Characteristics of the cholesterol efflux induced by novel seminal phospholipid-binding proteins.
Our recent results indicated that the major proteins of bovine seminal plasma (collectively called BSP proteins) stimulate cholesterol efflux from fibroblasts and that this process shows many differences compared to the efflux induced by apolipoprotein A-I (apoA-I)-containing lipoproteins. The present study was undertaken to investigate the BSP-mediated efflux mechanism. Compared to the slow and constant rate of cholesterol efflux induced by apoA-I-containing lipoproteins, the BSP proteins stimulated a rapid efflux that gradually reached a plateau. The addition of purified BSP proteins after the establishment of the plateau resulted in a further cholesterol efflux indicating that cellular cholesterol was still available for efflux. Incubation of unlabeled fibroblast culture with the spent medium containing BSP-generated lipid ([(3)H]cholesterol) particles obtained after the establishment of the plateau did not result in any cholesterol influx. Therefore, the plateau did not correspond to an equilibrium of the radiolabel between the medium and the cells but rather to a saturation of the efflux particles with cholesterol. Numerous studies have indicated that the cholesterol efflux induced by apoA-I-containing lipoproteins involves cell-surface receptor, caveolae and intracellular cholesterol mobilization. Therefore, we investigated these characteristics for the BSP-mediated cholesterol efflux. Binding of BSP proteins to cells (evaluated by immunoblotting) reached saturation rapidly and remained constant thereafter. However, after several washings the cell-bound BSP proteins were unable to promote significant cholesterol efflux. Both results indicate no correlation of cholesterol efflux with cell binding. Moreover, in comparison to apoA-I-mediated cholesterol efflux, BSP-mediated efflux was not abolished at temperatures below 22 degrees C indicating that the BSP-induced cholesterol efflux does not involve intracellular cholesterol mobilization. High-density lipoprotein- and apoA-I-mediated cholesterol efflux was inhibited by preincubating fibroblasts with progesterone, whereas the cholesterol efflux by BSP proteins was not, indicating that cell-surface caveolae do not participate in BSP-mediated cholesterol efflux. Our results indicate that the mechanism of cholesterol efflux by BSP proteins is unidirectional and is strikingly different from that mediated by apoA-I-containing lipoproteins.
Moreau
R
R
Department of Medicine, University of Montreal and Guy Bernier Research Centre, Maisonneuve-Rosemont Hospital, 5415 L'Assomption, Montreal, Quebec, Canada H1T 2M4.
Manjunath
P
P
D016632:Apolipoprotein A-I / Q000494:pharmacology; D001665:Binding Sites; D001692:Biological Transport; D002352:Carrier Proteins / Q000494:pharmacology*; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism*; D002794:Choline / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008075:Lipoproteins, HDL / Q000494:pharmacology; D008565:Membrane Proteins / Q000494:pharmacology*; D049294:Phospholipid Transfer Proteins; D011374:Progesterone / Q000494:pharmacology; D011506:Proteins / Q000494:pharmacology*; D029589:Seminal Plasma Proteins; D013696:Temperature; D014316:Tritium
D016632:Apolipoprotein A-I; D002352:Carrier Proteins; D008075:Lipoproteins, HDL; D008565:Membrane Proteins; D049294:Phospholipid Transfer Proteins; D011506:Proteins; D029589:Seminal Plasma Proteins; D014316:Tritium; D011374:Progesterone; D002784:Cholesterol; D002794:Choline