Effect of diindolylmethane on Ca2+ homeostasis and viability in MDCK renal tubular cells

Abstract

The effect of the natural product diindolylmethane (DIM) on cytosolic Ca(2+) concentrations ([Ca(2+)]i) and viability in MDCK renal tubular cells was explored. The Ca(2+)-sensitive fluorescent dye fura-2 was applied to measure [Ca(2+)]i. DIM at concentrations 1-50 μM induced a [Ca(2+)]i rise in a concentration-dependent manner. The response was reduced partly by removing Ca(2+). DIM induced Mn(2+) influx leading to quenching of fura-2 fluorescence. DIM-evoked Ca(2+) entry was suppressed by nifedipine, econazole, SK&F96365 and protein kinase C modulators. In the absence of extracellular Ca(2+), incubation with the endoplasmic reticulum Ca(2+) pump inhibitor thapsigargin (TG) or 2,5-di-tert-butylhydroquinone (BHQ) greatly inhibited DIM-induced [Ca(2+)]i rise. Incubation with DIM abolished TG or BHQ-induced [Ca(2+)]i rise. Inhibition of phospholipase C with U73122 reduced DIM-induced [Ca(2+)]i rise by 50%. At 1, 10, 40 and 50 μM, DIM slightly enhanced cell proliferation. The effect of 50 μM DIM was reversed by chelating cytosolic Ca(2+) with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. In sum, in MDCK cells, DIM induced a [Ca(2+)]i rise by evoking phospholipase C-dependent Ca(2+) release from the endoplasmic reticulum and Ca(2+) entry via protein kinase C-sensitive store-operated Ca(2+) channels. DIM did not induce cell death.