Effects of serpins on cholesterol crystallization in model bile

Abstract

Background/Aims: The serine proteinase inhibitors (serpins), alpha-1-antitrypsin (AAT) and alpha-1-antichymotrypsin (ACT), are acute phase proteins synthesized by hepatocytes and excreted to some extent into bile. Their role in gallstone pathogenesis is unclear, and it was the aim of this study to determine their effect on cholesterol crystal growth rate in model bile. Methods: Purified AAT and ACT were added to model bile at concentrations from 0.5 to 500 μg/ml. Cholesterol crystal growth was analyzed daily by polarizing microscopy and spectrophotometrically at 650 nm. Serpin inhibitory activity was measured spectrophotometrically at 405 nm, and polymerization was studied on 7.5% SDS-PAGE under non-reducing conditions, by immunoelectrophoresis and Western blotting. Results: ACT added to model bile at a concentration of 0.5 μg/ml, inhibited cholesterol crystallization by 30%, had no influence at 5 μg/ml, and increased the crystallization rate 2–3 fold at concentrations of 50 and 500 μg/ml. AAT at a concentration of 0.5 μg/ml had a profound (50%) inhibitory effect on cholesterol crystal growth rate, lacked significant effect at both concentrations of 5 and 50 μg/ml, and showed stimulation of crystal growth up to 30% at a concentration of 500 μg/ml. Both serpins incubated in model bile polymerized and totally lost their inhibitory activity. Conclusions: Serpins can exhibit both inhibiting and promoting effects on the cholesterol crystallization rate in model bile, dependent on their concentrations. Since AAT and ACT are acute phase proteins, their concentrations may vary under certain pathological conditions, which may result in different actions of these serpins in gallstone formation.