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 modulated by cholesterol crystallization effector substances. In this study, we developed a method for quantitative assessment of vesicle fusion and used it to partly characterize the mechanisms of action of cholesterol crystallization effector proteins.


METHODS
Cholesterol:phospholipid (1:1) liposomes were prepared and labelled 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, immunoglobulins, apolipoprotein A-I and apolipoprotein B (all at 100 microg/mL).


RESULTS
Fusion of cholesterol/phospholipid liposomes was increased by 16 and 14% in the presence of concanavalin-A bound glycoprotein and immunoglobulins, respectively, and decreased by 21 and 9% after addition of apolipoprotein 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 modulate the stability of vesicles and, thus, affect cholesterol crystallization. Such modulation is based upon protein-vesicle association, which defines the physico-chemical metastability of vesicular cholesterol.
Hattori
Y
Y
First Department of Internal Medicine, Hiroshima University School of Medicine, Hiroshima, Japan.
Tazuma
S
S
Yamashita
G
G
Kajiyama
G
G
D016632:Apolipoprotein A-I / Q000737:chemistry; D001055:Apolipoproteins B / Q000737:chemistry; D001646:Bile / Q000737:chemistry*; D002784:Cholesterol / Q000737:chemistry*; D003460:Crystallization; D005456:Fluorescent Dyes; D006801:Humans; D007136:Immunoglobulins / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry*; D008561:Membrane Fusion; D008954:Models, Biological; D011506:Proteins / Q000737:chemistry*; D011952:Receptors, Concanavalin A / Q000737:chemistry; D012235:Rhodamines; D013997:Time Factors
D016632:Apolipoprotein A-I; D001055:Apolipoproteins B; D005456:Fluorescent Dyes; D007136:Immunoglobulins; D008081:Liposomes; D011506:Proteins; D011952:Receptors, Concanavalin A; D012235:Rhodamines; C027774:concanavalin A-binding glycoproteins; D002784:Cholesterol; C044542:octadecyl Rhodamine B chloride