Transport of Eicosapentaenoic Acid-Derived PGE3, PGF3α, and TXB3 by ABCC4

Background: Eicosapentaenoic acid-derived prostaglandin (PG) E-3, PGF(3 alpha), and thromboxane (TX) B-3 are bioactive lipid mediators which have anti-cancer and anti-inflammatory effects. To exert their effects, PGE(3), PGF(3 alpha), and TXB3 must be released to the extracellular space from cells, but the release mechanism has been unclear. We therefore investigated the contribution of ATP-binding cassette transporter C4 (ABCC4), which has been known as a prostanoids efflux transporter, to the release of PGE(3), PGF(3 alpha), and TXB3. Materials and Methods: ATP-dependent transport of PGE(3), PGF(3 alpha), and TXB3 via ABCC4 was investigated by using inside-out membrane vesicles prepared from ABCC4-overexpressing HEK293 cells. To evaluate the contribution of ABCC4 to the release of PGE(3), PGF(3 alpha), and TXB3, we measured the extracellular and intracellular levels of PGE(3), PGF(3 alpha), and TXB3 in A549 cells when we used ABCC4 inhibitors (dipyridamole, MK571, and probenecid) or ABCC4 siRNAs. The quantification of PGE(3), PGF(3 alpha), and TXB3 was performed by using liquid chromatography-tandem mass spectrometry. Results: The apparent K-m values for ABCC4-mediated transport were 2.9 +/- 0.1 mM for PGE(3), 12.1 +/- 1.3 mM for PGF(3 alpha), and 11.9 +/- 1.4 mM for TXB3 and the ATP-dependent accumulation of PGE(3), PGF(3 alpha), and TXB3 into vesicles was decreased by using typical substrates and inhibitors of ABCC4. ABCC4 inhibitors and ABCC4 knockdown showed the reduction of extracellular/intracellular ratio of PGE(3) (40-60% of control) and PGF(3 alpha) (60-80% of control) in A549 cells. Conclusions: Our results suggest that PGE(3), PGF3 alpha, and TXB3 are substrates of ABCC4 and ABCC4 partially contributes to the release of PGE(3) and PGF(3 alpha).