Human erythrocyte homeostasis involves the activation of a novel phospholipase C

NOSEDA DIEGO GABRIEL; GUIMENEZ G; FLORIN CHRISTENSEN J

Puerto Iguazu, Misiones

Congreso; Reunion Anual de la Sociedad Argentina de Investigaciones Bioquimicas (SAIB); 2004

Sociedad Argentina de Investigaciones Bioquímicas (SAIB)

We have investigated the metabolic fair of l-palmitoyl-2-(6-[(7- nitro-2-l,3-benzoxadiazol-4) amino]caproyl)-sn-glycero-3-phosphocholine (C6-NBD- PC) exposed to human red blood cells suspension. The method involved incubating at 37 C the fluorescent phospholipid with freshly washed human erythrocytes for 1 h. Afterwards, the red blood cells were washed again three times to remove any unincorporated C6-NBD-PC. The fluorescently labeled erythrocytes were added to an equal volume of plasma and incubated at 37 C with constant stirring. Aliquots were removed at different time points and centrifuged. Three kinds of samples were thus generated: 1) total erythrocyte plus plasma suspensions, 2) plasma supernatant, and 3) red blood cell pellets. Lipids from these samples were extracted by a modification of the procedure of Bligh and Dyer and analyzed by thin layer chromatography on silica gel plates, using a double developing system, as described by Florin-Christensen et al. The results show, surprisingly, a striking formation of diacylglycerol (DG) which was entirely recovered in the plasma samples. These results demonstrate for the first time, the occurrence of a PC-PLC in human erythrocytes and suggest that plasma lipoproteins possess high affinity for the DG. These observations open the possibility that plasma membrane DG from other type of cells could be removed by interaction with plasma lipoproteins, therefore constituting a novel termination mechanism for cellular DG signals.