γ-Irradiation induces P2X 7 receptor-dependent ATP release from B16 melanoma cells

Abstract

Background Ionizing irradiation causes not only growth arrest and cell death, but also release of growth factors or signal transmitters, which promote cancer malignancy. Extracellular ATP controls cancer growth through activation of purinoceptors. However, there is no report of radiation-induced ATP release from cancer cells. Here, we examined γ-irradiation-induced ATP release and its mechanism in B16 melanoma. Methods Extracellular ATP was measured by luciferin–luciferase assay. To investigate mechanism of radiation-induced ATP release, we pharmacologically inhibited the ATP release and established stable P2X 7 receptor-knockdown B16 melanoma cells using two short hairpin RNAs targeting P2X 7 receptor. Results Cells were exposed to 0.5–8 Gy of γ-rays. Extracellular ATP was increased, peaking at 5 min after 0.5 Gy irradiation. A selective P2X 7 receptor channel antagonist, but not anion transporter inhibitors, blocked the release of ATP. Further, radiation-induced ATP release was significantly decreased in P2X 7 receptor-knockdown cells. Our results indicate that γ-irradiation evokes ATP release from melanoma cells, and P2X 7 receptor channel plays a significant role in mediating the ATP release. General Significance We suggest that extracellular ATP could be a novel intercellular signaling molecule released from cancer cells when cells are exposed to ionizing radiation.