Survival of cultured cells after functional and structural disorganization of plasma membrane by bacterial haemolysins and phospholipases

Abstract

Lesions were induced in the plasma membranes of cultured human fibroblasts by membrane damaging toxins of bacterial origin (haemolysins). Structural disorganization of the membrane was measured as leakage of a radiolabelled small cytoplasmic marker and functional membrane damage was measured as decreased uptake of aminoisobutyrate. Cell survival was scored 24 and 48 hr later by measuring uptake of Trypan Blue and by light microscopical evaluation of cell morphology and proliferation. The membrane damage induced by most bacterial toxins was reversible upon removal of the toxin, since toxin-treated cells recovered and excluded Trypan Blue although they had been permeable to the dye immediately after the toxin treatment. Among ten bacterial toxins tested, the only exception of this general behavior was the Aeromonas hydrophila beta-haemolysin, which irreversibly damaged human fibroblasts. Thus, the action of bacterial haemolysins on cultured cells generally seems restricted to a plasma membrane permeabilization, which is reversible regardless of the mechanism of membrane damaging action of the toxin or of the relative size of the structural lesions induced. Furthermore, the use of Trypan Blue uptake as a measure of cell death caused by membrane damaging agents appears to be of limited value.