Mechanisms of membrane damage for CHO cells heated in suspension

Chinese hamster ovary cells were heated for 20 min at 45.5 degrees C in different conditions, and quantitative determinations of cellular membrane blebbing were performed for cells maintained at 25 degrees C and 37 degrees C after hyperthermia. The percentage of cells with blebs following heating was dependent upon the composition of the medium during heating and the posthyperthermia temperature after heating. The total extent of bleb formation after heating was independent of the calcium-ion concentration in the medium during heating; however, differences in the kinetics of bleb disappearance after heating point to the importance of Ca2+ concentration in the expression of heat damage. Without hyperthermia, blebs were formed on the cell-surface membrane with agents which block sulfhydryl groups or release calcium from cellular stores. The cells were protected from bleb formation when cells were incubated with glutathione before addition of sulfhydryl-blocking agents or heat treatment. Oligomycin did not prevent the formation of blebs, suggesting that this phenomenon is not energy-dependent. Only a small percentage of cells were covered with blebs when they were heated in saline solution. When cells were incubated with dbcAMP before heat, blebs did not appear at 25 degrees C. A possible interpretation for these observations is presented.