Abstract
Purpose: :
To investigate the effect of levocabastine, an H1 receptor antagonist currently marketed for treatment of ocular allergy, on cytokine release from eosinophils using high-throughput multiplex analysis.
Methods: :
EoL-1 (human eosinophilic leukaemia cell line) cells were used. Each experiment was performed in triplicate and carried out in parallel with both PMA-differentiated (25 ng/mL) and non-differentiated eosinophils. Cells were suspended in low-serum medium. TNF-α was used to induce cytokine release. To evaluate the relationship between time and concentration in TNF-α stimulation and cytokine release, 5, 10, or 25 ng/mL of TNF-α was administered at 0, 0.5, 1, 2, 3, 6, 12, or 24 h, and cytokine analysis was performed. Subsequently, the effect of levocabastine on TNF-α cytokine release was evaluated by exposing differentiated and non-differentiated EoL-1 cells to levocabastine (0.1 to 2.3 mM), and the content of several different cytokines was evaluated at 12 and 24 h. Samples were analyzed using Luminex 200TM (Luminex, Austin, TX) and Beadview software v1.0 (Upstate Cell Signaling Solutions, Temecula, CA). Statistical analysis was performed by ANOVA and Newman-Keuls post-hoc test using GraphPad (San Diego, CA).
Results: :
Levocabastine treatment resulted in a significantly dose-dependent decrease in TNF-α-induced release of IL-1β, IL-1ra, IL-6, IL-8, IL-12p40, IP-10, and VEGF. On the other hand, levocabastine did not inhibit the release of IL-1α, IL-5, IL-10, GM-CSF, G-CSF, MCP-1, MIP-1α, MIP-1β, RANTES, TGFα, or fractalkine.
Conclusions: :
This study demonstrates that levocabastine exerts beneficial cellular effects beyond H1 receptor antagonism, suggesting that the clinical benefit of levocabastine may extend to modulation of pro-inflammatory mediators. Therefore, it is possible that levocabastine may provide benefit to patients with allergic conjunctivitis by inhibiting pro-inflammatory cytokine secretion; this hypothesis will require further clinical interrogation for confirmation.
Keywords: cytokines/chemokines • inflammation • flow cytometry