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Nucleotides such as ATP can act as extracellular effector molecules by interaction with specific cellular receptors known as P2-purinoceptors. Recently, we cloned the human P2U purinoceptor from osteoclastoma and demonstrated its expression in skeletal tissues. In the current study we have investigated the expression of P2U purinoceptors in human breast tumour cell lines and examined functional effects of extracellular nucleotides on these cells. By reverse transcription-linked polymerase chain reaction (RT-PCR) the expression of mRNA for P2U purinoceptors was demonstrated in four human breast cancer cell lines, Hs578T, MCF-7, SK-Br3 and T47-D. In MCF-7 cells, extracellular ATP (1-100 microM) elevated intracellular free calcium concentration [Ca2+]i, indicating that these cells express functional P2-purinoceptors. UTP elevated [Ca2+]i in an identical manner to ATP, whereas 2-methylthioATP was completely ineffective, and ADP only partially effective. This pharmacological profile suggests that the P2U subtype may be the only P2-purinoceptor expressed by these cells. The functional significance of P2U purinoceptor expression by MCF-7 cells was investigated by analysing the effects of extracellular ATP on cell proliferation. The slowly hydrolysed analogue of ATP, ATPgammaS (which was also shown to elevate [Ca2+]i), induced proliferation of MCF-7 cells when added daily to serum-free cultures over a period of 3 days. ATPgammaS-induced proliferation was demonstrated by three separate methods, detection by scintillation counting of [3H]thymidine incorporation, immunocytochemical detection of 5-bromo-2-deoxyuridine incorporation and direct counting of cell numbers. These data suggest that ATP, possibly released at sites of tissue injury or inflammation, may be capable of growth factor action in promotion of tumour proliferation or progression.

Extracellular nucleotides stimulate proliferation in MCF-7 breast cancer cells via P2-purinoceptors