Reactions of the alkylating agent tris(2-chloroethyl)-amine with the erythrocyte membrane: Effects on shape changes of human erythrocytes and ghosts

Abstract

The influence of tris(2-chloroethyl)amine on shape changes of human erythrocytes and ghosts was studied in vitro and correlated with alterations in the molecular structure of the membrane. 1. (1) Reaction with 1–2 mM tris(2-chloroethyl)amine, a concentration which caused polymerisation of spectrin as detected by sodium dodecyl sulphate polyacrylamide gel electrophoresis, protected intact erythrocytes against metabolically induced shape changes. 2. (2) When induced by Mg 2+-ATP, ghosts prepared from alkylated erythrocytes underwent normal changes in shape. However, when ghosts were treated directly with tris(2-chloroethyl)amine, no Mg 2+-ATP-induced shape changes occurred. This fixation in shape appeared to be due to a higher degree of reaction with the alkylating agent. 3. (3) The amount of chlorpromazine necessary for transformation of erythrocytes into stomatocytes was increased for tris(2-chloroethyl)amine-pretreated cells and was dependent on the degree of reaction with tris(2-chloroethyl)-amine. 4. (4) Deformability of red cells after reaction with tris(2-chloroethyl)amine was estimated by measuring their rheological behaviour in glass capillary arrays. A slight reduction of the flow rate was observed for cells alkylated with 1–2 mM tris(2-chloroethyl)amine. 5. (5) Extractability of spectrin was diminished and corresponded to the degree of spectrin polymerisation.