Effects of Drugs, Salts, and Phospholipid Vesicles on Hemoglobin Release from Hydrostatic Pressure-Treated Human Erythrocytes

Abstract

When human erythrocytes subjected to a pressure of 2.0 kbar were incubated at 0 degree C and atmospheric pressure, hemoglobin was released from the membrane. Effects of drugs, salts, and phospholipid vesicles on hemoglobin release were examined. The hemoglobin release was suppressed by the addition of amphiphathic drugs such as chlorpromazine, trifluoperazine, tetracaine, indomethacin, and phenylbutazone or by a hypertonic condition. The suppressive effect induced by these drugs or hypertonic buffer was enhanced by the addition of aminophospholipid vesicles containing phosphatidylserine (PS) and/or phosphatidylethanolamine (PE) but decreased by vesicles composed of phosphatidylcholine (PC) and sphingomyelin. In the isotonic buffer without these drugs, such effects of vesicles were small. The suppressive effects of PS and PE were decreased by chemical modification of carboxylic groups and amino groups with 1-ethyl-3-(3-dimethylaminopropyl)carbodimide and 2,4,6-trinitrobenzenesulfonate, respectively. The size of vesicles as well as phospholipid composition was also an important factor in the suppression of hemoglobin release. The ESR spectrum of pressure-treated membrane spin-labeled with a maleimide was changed by PC vesicles but not by PS vesicles. These results indicate that the resealing of pressure-induced membrane holes by amphipathic drugs or hypertonic medium is synergistic with aminophospholipid vesicles and the hole size is affected by the vesicle-membrane protein interaction.