Toxicological Sciences, Vol 48, 206-217, Copyright © 1999 by Society of Toxicology
TS Manetz and BJ Meade
These studies were conducted to investigate the potential use of a flow
cytometric analysis method for the identification and differentiation of
chemicals with the capacity to induce irritation, IgE- or T cell- mediated
hypersensitivity responses. An initial study investigated the ability of
equally sensitizing concentrations (determined by local lymph node assay)
of IgE-mediated (Toluene Diisocyanate-TDI) and T cell- mediated
(Dinitrofluorobenzene-DNFB) allergens to differentially modulate the
IgE+B220+ population in the lymph nodes draining the dermal exposure site.
Sodium lauryl sulfate (SLS) was also tested as a nonsensitizing irritant
control. Female B6C3F1 mice were dermally exposed once daily for 4
consecutive days, with the optimum time point for analysis determined by
examining the IgE+B220+ population 8, 10, and 12 days post-initial chemical
exposure. At the peak time point, day 10, the IgE+B220+ population was
significantly elevated in TDI (41%), while moderately elevated in DNFB
(18%) exposed animals when compared to the vehicle (0.8%), and remained
unchanged in SLS (2.2%) exposed animals when compared to the ethanol
control (2.5%). Experiments in our laboratory and others have demonstrated
that the draining lymph node B220+ population becomes significantly
elevated following exposure to allergens (IgE- and T cell-mediated), not
irritants, allowing for their differentiation. An existing mouse ear
swelling assay was used to identify chemical irritants. Therefore, using
the endpoints of percent ear swelling, percent B220+ cells, and percent
IgE+B220+ cells, a combined irritancy/phenotypic analysis assay was
developed and tested with tetradecane (irritant), toluene diisocyanate,
trimellitic anhydride (IgE-mediated allergens), benzalkonium chloride,
dinitrofluorobenzene, oxazolone, and dinitrochlorobenzene (T cell- mediated
allergens) over a range of concentrations. Based upon the pattern of
response observed, a paradigm was developed for continued evaluation:
Irritant exposure will result in significant ear swelling without altering
the B220+ or IgE+B220+ populations. Exposure to sensitizers (IgE-mediated
or T cell-mediated) will increase the B220+ population and the percent ear
swelling will remain unchanged or will significantly increase, depending on
the irritancy capacity of the chemical. Both the IgE+B220+ and B220+
populations will become elevated at the same test concentration following
exposure to IgE-mediated, hypersensitivity inducing allergens. At its peak,
the percent of IgE+B220+ cells will be equal to the percent of B220+ cells.
The B220+ population will increase at a lower test concentration than the
IgE+B220+ population, following exposure to T cell-mediated,
hypersensitivity inducing allergens. At its peak, the percent of IgE+B220+
cells will reach less than half that of the percent of B220+ cells. The
irritancy/phenotypic analysis method may represent a single murine assay
able to identify and differentiate chemicals with the capacity to induce
irritation, or IgE-mediated or T cell-mediated responses.
ARTICLES
Development of a flow cytometry assay for the identification and differentiation of chemicals with the potential to elicit irritation, IgE-mediated, or T cell-mediated hypersensitivity responses
Department of Pharmacology and Toxicology, Medical College of Virginia/VCU, Richmond, USA.
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