5-hydroxy-6-8-11-14-eicosatetraenoic-acid and 4-bromophenacyl-bromide

Serum-cultured rat W256 carcinosarcoma cells of the monocytoid origin undergo rapid apoptosis in response to the lipoxygenase inhibitor NDGA (nordihydroguaiaretic acid). Exogenous arachidonic acid (AA), in a time- and dose-dependent fashion, suppressed NDGA-induced W256 cell apoptosis as well as DNA fragmentation, with the maximal effect observed at approximately 25 microM. Mobilization of endogenous AA by calcium ionophore A23187 provided an even stronger and longer-lasting protection against NDGA-caused cell death. The A23187 effect on AA release as well as W256 cell death can be blocked by bromophenacyl bromide, thus suggesting involvement of phospholipase A2 activation. Serum withdrawal similarly caused W256 cells to undergo typical apoptosis, which was not rescued by several growth factors commonly found in serum. However, exogenous AA suppressed serum starvation-induced W256 cell apoptosis and significantly extended cell survival in a dose-dependent manner. Lipoxygenase products, 12(S)- and 15(S)-, but not 5(S)-hydroxyeicosatetraenoic acid (HETE), in a dose-dependent fashion, also prevented both NDGA- and serum-starvation-induced W256 cell apoptosis. AA appears to suppress W256 cell apoptosis via distinct signaling pathway(s) since it does not prevent cell death triggered by several other inducers. Examination of a panel of polyunsaturated fatty acids revealed that alpha-linolenic and linoleic acid can also suppress NDGA-induced W256 cell apoptosis. Our data suggest that AA and other polyunsaturated fatty acids and/or their metabolites may enhance tumor growth not only by promoting cell proliferation but also by suppressing apoptosis.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Acetophenones; Animals; Apoptosis; Arachidonic Acid; Calcimycin; Carcinosarcoma; Cell Line; Cell Survival; Cytokines; DNA Fragmentation; Fatty Acids, Nonesterified; Growth Substances; Hydroxyeicosatetraenoic Acids; Kinetics; Masoprocol; Rats; Tumor Cells, Cultured

The cross-talk of arachidonic acid (AA) and its lipoxygenase products with protein kinase C beta (PKC beta) mRNA levels during the action of gonadotropin-releasing hormone (GnRH) was investigated in the pituitary alpha T3-1 cell line. The addition of AA or its 5-lipoxygenase products 5-hydroxyeicosatetraenoic acid (5-HETE) or leukotriene C4 (LTC4) for 30 or 60 min stimulated PCK beta, but not PKC alpha mRNA levels (3-5-fold); PCK gamma is not expressed by the cells. Other HETEs or leukotrienes tested showed no significant effect. The range of effective concentration for LTC4 and 5-HETE (around 10(-10) M) is the range found in GnRH-stimulated pituitary cells. Although PKC beta mRNA levels were preferentially elevated by LTC4 and 5-HETE at early time points, PKC alpha mRNA levels were elevated at 6-12 h of incubation when PKC beta mRNA levels returned to basal levels. The addition of the phospholipase A2 inhibitor 4-bromophenacyl bromide or the selective 5-lipoxygenase inhibitor L-656,224 abolished [D-Trp6]GnRH (GnRH-A) elevation of PKC beta mRNA levels, whereas PKC alpha mRNA levels were not increased by this neurohormone. The cyclo-oxygenase inhibitor indomethacin elevated basal PKC beta mRNA levels and potentiated the GnRH-A response. Cross-talk exists between AA and some of its lipoxygenase products and PKC beta gene expression during cell signalling. AA, 5-HETE and LTC4 participate in the rapid stimulation of PKC beta mRNA levels by GnRH.

Topics: Acetophenones; Anti-Inflammatory Agents, Non-Steroidal; Arachidonic Acid; Benzofurans; Cell Line; Cyclooxygenase Inhibitors; Enzyme Inhibitors; Gonadotropin-Releasing Hormone; Hydroxyeicosatetraenoic Acids; Indomethacin; Kinetics; Leukotriene C4; Lipoxygenase; Lipoxygenase Inhibitors; Pituitary Gland; Protein Kinase C; Protein Kinase C beta; RNA, Messenger

Incubation of rocker-cultured neonatal rat heart cells with 3 mM L(+)-lactate led to a sharp increase in the sensitivity of cardiomyocytes to the beta-adrenergic agonist isoprenaline, as measured by their chronotropic response. This effect was accompanied by a reduction in the arachidonic acid content of the total phospholipids. The phospholipase A2-activator melittin as well as free arachidonic acid induced this supersensitivity to the same degree. On the other hand, the L(+)-lactate-evoked supersensitivity could be blocked by the phospholipase A2 inhibitors mepacrine and n-bromophenacyl-bromide, suggesting an involvement of phospholipase A2 in the process of beta-adrenergic sensitization. The sensitizing action of arachidonic acid was blocked by the lipoxygenase inhibitors esculetin and nordihydroguaiaretic acid, but not by the cyclo-oxygenase inhibitor indomethacin. Supersensitivity was likewise evoked by 15-S-hydroxyeicosatetraenoic acid (15-S-HETE), but not by 5-S-HPETE or 5-S-HETE. These findings suggest that the phospholipase A2-15-lipoxygenase pathway plays a role in the induction of beta-adrenergic supersensitivity in the cultured cardiomyocytes and point to a new physiological role of the lipoxygenase product 15-S-HETE.

Topics: Acetophenones; Animals; Arachidonic Acid; Arachidonic Acids; Cells, Cultured; Chronobiology Phenomena; Dose-Response Relationship, Drug; Enzyme Activation; Heart; Hydroxyeicosatetraenoic Acids; Isoproterenol; Lactates; Leukotrienes; Melitten; Phospholipases A; Phospholipases A2; Prostaglandins D; Quinacrine; Rats; Rats, Inbred Strains; Receptors, Adrenergic, beta