Effect of diindolylmethane on Ca2+ movement and viability in HA59T human hepatoma cells

The effect of diindolylmethane, a natural compound derived from indole-3-carbinol in cruciferous vegetables, on cytosolic Ca2+ concentrations ([Ca2+](i)) and viability in HA59T human hepatoma cells is unclear. This study explored whether diindolylmethane changed [Ca2+](i) in HA59T cells. The Ca2+-sensitive fluorescent dye fura-2 was applied to measure [Ca2+](i). Diindolylmethane at concentrations of 1-50 mu M evoked a [Ca2+](i) rise in a concentration-dependent manner. The signal was reduced by removing Ca2+. Diindolylmethane-induced Ca2+ influx was not inhibited by nifedipine, econazole, SK&F96365, and protein kinase C modulators but was inhibited by aristolochic acid. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitors thapsigargin or 2,5-di-tert-butylhydroquinone (BHQ) inhibited or abolished diindolylmethane-induced [Ca2+](i) rise. Incubation with diindolylmethane inhibited thapsigargin or BHQ-induced [Ca2+](i) rise. Inhibition of phospholipase C with U73122 reduced diindolylmethane-induced [Ca2+](i) rise. At concentrations of 10-75 mu M, diindolylmethane killed cells in a concentration-dependent manner. The cytotoxic effect of diindolylmethane was not reversed by chelating cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Propidium iodide staining data suggest that diindolylmethane (25-50 mu M) induced apoptosis in a concentration-dependent manner. Collectively, in HA59T cells, diindolylmethane induced a [Ca2+](i) rise by causing phospholipase C-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ influx via phospholipase A(2)-sensitive channels. Diindolylmethane induced cell death that may involve apoptosis.