Introduction

Glycerolipids and glycerophospholipids are key molecules in many inter- and intracellular signal transduction pathways. Some of these processes, such as the phospholipase C (PLC)-driven phosphatidylinositol cycle, have been known to participate in signal transduction since the 1950s (3). Phosphatidic acid (PA) and diacylglycerol (DAG) participate in signaling pathways initiated by growth factors and G protein-coupled receptors (GPCRs) as well. However, the importance of additional lipid classes as cellular signals has only more recently been appreciated. These include the participation of lysophosphatidic acid in apoptosis and lysophospholipids as ligands for certain GPCRs involved in cardiac, neuronal, and immunological processes (4-7).

Until recently, the detection and identification of low concentration lipids was quite difficult. Thin layer chromatography (TLC) was utilized for many decades to separate lipid classes. With the advent of gas chromatography and gas chromatography mass spectrometry (GC-MS), class separation by TLC followed by hydrolysis and derivatization made it possible to identify individual fatty acid species. One of the drawbacks to this method was that large amounts of lipids were normally required. With the introduction of fast atom bombardment mass spectrometry (FAB-MS), routine analysis of intact phospholipids was possible (8). More recently, electrospray ionization mass spectrometry (ESI-MS) has greatly simplified the procedures for lipid analysis. The soft ionization process associated with ESI-MS results in decreased molecular ion decomposition and lower detection limits compared to FAB-MS (9, 10).

A primary goal of these studies has been to identify the phospholipids participating in the cellular signaling pathways downstream of the AIG pathway in WEHI-231 cells as well as other ligands of interest to the AfCS. In this way, lipid signaling components can be integrated into the larger cellular signaling network, appreciated as part of the molecular response elements used by cells to transduce information from the cell surface, and better understood regarding the sometimes mysterious events occurring within cellular bilayers. As such, this report represents the most extensive analysis of cellular lipid content and changes determined to date. The ability to monitor changes in cellular lipid content in parallel with the ability of other AfCS laboratories to determine changes in gene expression, protein modifications, and production of second messengers represents a powerful new approach to understanding the contextual changes that determine cellular responses to complex biological stimuli.