Induction of Ca2+ signaling and cytotoxic responses of human lung fibroblasts upon an antihistamine drug oxatomide treatment and evaluating the protective effects of Ca2+ chelating.

Background: Oxatomide, an antihistamine drug of the diphenylmethylpiperazine family, has anti-inflammatory effects in airway disease. Because oxatomide was shown to cause diverse physiological responses in several cell models, the impact of oxatomide on Ca²⁺ signaling and its related physiological effects has not been explored in IMR-90 human fetal lung fibroblasts.

Objectives: This study assessed the effect of oxatomide on cell viability and intracellular free Ca²⁺ concentrations ([Ca²⁺]_i) and examined whether oxatomide-induced cytotoxicity through Ca²⁺ signaling in IMR-90 cells.

Methods: Cell viability was measured by the cell proliferation reagent (WST-1). [Ca²⁺]_i was measured by the Ca²⁺-sensitive fluorescent dye fura-2.

Results: Oxatomide (10-40 μM) concentration dependently reduced cell viability and induced [Ca²⁺]_i rises in IMR-90 cells. This cytotoxic effect was reversed by chelation of cytosolic Ca²⁺ with BAPTA-AM. In terms of Ca²⁺ signaling, oxatomide-caused Ca²⁺ entry was inhibited by modulators of store-operated Ca²⁺ channels (2-APB and SKF96365) and protein kinase C (PKC) inhibitor (GF109203X). Furthermore, oxatomide-induced Ca²⁺ influx was confirmed by Mn²⁺-induced quench of fura-2 fluorescence. In a Ca²⁺-free medium, preincubation with the endoplasmic reticulum Ca²⁺ pump inhibitor thapsigargin inhibited oxatomide-evoked [Ca²⁺]_i rises. Conversely, treatment with oxatomide abolished thapsigargin-induced [Ca²⁺]_i rises. Inhibition of phospholipase C (PLC) with U73122 also inhibited oxatomide-caused [Ca²⁺]_i rises.

Conclusion: In IMR-90 cells, oxatomide-induced cytotoxicity by preceding [Ca²⁺]_i rises involving PKC-sensitive store-operated Ca²⁺ entry and PLC-dependent Ca²⁺ release from the endoplasmic reticulum. BAPTA-AM, with its Ca²⁺ chelating effects, may be a potential compound for preventing oxatomide-induced cytotoxicity.