Role of phospholipases A(2) in growth-dependent changes in prostaglandin release from 3T6 fibroblasts.
Previously, we reported a growth-dependent change in prostaglandin production as a consequence of a marked growth-dependent alteration in arachidonic acid (AA) mobilization from phospholipids. Our present results show that fetal calf serum (FCS) and 4 beta-phorbol-12-myristate acetate (PMA) caused an enhancement of phospholipase A(2) (PLA(2)) activity in the membrane fraction of non-confluent cells allowing PLA(2) access to its substrate and the release of AA. Western blot analysis has shown that FCS and PMA increased secreted PLA(2) (sPLA(2)) expression in non-confluent 3T6 fibroblast cultures. Moreover, FCS and PMA induced dithiothreitol-sensitive and bromoenol lactone-sensitive PLA(2) activities in cytosol and membrane fraction. However, these stimuli did not modify significantly the PLA(2) activity in both fractions when 3T6 fibroblasts reached a high cell density. This could be associated with the impairment of AA mobilization in these cell culture conditions. On the other hand, we observed that FCS and PMA induced the same prostaglandin H synthase-2 induction in non-confluent and confluent culture conditions. Moreover, the prostaglandin E(2) levels reached in cell culture supernatants were independent of the degree of confluence when AA was added exogenously. These results suggest that the changes of intracellular distribution of PLA(2) activity of sPLA(2) and iPLA(2) stimulated by exogenous stimuli may be controlled by cell density conditions which constitute an important mechanism in the regulation of prostaglandin release.
Sánchez
T
T
Department of Physiology, School of Pharmacy, Barcelona University, Spain.
Moreno
J J
JJ
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division; D002460:Cell Line; D003470:Culture Media; D051546:Cyclooxygenase 2; D015232:Dinoprostone / Q000096:biosynthesis*; D005347:Fibroblasts / Q000166:cytology / Q000201:enzymology*; D054505:Group V Phospholipases A2; D054522:Group VI Phospholipases A2; D007425:Intracellular Membranes / Q000201:enzymology; D007527:Isoenzymes / Q000378:metabolism; D051379:Mice; D010741:Phospholipases A / Q000378:metabolism / Q000502:physiology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D003470:Culture Media; D007527:Isoenzymes; D016718:Arachidonic Acid; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D010741:Phospholipases A; D054505:Group V Phospholipases A2; D054522:Group VI Phospholipases A2; C516300:Pla2g5 protein, mouse; C516476:Pla2g6 protein, mouse; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate