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A NOVEL TECHNIQUE FOR QUANTITATIVE ESTIMATION OF UPTAKE OF DIESEL EXHAUST PARTICLES BY LUNG CELLS
Citation:
SAXENA, R. K., M. D. HAYS, AND M. I. GILMOUR. A NOVEL TECHNIQUE FOR QUANTITATIVE ESTIMATION OF UPTAKE OF DIESEL EXHAUST PARTICLES BY LUNG CELLS . Presented at Society of Toxicology Annual Meeting, Charlotte, NC, March 25 - 29, 2007.
Description:
While airborne particulates like diesel exhaust particulates (DEP) exert significant toxicological effects on lungs, quantitative estimation of accumulation of DEP inside lung cells has not been reported due to a lack of an accurate and quantitative technique for this purpose. In the present study, we developed a sensitive and reliable new method for quantitative estimation of uptake of DEP by lung epithelial cells and macrophages in culture. The principle of the method is to first isolate the DEP exposed cells by trypsinization and to separate them from free extracellular diesel exhaust particles by Ficoll density gradient centrifugation. Ficoll purified cells were dissolved in 1% sodium dodecyl sulfate solution and insoluble ingested DEP isolated by centrifugation. DEP pellets were subjected to an analysis on a carbon analyzer that could accurately measure the amount of elemental carbon in the sample. Since the DEP preparation used in our studies is about 90% elemental carbon, and normal cells do not have any elemental carbon, the amount of DEP taken up by the cells could be accurately estimated by this method. Using this technique, we studied the time course and dose dependence of uptake of DEP by lung epithelial cells and alveolar macrophages. Qualitative differences were found in the uptake of DEP and ultra fine carbon black (UFCB) by LA4 epithelial cells and MHS alveolar macrophages. Macrophages could ingest DEP as well as UFCB with equal efficiency, whereas epithelial cells were significantly more efficient in taking up DEP as compared to UFCB. This technique can now be used to estimate the deposition and clearance of inhale DEP and provide correlations between particle load and toxicity. This abstract does not reflect EPA policy.