Role of Na⁺/Ca²⁺ Exchangers in Therapy Resistance of Medulloblastoma Cells

Background/Aims: Alterations of cytosolic Ca²⁺-activity ([Ca²⁺]_i) are decisive in the regulation of tumor cell proliferation, migration and survival. Transport processes participating in the regulation of [Ca²⁺]_i include Ca²⁺ extrusion through K⁺-independent (NCX) and/or K⁺-dependent (NCKX) Na⁺/Ca²⁺-exchangers. The present study thus explored whether medulloblastoma cells express Na⁺/Ca²⁺-exchangers, whether expression differs between therapy sensitive D283 and therapy resistant UW228-3 medulloblastoma cells, and whether Na⁺/Ca²⁺-exchangers participate in the regulation of cell survival. Methods: In therapy sensitive D283 and therapy resistant UW228-3 medulloblastoma cells transcript levels were estimated by RT-PCR, protein abundance by Western blotting, cytosolic Ca²⁺-activity ([Ca²⁺]_i) from Fura-2-fluorescence, Na⁺/ Ca²⁺-exchanger activity from the increase of [Ca²⁺]_i (Δ[Ca²⁺]_i) and from whole cell current (I_ca) following abrupt replacement of Na⁺ containing (130 mM) and Ca²⁺ free by Na⁺ free and Ca²⁺ containing (2 mM) extracellular perfusate as well as cell death from PI -staining and annexin-V binding in flow cytometry. Results: The transcript levels of NCX3, NCKX2, and NCKX5, protein abundance of NCX3, slope and peak of Δ[Ca²⁺]_i as well as I_ca were significantly lower in therapy sensitive D283 than in therapy resistant UW228-3 medulloblastoma cells. The Na⁺/Ca²⁺-exchanger inhibitor KB-R7943 (10 µM) significantly blunted Δ[Ca²⁺]_i, and augmented the ionizing radiation-induced apoptosis but did not significantly modify clonogenicity of medulloblastoma cells. Apoptosis was further enhanced by NCX3 silencing. Conclusions: Na⁺/Ca²⁺-exchanger activity significantly counteracts apoptosis but does not significantly affect clonogenicity after radiation of medulloblastoma cells.