Chemical co-treatments and intramembrane particle patching in the poly(ethylene glycol)-induced fusion of turkey and human erythrocytes

Abstract

Several chemical co-treatments were used to lower the threshold concentrations of poly(ethylene glycol) (PEG) required to induce fusion between turkey erythrocytes and between human erythrocytes. Concanavalin A was used in conjuction with 25% (w/w) PEG to induce turkey erythrocyte fusion. The fusion percentage increased with increasing concentrations of concanavalin A and the duration of concanavalin A treatment. In samples with high percentages of fusion, numerous hemispherical intramembraine particle-free zones (bubbles) in the plasma membrane were revealed by freeze-fracture electron microscopy. However, concanavalin A treatment did not facilitate fusion between human erythrocytes even at 35% PEG, although slight intramembrane particle patching was observed under this condition. Spermidine (0.05% w/v), trichloroacetic acid (100 mM) and ethanol (4% v/v) were found to promote fusion of human erythrocytes in 25% PEG. In all of these cases, intramembrane particle patching was observed by freeze-fracture electron microscopy in the presence of PEG. When applied alone, only ethanol caused a slight intramembrane particle patching. Neither dimethylsulfoxide (2% v/v), lysophosphatidylcholine (lysoPC, 0.15 mM), nor polylysine (mol. wt. 1000–4000, 0.05% w/v) promoted fusion of human erythrocyte in 25% PEG. None of these chemical treatments, alone, or in combination with PEG, caused intramembrane particle patching. We conclude that the positive effect of chemical treatments on PEG-induced cell fusion is closely related to the formation of intramembrane particle-free zones on the plasma membrane.