The membranes of healthy lymphocytes normally resist hydrolysis by secretory
phospholipase A(2). However, they become susceptible during the process of apoptosis. Previous experiments have demonstrated the importance of certain physical changes to the membrane during cell death such as a reduction in
membrane lipid order and exposure of
phosphatidylserine on the membrane surface. Nevertheless, those investigations also showed that at least one additional factor was required for rapid hydrolysis by the human group IIa
phospholipase isozyme. This study was designed to test the possibility that oxidation of
membrane lipids is the additional factor. Flow cytometry and confocal microscopy with a
fluorescent probe of oxidative potential suggested that oxidation of the plasma membrane occurs during apoptosis stimulated by
thapsigargin. When oxidative potential was high, the activity of human group IIa secretory
phospholipase A(2) was enhanced 30- to 100-fold compared to that observed with conditions sufficient for maximal hydrolysis by other secretory
phospholipase A(2)
isoforms. Direct oxidation of cell membranes with either of two
oxidizing agents also stimulated hydrolysis by secretory
phospholipase A(2). Both oxidizers caused externalization of
phosphatidylserine, but a change in
lipid order did not always occur. These results demonstrated that membrane oxidation strongly stimulates human group IIa secretory
phospholipase A(2) activity toward apoptotic cells. Interestingly, the change in membrane order, previously thought to be imperative for high rates of hydrolysis, was not required when
membrane lipids were oxidized. Whether
phosphatidylserine exposure is still necessary with oxidation remains unresolved since the two events could not be deconvoluted.