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

Abstract 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 Ca 2+ 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 Ca 2+ concentrations ([Ca 2+ ] i ) and examined whether oxatomide‐induced cytotoxicity through Ca 2+ signaling in IMR‐90 cells. Methods Cell viability was measured by the cell proliferation reagent (WST‐1). [Ca 2+ ] i was measured by the Ca 2+ ‐sensitive fluorescent dye fura‐2. Results Oxatomide (10–40 μM) concentration dependently reduced cell viability and induced [Ca 2+ ] i rises in IMR‐90 cells. This cytotoxic effect was reversed by chelation of cytosolic Ca 2+ with BAPTA‐AM. In terms of Ca 2+ signaling, oxatomide‐caused Ca 2+ entry was inhibited by modulators of store‐operated Ca 2+ channels (2‐APB and SKF96365) and protein kinase C (PKC) inhibitor (GF109203X). Furthermore, oxatomide‐induced Ca 2+ influx was confirmed by Mn 2+ ‐induced quench of fura‐2 fluorescence. In a Ca 2+ ‐free medium, preincubation with the endoplasmic reticulum Ca 2+ pump inhibitor thapsigargin inhibited oxatomide‐evoked [Ca 2+ ] i rises. Conversely, treatment with oxatomide abolished thapsigargin‐induced [Ca 2+ ] i rises. Inhibition of phospholipase C (PLC) with U73122 also inhibited oxatomide‐caused [Ca 2+ ] i rises. Conclusion In IMR‐90 cells, oxatomide‐induced cytotoxicity by preceding [Ca 2+ ] i rises involving PKC‐sensitive store‐operated Ca 2+ entry and PLC‐dependent Ca 2+ release from the endoplasmic reticulum. BAPTA‐AM, with its Ca 2+ chelating effects, may be a potential compound for preventing oxatomide‐induced cytotoxicity.