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Updated: Jun 8, 2026
Measurement of T Cell Alloreactivity Using Imaging Flow Cytometry
Published on: April 19, 2017
Flow cytometry imaging identifies rare T(H)2 cells expressing thymic stromal lymphopoietin receptor in a
Amanda J Reefer1, Kathryn E Hulse, Josephine A Lannigan
1Asthma and Allergic Diseases Center, University of Virginia Health System, Charlottesville, VA 22908-1355, USA.
Background:
Thymic stromal lymphopoietin (TSLP) is expressed at sites of allergic inflammation, including eczematous skin. This cytokine has been reported to exert its T(H)2-inducing properties through dendritic cells. Expression of TSLP receptor on the surface of activated T(H)2 cells could amplify T(H)2 responses at inflamed sites through the direct actions of TSLP.
Objective:
To test rigorously whether T(H)2 cells induced by "proallergic" factors express TSLP receptor and characterize these cells using an experimental platform that combines flow cytometry with microscopic capabilities.
Methods:
CD4(+) T cells isolated from patients with atopic dermatitis or normal healthy controls were cocultured with autologous dendritic cells in the presence of T(H)2-promoting stimuli (TSLP ± allergen and staphylococcal enterotoxin B ± TSLP). Surface expression of TSLP receptor was analyzed by image-based flow cytometry, and responsiveness of purified T cells to TSLP was assessed by phosphorylation of signal transducer and activator of transcription-5 and cytokine secretion.
Results:
T(H)2-promoting stimuli induced a robust population of activated T(H)2 cells (CD25(+)IL-4(+)). Regardless of the nature of the stimulus, flow cytometry imaging confirmed that T cells expressing TSLP receptor were rare, constituting a minor fraction of the IL-4(+) T cell pool; however, TSLP responsiveness was nonetheless detectable. Analysis of cell size and nuclear morphology revealed preferential expression of TSLP receptor on IL-4-expressing cells undergoing mitosis. Analysis of lesional skin in atopic dermatitis supported the view that rare IL-4(+) T cells expressing TSLP receptor are present at inflamed sites.
Conclusion:
In a "proallergic" milieu, TSLP receptor is preferentially expressed on rare actively dividing T(H)2 cells. The direct action of TSLP on T cells could amplify T(H)2 responses at sites of allergic inflammation.
