The effect of fendiline on cytosolic free Ca2+ concentrations ([Ca2+]i) and proliferation has not been explored in human oral cancer cells. This study examined whether fendiline altered Ca2+ levels and caused cell death in OC2 human oral cancer cells. [Ca2+]i and cell viability were measured using the fluorescent dyes fura-2 and WST-1, respectively. Fendiline at concentrations above 10 μM increased [Ca2+]i in a concentration-dependent manner. The Ca2+ signal was reduced partly by removing extracellular Ca2+. The fendiline-induced Ca2+ influx was sensitive to blockade of L-type Ca2+ channel blockers. In Ca2+-free medium, after pretreatment with 50 μM fendiline, 1 μM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor)-induced [Ca2+]i rises were inhibited; and conversely, thapsigargin pretreatment nearly abolished fendiline-induced [Ca2+]i rises. Inhibition of phospholipase C with 2 μM U73122 did not change fendiline-induced [Ca2+]i rises. At concentrations between 5 and 25 μM, fendiline killed cells in a concentration-dependent manner. The cytotoxic effect of 15 μM fendiline was not reversed by prechelating cytosolic Ca2+ with BAPTA/AM. Collectively, in OC2 cells, fendiline induced [Ca2+]i rises by causing Ca2+ release from the endoplasmic reticulum and Ca2+ influx from L-type Ca2+ channels. Furthermore, fendiline-caused cytotoxicity was not via a preceding [Ca2+]i rise.

Resveratrol is a natural compound that affects cellular calcium (Ca2+) homeostasis and viability in different cells. This study examined the effect of resveratrol on cytosolic free Ca2+ concentrations ([Ca2+]i) and viability in OC2 human oral cancer cells. The Ca2+-sensitive fluorescent dye fura-2 was used to measure [Ca2+]i, and water-soluble tetrazolium-1 was used to measure viability. Resveratrol evoked concentration-dependent increase in [Ca2+]i. The response was reduced by removing extracellular Ca2+. Resveratrol also caused manganese-induced fura-2 fluorescence quench. Resveratrol-evoked Ca2+ entry was inhibited by nifedipine and the protein kinase C (PKC) inhibitor GF109203X but was not altered by econazole, SKF96365, and the PKC activator phorbol 12-myristate 13 acetate. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor 2,5-di- tert-butylhydroquinone (BHQ) abolished resveratrol-evoked [Ca2+]i rise. Conversely, treatment with resveratrol inhibited BHQ-evoked [Ca2+]i rise. Inhibition of phospholipase C (PLC) with U73122 abolished resveratrol-evoked [Ca2+]i rise. At 20–100 μM, resveratrol decreased cell viability, which was not affected by chelating cytosolic Ca2+with 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid-acetoxymethyl ester. Annexin V-fluorescein isothiocyanate staining data suggest that resveratrol at 20–40 μM induced apoptosis in a concentration-dependent manner. Collectively, in OC2 cells, resveratrol induced [Ca2+]i rise by evoking PLC-dependent Ca2+ release from the endoplasmic reticulum and by causing Ca2+ entry via nifedipine-sensitive, PKC-regulated mechanisms. Resveratrol also caused Ca2+-independent apoptosis.

Cyfluthrin is a pyrethroid insecticide and common household pesticide. The effect of cyfluthrin on Ca2+-related physiology in human osteosarcoma is unclear. This study investigated the effect of cyfluthrin on cytosolic-free Ca2+ concentrations ([Ca2+]i) and viability in MG63 human osteosarcoma cells. Cyfluthrin concentration-dependently induced [Ca2+]i rises. Cyfluthrin-induced Ca2+ entry was confirmed by the Mn2+-induced quench of fura-2 fluorescence. Cyfluthrin at concentrations of 10–100 μM induced [Ca2+]i rises. Ca2+ removal reduced the signal by approximately 50%. Cyfluthrin (100 μM) induced Mn2+ influx suggesting Ca2+ entry. Cyfluthrin-induced Ca2+ entry was inhibited 50% by protein kinase C (PKC) activator (phorbol 12-myristate 13-acetate) and inhibitor (GF109203X) and also by three inhibitors of store-operated Ca2+ channels: nifedipine, econazole, and SKF96365. In Ca2+-free medium, treatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin (TG) completely inhibited cyfluthrin-evoked [Ca2+]i rises. Conversely, treatment with cyfluthrin abolished TG-evoked [Ca2+]i rises. Inhibition of phospholipase C (PLC) with 1-[6-[((17β)-3-methoxyestra-1,3,5[10]-trien-17-yl)amino]hexyl]-1H-pyrrole-2,5-dion abolished cyfluthrin-induced [Ca2+]i rises. Cyfluthrin at 25–65 μM decreased cell viability, which was not reversed by pretreatment with the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid–acetoxymethyl ester. Together, in MG63 cells, cyfluthrin induced [Ca2+]i rises by evoking PLC-dependent Ca2+ release from the endoplasmic reticulum and Ca2+ entry via PKC-sensitive store-operated Ca2+ entry. Cyfluthrin also caused Ca2+-independent cell death.

The effects of the environmental toxicant, triethyltin, on Ca2 + mobilization in Madin–Darby canine kidney (MDCK) cells have been examined. Triethyltin induced an increase in cytosolic free Ca2 + levels ([Ca2 +]i) at concentrations larger than 2 mM in a concentrationdependent manner. Within 5 min, the [Ca2 +]i signal was composed of a gradual rise and a sustained phase. The [Ca2 +]i signal was partly reduced by removing extracellular Ca2 +. In Ca2 +-free medium, pretreatment with thapsigargin (1 mM), an endoplasmic reticulum Ca2 + pump inhibitor, reduced 50 mM triethyltin-induced [Ca2 +]i increase by 80%. Conversely, pretreatment with triethyltin abolished thapsigargin-induced Ca2 + release. Pretreatment with U73122 (2 mM) to inhibit phospholipase C-coupled inositol 1,4,5-trisphosphate formations failed to alter 50 mM triethyltin-induced Ca2 + release. Incubation with triethyltin at a concentration (1 mM) that did not increase basal [Ca2 +]i for 3 min did not alter ATP (10 mM)and bradykinin (1 mM)-induced [Ca2 +]i increases. Collectively, this study shows that triethyltin altered Ca2 + movement in renal tubular cells by releasing Ca2 + from multiple stores in an inositol 1,4,5-trisphosphate-independent manner, and by inducing Ca2 + influx.

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.