Influence of cholesterol crystallization effector proteins on vesicle fusion in supersaturated model bile

Background: In lithogenic bile, cholesterol-rich vesicles rapidly aggregate and fuse to eventually form cholesterol crystals. This process is modulate d by cholesterol crystallization effector substances. In this study, we dev eloped a method for quantitative assessment of vesicle fusion and used it t o partly characterize the mechanisms of action of cholesterol crystallizati on effector proteins. Methods: Cholesterol:phospholipid (1:1) liposomes were prepared and labelle d with octadecyl rhodamine B chloride (R18). Fusion of these liposomes was detected by the increase of R18 fluorescence after incubation with various proteins, such as albumin, concanavalin-A bound glycoprotein, immunoglobuli ns, apolipoprotein A-I and apolipoprotein B (all at 100 mu g/mL). Results: Fusion of cholesterol/phospholipid liposomes was increased by 16 a nd 14% in the presence of concanavalin-h bound glycoprotein and immunoglobu lins, respectively, and decreased by 21 and 9% after addition of apolipopro tein A-I and apolipoprotein B, respectively. The effect of each protein on vesicle fusion was correlated with its hydrophobicity. Conclusions: These results suggest that nucleation effector proteins modula te the stability of vesicles and, thus, affect cholesterol crystallization. Such modulation is based upon protein-vesicle association, which defines t he physico-chemical metastability of vesicular cholesterol. (C) 1999 Blackw ell Science Asia Pty Ltd.