Effect of methoxsalen on Ca ²⁺ homeostasis and viability in human osteosarcoma cells

Methoxsalen is a natural compound found in many seed plants. The effect of methoxsalen on Ca ²⁺ -related physiology in human osteosarcoma is unclear. This study investigated the effect of methoxsalen on cytosolic free Ca ²⁺ concentrations ([Ca ²⁺ ] _i ) in MG63 human osteosarcoma cells. Methoxsalen induced [Ca ²⁺ ] _i rises concentration-dependently. Methoxsalen-induced Ca ²⁺ entry was confirmed by Mn ²⁺ -induced quench of fura-2 fluorescence. This Ca ²⁺ entry was suppressed by nifedipine, econazole, and SKF96365. In Ca ²⁺ -free medium, incubation with the endoplasmic reticulum Ca ²⁺ pump inhibitor 2,5-di-tert-butylhydroquinone (BHQ) inhibited methoxsalen-evoked [Ca ²⁺ ] _i rises by 96%. In contrast, incubation with methoxsalen abolished BHQ-evoked [Ca ²⁺ ] _i rises. Inhibition of phospholipase C (PLC) with U73122 abolished methoxsalen-induced [Ca ²⁺ ] _i rises. Methoxsalen was cytotoxic at 300-700 μM in a concentrationdependent fashion. Chelating cytosolic Ca ²⁺ with 1,2-bis(2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid/acetoxymethyl ester (BAPTA/AM) did not prevent methoxsalen-induced cytotoxicity. Collectively, our data suggest that in MG63 cells, methoxsalen induced [Ca ²⁺ ] _i rises by evoking PLC-dependent Ca ²⁺ release from the endoplasmic reticulum, and Ca ²⁺ entry via store-operated Ca ²⁺ entry. Methoxsalen also induced Ca ²⁺ -disassociated cell death.