Phorbol ester stimulates ethanolamine release from the metastatic basal prostate cancer cell line PC3 but not from prostate epithelial cell lines LNCaP and P4E6.

Malignancy alters cellular complex lipid metabolism and membrane lipid composition and turnover. Here, we investigated whether tumorigenesis in cancer-derived prostate epithelial cell lines influences protein kinase C-linked turnover of ethanolamine phosphoglycerides (EtnPGs) and alters the pattern of ethanolamine (Etn) metabolites released to the medium. Prostate epithelial cell lines P4E6, LNCaP and PC3 were models of prostate cancer (PCa). PNT2C2 and PNT1A were models of benign prostate epithelia. Cellular EtnPGs were labelled with [1-(3)H]-Etn hydrochloride. PKC was activated with phorbol ester (TPA) and inhibited with Ro31-8220 and GF109203X. D609 was used to inhibit PLD (phospholipase D). [(3)H]-labelled Etn metabolites were resolved by ion-exchange chromatography. Sodium oleate and mastoparan were tested as activators of PLD2. Phospholipase D activity was measured by a transphosphatidylation reaction. Cells were treated with ionomycin to raise intracellular Ca(2+) levels. Unstimulated cell lines release mainly Etn and glycerylphosphorylEtn (GPEtn) to the medium. Phorbol ester treatment over 3h increased Etn metabolite release from the metastatic PC3 cell line and the benign cell lines PNT2C2 and PNT1A but not from the tumour-derived cell lines P4E6 and LNCaP; this effect was blocked by Ro31-8220 and GF109203X as well as by D609, which inhibited PLD in a transphosphatidylation reaction. Only metastatic PC3 cells specifically upregulated Etn release in response to TPA treatment. Oleate and mastoparan increased GPEtn release from all cell lines at the expense of Etn. Ionomycin stimulated GPEtn release from benign PNT2C2 cells but not from cancer-derived cell lines P4E6 or PC3. Ethanolamine did not stimulate the proliferation of LNCaP or PC3 cell lines but decreased the uptake of choline (Cho). Only the metastatic basal PC3 cell line specifically increased the release of Etn on TPA treatment most probably by PKC activation of PLD1 and increased turnover of EtnPGs. The phosphatidic acid formed will maintain a cancer phenotype through the regulation of mTOR. Ethanolamine released from cells may reduce Cho uptake, regulating the membrane PtdEtn:PtdCho ratio and influencing the action of PtdEtn-binding proteins such as RKIP and the anti-apoptotic hPEBP4. The work highlights a difference between LNCaP cells used as a model of androgen-dependent early stage PCa and androgen-independent PC3 cells used to model later refractory stage disease.