Cholesterol nucleation-influencing activity in t-tube bile

Abstract

Nucleation-influencing activity was determined in T-tube bile samples derived from patients with obstructive jaundice. Since native T-tube bile samples do not nucleate, nucleation-influencing activity was determined by measuring the influence of T-tube bile on the nucleation time of model bile. In the assay, T-tube bile was mixed with model bile, and the nucleation time of this mixture was compared with the nucleation time of a model bile supplemented with the same amount of lipid as present in the bile sample. The results were expressed as ratio of the nucleation time of the mixture and the nucleation time of the control (NT_m/NT_c). There was a significant difference (p < 0.01) between bile samples from patients with cholesterol gallstones and samples from patients with biliary obstruction due to other causes. More than 80% of the 33 samples from eight patients with stones were nucleation-promoting (NT_m/NT_c ≤ 0.6). Of the 40 bile samples from patients without stones, 7 were nucleation-promoting, 25 had no effect (NT_m/NT_c = 0.8 to 1.2) and 8 bile samples were nucleation-inhibiting (NT_m/NT_c ≥ 1.4). There was no correlation between the lipid or protein content of a T-tube bile sample and its nucleation-influencing activity. The presence of both nucleation-promoting and nucleation-inhibiting activity in the same T-tube bile was demonstrated by chromatography on concanavalin A-Sepharose. More than 75% of the biliary protein did not bind to the column. This fraction showed nucleation-inhibiting activity. However, nucleation inhibitor present in this fraction was unstable, and after destruction of the inhibitor, the fraction showed promoting activity. This suggests that, in addition to inhibitor, this fraction also contained nucleation-promoting activity. About 10% of biliary protein did bind to concanavalin A-Sepharose and could be eluted with α-D-methylmannopyranoside. This fraction always contained nucleation-promoting activity. Both this factor and the nucleation-promoting factor that did not bind to concanavalin A were resistant to treatment with the proteolytic enzyme pronase. We conclude that T-tube bile contains at least three different classes of nucleation-influencing factors. The balance between promoting and inhibiting activity in a particular bile sample determines the overall nucleation-influencing effect of the sample. In bile from cholesterol gallstone patients, this balance is shifted towards promoting activity. One of the promoting factors probably is a glucose-/mannose-containing glycoprotein.