4.

Blockade of store-operated Ca²⁺ channels inhibiting osteosarcoma cell growth is associated with down-regulation of nuclear factor of activated T-cells cytoplasmic 1 (NFATc1). (A) Representative time course recording of store-operated calcium entry (SOCE) in the presence of SKF96365 in osteosarcoma cells. Cells were treated with cyclopiazonic acid (CPA) to passively deplete the intracellular Ca²⁺ stores followed by application of 2 mmol/L Ca²⁺; (B) Quantification of CPA-induced SOCE peak value (delta R: Ratio_max−Ratio_base; N=45 cells for each group); (C) NFATc1 luciferase activity. 143B and U2OS cells transfected with reporter plasmids were treated with SKF96365 20 μmol/L, cyclosporine A (CsA) 10 μmol/L or VIVIT 5 μmol/L; (D) Autotaxin (ATX) protein levels in osteosarcoma cells. Cytoplasmic extracts from cells treated with SKF96365 (20 μmol/L) for 24 h were analyzed by Western blot hybridization using anti-ATX and anti-GAPDH antibodies; (E) Quantitation of protein levels by densitometry. Data are presented as from three independent experiments. *, P<0.05 (vs. Control).