaperonin, were decreased 2- and 3.6-fold, respectively. The ribosomal S10 and eukaryotic translation factor (eIf4G1), which are involved in protein synthesis, were down-regulated 2.6- and 3.7-fold, respectively. Also, proteins involved in splicing of pre-mRNA and in transcription, splicing factor arginine/serine-rich 2 and chromodomain-helicase-DNA binding protein 4 (Chd4), were decreased 2.6- and 4.5 times, respectively. Nuclear RNA helicase II was reduced 2.8-fold. Finally, prothymosin-alpha (ProTalpha), an essential protein for cell proliferation, and a protein similar to ProTalpha, (with a molecular weight and a pI (3.54) comparable to that of ProTalpha) were also down-regulated 6-and 8-fold, respectively. We propose that the observed down-regulation of the expression level of ProTalpha in the TBTO-exposed cells could account for the previously reported anti-proliferative effect of TBTO.

New techniques are needed to broaden the understanding of the food allergic response. The capacity of peanut extract to influence gene expression profiles was investigated in a Brown Norway rat model for food allergy. Brown Norway rats were sensitized to peanut extract (0, 1 and 10 mg/rat/d) by daily oral gavage and were dissected after 3, 7 or 14 days of exposure. RNA extracted from mesenteric lymph nodes of individual rats were hybridized against a common reference pool on Agilent whole rat genome (4*44k) arrays. The raw data were normalized and statistically analyzed using the statistical program R. A False Discovery Rate of 10% and a Fold Ratio of - 1.5 < or = Fold Ratio or Fold Ratio > or = 1.5 between the experimental groups and their respective control groups were applied. Differentially expressed genes were clustered into a heatmap. Functional annotation and GeneOntology term enrichment were examined. Furthermore, the involvement of the differentially expressed genes in specific cellular pathways was investigated with MetaCore. Gene expression changes, which were both dose- and time-dependent, were detected after sensitization to peanut. A total of 64 genes were differentially expressed, of which 60 were up-regulated and four were down-regulated. These changes were related to the regulation of immunological processes, most notably increased cell division. The findings indicate that responses to peanut include proliferation of immunologically relevant tissues, which can be identified by analysis of gene expression profiles. This may lay a basis for further research into possibilities for discrimination of allergenic from non-allergenic proteins.

In the Local Lymph Node Assay (LLNA), a stimulation index of 3 (SI = 3) is established as a threshold value for hazard identification of sensitization. The corresponding EC3 value, the effective concentration inducing a threefold increase compared to controls, can possibly predict threshold levels for sensitization in humans. Exposure to a dose below the threshold dose would not result in an induction of an immune response. Each repeated contact would be considered and viewed as a new contact and as long as the dose is below the threshold there will be no response, even after repeated exposures. However, repeated exposure may result in local accumulation eventually resulting in a dose that induces a response above the threshold for immunization. We investigated lymph node responses after short and prolonged exposure to formaldehyde donors, chemicals that are highly reactive with proteins and may thus persist in the skin. The studies were performed with formaldehyde and formaldehyde releasers (formaldehyde, paraformaldehyde, Quaternium-15, 2-Chloro-N-(hydroxymethyl)acetamide, and hexamethylenetetramine), at concentrations that induce a SI = 2, i.e., below the threshold for hazard identification. For all test chemicals investigated enhanced lymph node responses were obtained when comparing long-term prolonged exposure to short-term exposure, while three of five chemicals induced responses above SI = 3. Our results show that repeated and prolonged exposure to doses below the EC3 value can induce reactions above the SI = 3, the hazard identification threshold for sensitization in mice. So, when discussing the possible use of the EC3 as benchmark for risk assessment, one should consider duration of exposure and the possibility of local accumulation of the chemical under investigation.