influx. The distribution of P2X (P2X1-7) and P2Y (P2Y1, P2Y2, P2Y4, P2Y6, P2Y11) receptors was different between lung cancer cells and normal cells. Proapoptotic P2X7 was nearly undetectable in lung cancer cells, which may explain why lung cancer cells showed decreased cytotoxicity when treated with high concentration of ATP. The Bcl-2/Bax ratio was increased in lung cancer cells following treatment with ATP; however, the antiapoptotic protein Bcl-2 demonstrated more sensitivity to ATP than proapoptotic protein Bax. Decreasing extracellular Ca 2ϩ or chelating intracellular Ca 2ϩ with BAPTA-AM significantly inhibited ATP-induced increase in Bcl-2/ Bax ratio, indicating that a rise in [Ca 2ϩ ]cyt through Ca 2ϩ influx is the critical mediator for ATP-mediated increase in Bcl-2/Bax ratio. Therefore, despite high ATP levels in the tumor microenvironment, which would induce cell apoptosis in normal cells, the decreased P2X7 and elevated Bcl-2/Bax ratio in lung cancer cells may enable tumor cells to survive. Increasing the Bcl-2/Bax ratio by exposure to high extracellular ATP may, therefore, be an important selective pressure promoting transformation and cancer progression. purinergic receptor; calcium signaling; Bcl-2; Bax; cell apoptosis; cell proliferation THERE IS GROWING INTEREST in the role of ATP in the development of cancer. ATP, well known as an intracellular molecular energy source, also functions as an extracellular messenger (2). ATP receptors are purinergic receptors (P2 receptors) and include the ligand-gated ion channel family of P2 receptors (P2X1-7) as well as the G protein-coupled receptor (GPCR) family of P2 receptors (P2Y1-2, P2Y 4 , P2Y 6 and P2Y11-14) (3, 31). Activation of P2X receptors, which are nonselective cation channels formed by three homomeric or heteromeric P2X subunits, directly results in Na ϩ and Ca 2ϩ influx through the cell plasma membrane, leading to membrane depolarization, which in turn activates voltage-gated Na ϩ and Ca 2ϩ channels and causes the firing of action potentials. Activation of P2Y receptors, which are GPCRs, increases cytosolic free Ca 2ϩ concentration ([Ca 2ϩ ] cyt ) by inducing Ca 2ϩ release from intracellular stores (e.g., sarcoplasmic or endoplasmic reticulum) and Ca 2ϩ influx through store-operated (SOC) and/or receptor-operated (ROC) Ca 2ϩ channels. P2X and P2Y signaling is not only responsible for inducing action potentials in excitable cells (e.g., neurons), but also has been implicated in cell proliferation, differentiation, and apoptosis in nonexcitable cells (e.g., epithelial cells) (5). In a lung cancer cell line, A549, extracellular ATP, UTP, and UDP have been shown to stimulate proliferation that is dependent on the P2Y 2 and P2Y 6 receptors (44).Extracellular or intercellular ATP concentration is reported to be very low (1-5 M) in normal healthy tissues; however, it is significantly increased (to Ͼ100 M) in the tumor microenvironment (26). The effect of increased extracellular ATP on cancer cell function is, however, dependent of the expression of...

ATP (extracellular messenger) is in much higher concentration in tumor microenvironment. It regulates cell functions through acting on purinergic receptors to activate a series of intracellular signal transduction pathways, including Ca2+ signaling pathway. Here we report that ATP caused a long‐duration Ca2+ influx in lung cancer cells compared to normal control cells. Ionotropic (P2X3, P2X4 and P2X5) and metabotropic (P2Y2, P2Y4 and P2Y6) receptors were expressed in much higher lever in lung cancer cells than in normal control cells. UTP (P2Y receptor agonist) induced a transient increase in [Ca2+]cyt without obvious Ca2+ influx in lung cancer cells. Activation of P2X receptor with α,β‐meATP and D‐β,γ‐meATP caused Ca2+ influx in lung cancer cells. PLC inhibitor U73122 inhibited Ca2+ transient, while P2X receptor inhibitor TNP‐ATP significantly attenuated Ca2+ influx. Store‐operated Ca2+ entry was not significantly enhanced in lung cancer cells. ATP also increased Bcl‐2/Bax ratio in lung cancer cells, but not in normal control cells. BAPTA‐AM abolished and significantly inhibited ATP‐mediated increase in [Ca2+]cyt and Bcl‐2/Bax ratio in lung cancer cells. The ATP‐mediated increase in Bcl‐2/Bax ratio was also significantly attenuated by removing extracellular Ca2+. These data indicate that upregulated expression of P2X3, P2X4 and P2X5 receptors in lung cancer cells leads to a long‐duration Ca2+ influx induced by ATP that subsequently causes a sustained increase in Bcl‐2/Bax ratio and eventually a significant increase in cell viability or cell proliferation.

No abstract