Immune cell profiles associated with measured exposure to phthalates in the Norwegian EuroMix biomonitoring study – A mass cytometry approach in toxicology

Unni C Nygaard,Emilie S Ulriksen,Hege Hjertholm,Friederike Sonnet,Anette K Bølling,Monica Andreassen,Trine Husøy,Hubert Dirven

doi:10.1016/j.envint.2020.106283

Abstract

BackgroundPhthalate exposure has been associated with immune-related diseases such as asthma and allergies, but there is limited knowledge on mechanisms, effect biomarkers and thus biological support of causality. ObjectivesTo investigate associations between exposure to the phthalates DEHP (di(2-ethylhexyl) phthalate) and DiNP (diisononyl phthalate) and functional immune cell profiles. MethodsPeripheral blood mononuclear cells (PBMCs) from 32 healthy adult Norwegian participants in the EuroMix biomonitoring study were selected based on high or low (n = 16) levels of urine metabolites of DEHP and DiNP. High-dimensional immune cell profiling including phenotyping and functional markers was performed by mass cytometry (CyTOF) using two broad antibody panels after PMA/ionomycin-stimulation. The CITRUS algorithm with unsupervised clustering was used to identify group differences in cell subsets and expression of functional markers, verified by manual gating. ResultsThe group of participants with high phthalate exposure had a higher proportion of some particular innate immune cells, including CD11c positive NK-cell and intermediate monocyte subpopulations. The percentage of IFNγ TNFα double positive NK cells and CD11b expression in other NK cell subsets were higher in the high exposure group. Among adaptive immune cells, however, the percentage of IL-6 and TNFα expressing naïve B cell subpopulations and the percentage of particular naïve cytotoxic T cell populations were lower in the high exposure group. DiscussionCell subset percentages and expression of functional markers suggest that DEHP and DiNP phthalate exposure may stimulate subsets of innate immune cells and suppress adaptive immune cell subsets. By revealing significant immunological differences even in small groups, this study illustrates the promise of the broad and deep information obtained by high-dimensional single cell analyses of human samples to answer toxicological questions regarding health effects of environmental exposures.

Highlights

The immune system is a fine-tuned and complex system, and dysre gulation plays a fundamental role in a variety of diseases and conditions, including several non-communicable diseases (NCDs; (Davis et al, 2017))
Immune cell profiles associated with phthalate exposure levels in unstimulated cells
For the adaptive immune cells, the % of particular subpopulations were lower in the high di(2-ethylhexyl) phthalate (DEHP) and diisononyl phthalate (DiNP) exposure group, including IL-6 TNFα double positive B cell sub populations and less consistently various naïve cytotoxic and helper T cell populations

Summary

Introduction

The immune system is a fine-tuned and complex system, and dysre gulation plays a fundamental role in a variety of diseases and conditions, including several non-communicable diseases (NCDs; (Davis et al, 2017)). Identification of causal relationships as well as appropriate biological assays and biomarkers of effect to assess adverse effects on the immune system is in high demand Such knowledge is required to motivate development and implementation of effective policies for reduced chemical exposure, and a subsequent reversion of the present increases in disease incidence and socioeconomic costs (EFSA CEF Panel (EFSA Panel on Food Contact Materials et al, 2019). Phthalates are a group of chemicals commonly used as plasticizers, lubricants, and solvents in a variety of consumer products, such as food contact materials, building materials, cosmetics and toys (Luo et al, 2018; Wittassek et al, 2011). Discussion: Cell subset percentages and expression of functional markers suggest that DEHP and DiNP phthalate exposure may stimulate subsets of innate immune cells and suppress adaptive immune cell subsets. By revealing significant immunological differences even in small groups, this study illustrates the promise of the broad and deep information obtained by high-dimensional single cell analyses of human samples to answer toxicological questions regarding health effects of environmental exposures

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