Abstract
In order to study the mechanism of the accumulation of phosphatidyl choline (PC) in erythrocytes with abnormal erythrocyte phospholipids from patients with a hereditary hemolytic anemia, the phospholipids of the erythrocytes were labeled radioactively. Labeling of phosphatides was achieved by both passive equilibration with preformed phosphatides, and active "acylase"-mediated incorporation of fatty acid (FA) in the presence of glucose, ATP and coenzyme A. The labeled cells were then reincubated in fresh compatible sera and the catabolism of the labeled erythrocyte phospholipids was followed. In addition, total acylase capacity of erythrocyte stroma was determined under optimal conditions in a system with excess lysophosphatide, FA, ATP, CoA, and Mg++. No differences in passive uptake or release of phosphatides were found between the patients’ erythrocytes and comparable reticulocyte-rich controls. On the other hand, overall active incorporation of FA into PC was abnormally increased in the patients’ erythrocytes, whereas incorporation of FA into phosphatidyl-ethanolamine (PE) was decreased. However, acylase capacity for both lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE) was normal in the patients’ cells. This apparent paradox could be explained by the subsequent turnover of actively incorporated PC-FA which was found to be reduced. A brief labeling experiment designed to approximate pulse-chase conditions and to label primarily PC showed a considerable inhibition of the subsequent transfer of PC-FA to PE upon reincubation in fresh serum. This transfer has previously been shown to be responsible for a significant portion of PC-FA catabolism. Reincubation in hyperlipemic sera obtained from patients with liver disease or artificially enriched with PC did not influence the abnormal outflow of phosphatide-FA in actively labeled cells. The findings were consistent with the concept that PC accumulated in these cells because of a defect in the catabolism of actively incorporated PC-FA. This defect appeared to be in the transfer of PC-FA to PE prior to final release from the cell. Passive exchange pathways and the active anabolic acylase pathway were not abnormal in these patients’ erythrocytes.
