MOLECULAR-SPECIES ANALYSIS OF 1,2-DIGLYCERIDES ON PHORBOL ESTER STIMULATION OF LA-N-1 NEUROBLASTOMA-CELLS DURING PROLIFERATION AND DIFFERENTIATION

1,2-Diacyl-sn-glycerol (DAG) is a product of cell activation that has emerged as an important intracellular messenger whose primary function appears to be the activation of protein kinase C. They originate by t he activation of phospholipases, which hydrolyze different phospholipi ds depending on the external stimulus and the nature of the cells, lea ding to the production of different molecular species. In the present study the quantitative changes in the total mass and the molecular spe cies of DAG formed on phorbol ester (12-O-tetradecanoyl-phorbol 13-ace tate) stimulation were investigated in proliferating and retinoic acid (RA)-differentiated human LA-N-1 cells, The TPA treatment of both cel l types elicited an increase in the total amount of DAG. The increase was biphasic; i.e., an initial peak at 2-5 min was followed by a susta ined increase that persisted for >30 min. The analysis of the molecula r species of DAG and phospholipids showed that in proliferating LA-N-1 cells, the DAG increase corresponds to the production of mainly satur ated/monounsaturated (16:0-18:1, 18:0-18:1) and saturated/saturated (1 6:0-16:0, 16:0-18:0) species, suggesting that they originate essential ly from the hydrolysis of phosphatidylcholine. In contrast, RA-differe ntiated cells responded to TPA treatment by increasing the level of sa turated/polyunsaturated (16:1-22:6, 18:0-22:6, 16:0-20:4, 18:0-20:4) a nd monounsaturated/monounsaturated (18:1-18:1) species, suggesting mai nly a phosphatidylethanolamine origin. These findings indicate that th e treatment of LA-N-1 cells with TPA generates different molecular spe cies of DAG depending on their physiological state, These observations suggest in turn that different phospholipases are activated by TPA in proliferating and RA-differentiated cells.