Differential effect of residual oil fly ash (ROFA), ambient particulate matter and diesel exhaust particle (DEP) on the human pulmonary alveolar epithelium

Abstract

Epidemiological studies have shown that exposure to particulate matter (PM) contributes to cardiopulmonary morbidity and mortality. Evidence suggests that the diesel exhaust in ambient air in major cities mediates a significant proportion of the adverse health effects. However, it has been speculated that these effects might be increased in cities within the oil and gas producing countries as a result of a combined effect of inhaling both diesel exhaust particles (DEP) and residual oil fly ash (ROFA) It is hypothesised that the nature and degree of alveolar epithelial reactivity will depend on the physico-chemistry of the particles, and that DEP/ROFA mixture will result in greater bio-reactivity compared to the same amount of DEP and ROFA alone. The specific aim is to determine the physico-chemical characteristics of ambient PM, DEP, ROFA and DEP/ROFA mixture and to compare their cellular reactivity/effects on human alveolar epithelium in vitro. The physico-chemical composition of the particles was determined using transmission electron microscope and energy dispersive x-ray spectrometer (TEM/EDX) analysis. Primary human alveolar type 2 epithelial cells (AT2) and a transformed human alveolar type 1 (TT1) epithelial cell line were exposed to ambient particles, ROFA, DEP and a ROFA/ DEP (1:1) mixture. Reactive oxygen species (ROS) production and cell viability were evaluated using the dichlorofluorescin (DCFDA) and 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium (MTT) assay respectively. Enzyme-linked immunosorbent assay (ELISA) was used to quantify release of CXCL8, IL-6, and CCL2. TEM/EDX results revealed that transition metals and other reactive elements were present in the samples. The result also showed that ambient PM has an amorphous structure which made particle size difficult to determine. The estimated average diameters of ROFA and DEP were 74 ±17nm and 65 ± 2nm respectively. When mixed, it was 76 ± 24nm. Morphologically, these particles have large spherical, agglomerate and crystalline structures. Cellular exposures to PM were associated with significant increases in cytokine and chemokine release compared to non-treated controls (P< 0.002). At high concentrations (50 and 100μg/ml) the ROFA/DEP mixtures caused a significant increase in IL-6, CXCL8, and CCL2 release by TT-1 cells compared to exposure to an equivalent amount of DEP or ROFA alone (P<0.05). Ambient PM induced a higher level of mediator release compared to those observed with DEP while the DEP/ROFA mixture reduced cell viability and also triggerd rapid intracellular ROS release greater than that observed with ambient PM, DEP or ROFA alone. All these changes are concentration- and exposure time-dependent. However, the results are not statistically significant. In conclusion, DEP/ROFA mixture resulted in significant changes in cytokines/chemokines induction, ROS release and reduction in cell viability compared to DEP and ROFA alone (as well as PM). There was very little difference in particle size and shape, suggesting that differences between the reactivity of the test materials could be related to surface chemistry, for example elemental composition, although this could not be deduced in the current study. Other organic components might also be important, although they were not analysed in this investigation. Consequently, future work is needed to unravel the specific role of transition metals, as well as the organic and other inorganic components of the particles, in their cellular reactivity. It is also possible that mixing of ROFA and DEP causes reactions between volatile chemical components that enhances their cellular reactivity. These unique findings are of significance to those who live or work in cities where there are likely to be high levels of both types of emissions, which might have significant health effects.