Density-associated changes in platelet-activating factor acetylhydrolase activity and membrane fluidity of human...

Platelet-activating factor acetylhydrolase is known to degrade oxidatively fragmented phospholipids which are similar in structure to platelet-activating factor. We examined changes of acetylhydrolase activity during in vivo aging of human erythrocytes and tried to assess its role in maintaining the membrane properties of erythrocytes. Higher-density erythrocytes are enriched with older cells. Erythrocytes obtained from seven healthy colleagues were separated into four density fractions by centrifugation in discontinuous Percoll density gradients. Both membrane and cytosolic acetylhydrolase decreased with increasing erythrocyte density. Membrane and cytosolic acetylhydrolase activities in the lightest fraction were 2.0 +/- 1.0 (SD) nkat/g protein and 362 +/- 58 pkat/g protein, respectively, and these values were significantly higher than those in the densest fraction: 1.3 +/- 0.7 nkat/g protein and 286 +/- 70 pkat/g protein, respectively. Membrane acyltransferase activity also decreased with red cell density and the average values in the lightest and densest fractions were 51.2 +/- 23.6 and 27.0 +/- 20.2 mukat/g protein, respectively. Generation of thiobarbituric acid-reactive substances induced by t-butyl hydroperoxide treatment decreased with increasing cell density, and the inhibition of acetylhydrolase with diisopropylfluorophosphate resulted in enhanced peroxide-induced lipid oxidation, particularly in lower-density fractions. There was no significant change in basal levels of thiobarbituric acid-reactive substances in red cell membrane. Membrane fluidity was evaluated by fluorescence recovery after photo-bleaching and it decreased as erythrocyte density increased. We conclude that the activity of the deacylation/reacylation cycle maintained by acetylhydrolase and acyltransferase is gradually reduced during in vivo aging of erythrocytes. This may be connected with decreases of polyunsaturated fatty acids and membrane fluidity in old erythrocytes.