Quantification of selective phosphatidylserine oxidation during apoptosis.

Abstract

Membrane phospholipids are gaining increasing attention as important mediators in a variety of signal transduction processes. Oxidation and changes in membrane topography of lipids are probably important elements in the regulation of phospholipid-dependent signaling. Phosphatidylserine (PS), in particular, is implicated in the regulation of macrophage-dependent clearance of apoptotic cell "corpses" in a pathway probably mediated by selective oxidation and translocation of PS in the plasma membrane. Here we describe our highly sensitive and specific assay to measure differential lipid peroxidation in individual phospholipid classes in live cells using metabolic integration of the fluorescent oxidation-sensitive fatty acid analog cis-parinaric acid (cis-PnA) and resolution of specific phospholipids by high-performance liquid chromatography (HPLC). These experimental approaches can provide insight into the roles and mechanisms of PS oxidation in the identification and clearance of apoptotic cells.