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USE OF THE RIBONUCLEASE PROTECTION ASSAY FOR IDENTIFYING CHEMICALS WHICH ELLICIT HYPERSENSITIVITY RESPONSES
Citation:
Plitnick, L. M., D M. Sailstad, AND R J. Smialowicz. USE OF THE RIBONUCLEASE PROTECTION ASSAY FOR IDENTIFYING CHEMICALS WHICH ELLICIT HYPERSENSITIVITY RESPONSES. Presented at SOT, San Francisco, CA, March 25 - 29, 2001.
Description:
Use of the Ribonuclease Protection Assay (RPA) for Identifying Chemicals that Elicit Hypersensitivity Responses. L.M. Plitnick, 1, D.M. Sailstad, 2, and R.J. Smialowicz, 2 1UNC, Curriculum in Toxicology, Chapel Hill, NC and 2USEPA, NHEERL, RTP, NC.
The incidence of allergy and asthma has increased dramatically over the last 20 years and there is concern that exposure to chemicals such as diisocyanates and acid anhydrides in both domestic and occupational settings may contribute to this rise. Methods are needed to efficiently screen chemicals for the potential to cause airway hypersensitivity (AHS) reactions. AHS is driven by Th2 cells, whereas the contact response is driven by Th1 cells and the distinct cytokine profiles produced by these cells provide a means of distinguishing respiratory from contact sensitizers. ELISA assays have previously been conducted for this purpose but we have used the more efficient RPA to examine cytokine profiles. In these studies, mice were topically exposed to the known airway sensitizer trimellitic anhydride (TMA), and the contact sensitizers dinitrofluorobenzene (DNFB) and dinitrochlorobenzene (DNCB) on days 0 and 5 and challenged on days 10-12. At various times following challenge, total mRNA was isolated from draining lymph nodes and analyzed by RPA. Cytokines produced by Th2 cells (IL4, IL10 and IL13) were significantly increased in response to TMA as compared to DNCB and DNFB. Levels of the Th1 cytokine IFNg were significantly elevated over TMA in DNCB and DNFB treated mice however, this occurred at only one point during the timecourse. A dose response experiment confirmed these results and showed that TMA induces higher levels of IL4, IL10 and IL13 than DNCB and DNFB over a range of doses. The RPA appears to be an effective method for the detection of cytokines characteristic of AHS responses following dermal sensitization and elicitation with a respiratory sensitizer which is more efficient than traditional ELISA methods in that multiple cytokines can be assessed at one time and an additional in vitro stimulation step is not required. (This abstract does not reflect EPA policy. This work was supported in part by The Dow Chemical Co. & DuPont Co.)