title: Role of endogenous prostaglandin E2 in erythropoietin production and dome formation by human renal carcinoma cells in culture.
abstract: Studies were carried out on the role of endogenous prostaglandin E2 (PGE2) in erythropoietin (Ep) production and dome formation in primary monolayer cultures of a human renal carcinoma from a patient with erythrocytosis that has been serially transplanted into BALB/c athymic nude mice. The metabolism of [14C]arachidonic acid (14C-AA) by cultured renal carcinoma cells, which were plated in 25-cm2 flasks at a density of 2 X 10(4) cells/cm2 and grown for 6, 12 (confluence, 13 X 10(4) cells/cm2), 16, 24, and 30 d in Eagle's minimum essential medium (MEM) supplemented with 10% fetal bovine serum, was examined by using radiometric thin-layer chromatography (TLC). TLC revealed PGE2 to be the major metabolite of 14C-AA produced by the cultured cells throughout the 30 d of cultivation. In addition, the cultured cells at each time period were incubated for 24 h in 5 ml of serum-free Eagle's MEM and the levels of PGE2 and Ep in the incubated media were measured via radioimmunoassay. PGE2 levels in the serum-free media incubated with the cultured cells grown for 6 d were significantly (P less than 0.001) elevated (174 +/- 2.5 pg/ml, n = 5), compared with the unincubated control media (1.5 +/- 0.19 pg/ml, n = 5) and gradually decreased at each time period to 97.6 +/- 4.4 pg/ml (n = 5) at 30 d. On the other hand, the levels of Ep in the incubated media of the cells grown for 6 d were 11.5 +/- 0.52 mU/ml (n = 5) compared with 7.6 +/- 0.62 mU/ml (n = 5) in the control media. However, after the cultured cells became confluent, the levels of Ep in the incubated media showed a marked increase to 222.9 +/- 5.26 mU/ml (n = 5) at 30 d of cultivation. Multicellular hemicysts (domes) developed after the cultured cells reached confluence and their numbers increased with increasing time in confluence in parallel with the increase in Ep. Meclofenamate (MF) (3 X 10(-6)-3 X 10(-5) M), a prostaglandin synthesis inhibitor, produced a significant dose-related decrease in PGE2, Ep, and dome formation without producing a significant effect on cell viability in the 30-d cells. This inhibitory effect of MF on Ep production and dome formation was completely abolished by the addition of 10(-8) M PGE2 to the incubation medium. In conclusion, endogenous PGE2 plays an important role in supporting and/or stimulating Ep production and dome formation in cultured renal carcinoma cells.
mesh_terms: D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism; D002277:Carcinoma / Q000378:metabolism / Q000473:pathology* / Q000648:ultrastructure; D002471:Cell Transformation, Neoplastic / Q000187:drug effects / Q000648:ultrastructure; D002478:Cells, Cultured; D015232:Dinoprostone; D004921:Erythropoietin / Q000096:biosynthesis*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D007680:Kidney Neoplasms / Q000378:metabolism / Q000473:pathology* / Q000648:ultrastructure; D008469:Meclofenamic Acid / Q000494:pharmacology; D051379:Mice; D008819:Mice, Nude; D011448:Prostaglandin Antagonists / Q000494:pharmacology; D011458:Prostaglandins E / Q000096:biosynthesis / Q000502:physiology*; D011817:Rabbits
chemical_list: D001095:Arachidonic Acids; D011448:Prostaglandin Antagonists; D011458:Prostaglandins E; D004921:Erythropoietin; D016718:Arachidonic Acid; D008469:Meclofenamic Acid; D015232:Dinoprostone; D007213:Indomethacin