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Journal of Lipid Research, Vol. 43, 904-910, June 2002
Copyright © 2002 by Lipid Research, Inc.
Secretion of a lysophospholipase D activity by adipocytes
: involvement in lysophosphatidic acid synthesis
INSERM U317, Institut Louis Bugnard, Université Paul Sabatier, CHU Rangueil, Batiment L3, 31403, Toulouse cedex 04, France
1 To whom correspondence should be addressed. e-mail: saulnier{at}rangueil.inserm.fr
The aim of the present work was to depict the metabolic pathways involved in extracellular production of lysophosphatidic acid (LPA) by adipocytes. LPA was followed by quantifying the accumulation of LPA in the incubation medium (conditioned medium, CM) of 3T3F442A adipocytes or human adipose tissue explants using a radioenzymatic assay. Surprisingly, after separation from the cells, the amount of LPA present in CM could be significantly increased by further incubation at 37°C. This suggested the presence of a LPA-synthesizing activity (LPA-SA) in CM. LPA-SA appeared as a soluble activity which was inhibited by divalent ion chelators EDTA and phenanthrolin. The effect of EDTA was preferentially reverted by CoCl2, as described for a lysophospholipase D (lyso-PLD) activity previously identified in rat plasma. LPA concentration could also be increased by treatment with a bacterial PLD, demonstrating the presence of PLD-sensitive LPA precursors (mainly lysophosphatidylcholine) in adipocyte CM. LPA-SA could be increased by the addition of exogenous lysophosphatidylcholine, lysophosphatidylglycerol, or lyso-platelet activating factor, demonstrating that LPA-SA resulted from the action of a lyso-PLD. LPA-SA was not inhibited, but rather activated, by primary alcohol (ethanol and 1-butanol), suggesting that adipocyte lyso-PLD was not a classical PLD. Finally, LPA-SA was found to be weaker in CM of undifferentiated adipocyte (preadipocytes) compared with CM of differentiated adipocytes.
In conclusion, our results reveal the existence of a secreted lyso-PLD activity regulated during adipocyte-differentiation and involved in extra cellular production of synthesis of LPA by adipocytes.
Abbreviations: CHCl3, chloroform; CM, conditioned medium; LPA, lysophosphatidic acid; LPAAT, lysophosphatidic acid acyltransferase; LPAF, lyso-platelet activated factor; LPA-SA, lysophosphatidic acid-synthesizing activity; LPC, lysophosphatidylcholine; LPG, lysophosphatidylglycerol; MeOH, methanol; PA, phosphatidic acid; PC, phosphatidylcholine; PLA2, phospholipase A2; PLD, phospholipase D
Supplementary key words adipose tissue • 3T3F442A preadipocytes • human • mouse • phospholipase D • phospholipase A2 • conditioned medium • culture • lysophospholipids • lysophosphatidylcholine • phosphatidic acid • radioenzymatic assay
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