Inhaled BTEX Produces Pro-Inflammatory Cytokine Release in the Lung After Acute Exposure
On this page:
Background and Purpose:
Volatile organic compounds (VOCs) are ubiquitous environmental pollutants associated with adverse health effects on respiratory, immune, cardiovascular, and nervous systems. BTEX (benzene, toluene, ethylbenzene, and xylenes) are common VOCs emitted from solvents in paints and coatings, fires, gasoline, jet fuels, and kerosene. BTEX chemicals occur frequently in hazardous waste sites and significant concentrations are measured in ambient air, water, and soil. Atmospheric photochemical aging may alter composition and toxicity of BTEX and other VOCs. To understand the role of photochemical aging in BTEX toxicity, we first examined dose-response relationships of freshly emitted BTEX and acute inflammation, respiratory function, and genomic endpoints in mice.
Methods:
Mixtures of BTEX with proportions representative of environmental concentrations were generated (42% toluene, 34% xylenes, 18% benzene, 6% ethylbenzene), and 11-week old female Balb/cJ mice were exposed nose-only for 4 hours to clean air or target concentrations of 10, 31.6, 100, or 316 ppm total BTEX (n = 8/group). Real-time respiratory responses were measured throughout exposure (emka Scireq head-out plethysmography). Two hours after exposure, mice were euthanized, and blood was taken for hematological analysis. Right lung lobes were lavaged with HBSS, and bronchoalveolar lavage fluid (BALF) was analyzed for total cell count, cell differentials, biochemistry (protein, albumin, lactate dehydrogenase, g-glutamyl transpeptidase, N-acetyl glucosaminidase), and cytokines (macrophage inflammatory protein (MIP)-2, interleukin (IL)-6, tumor necrosis factor (TNF)-a). Samples of lateral and septal nasal epithelium and lung were taken for RNA sequencing using BioSpyder TempO-Seq whole transcriptome analysis.
Results:
Measured total BTEX exposure concentrations were 8.3, 30.6, 107.1, and 331.1 ppm (respectively 83%, 97%, 107%, and 105% of target concentrations); within these totals, the 4 chemical concentrations deviated from their targets by an average of 0.55 ppm (at 10 ppm total BTEX target), 0.78 ppm (at 31.6 ppm total), 2.00 ppm (at 100 ppm total), and 6.02 ppm (at 316 ppm total). There were no effects of BTEX on measures of pulmonary function during exposure in comparison with pre-exposure clean air baseline values. Two hours post-exposure, there were no significant effects of BTEX on complete blood counts, cell volumes, plateletcrit, or hematocrit. BTEX concentrations of 8.3, 30.6, 107.1, and 331.1 ppm all produced highly significant, but not dose-related, increases in BAL pro-inflammatory cytokines (MIP-2 respectively 5x, 7x, 5x, 7x air control of 27 ± 4 pg/mL; IL-6 respectively 80x, 98x, 66x, 114x air control of 1.4 ± 0.1 pg/mL; no change in TNF-a). There were no significant effects on BALF cell numbers or biochemistry. Whole transcriptome analysis reveals differentially expressed genes and altered pathways following BTEX exposure.
Conclusions:
We found that a single 4-hour exposure to environmentally representative proportions of BTEX chemicals induced highly significant releases of pro-inflammatory cytokines at all concentrations tested including the lowest measured level (8.3 ppm), but no other changes in respiratory responses or measures of toxicity 2 hours post-exposure. While the lowest concentration tested in this acute exposure is greater than most reported high occupational levels (0.1 – 1.0 ppm total BTEX), the consistency of the cytokine effect was striking. Chronic exposures at lower concentrations may elicit different toxicological pathways. Comparison of these responses with those from exposures to photochemically-aged BTEX mixtures will assist in understanding the effects of climate change and atmospheric aging on relative toxicity of VOC mixtures such as BTEX. (This abstract does not represent U.S. EPA policy)