Spontaneous Cholecysto- and Hepatolithiasis in Mdr2-/- Mice: A Model for Low Phospholipid-Associated Cholelithiasis

Abstract

Previously, we identified needle-like and filamentous, putatively "anhydrous" cholesterol crystallization in vitro at very low phospholipid concentrations in model and native biles. Our aim now was to address whether spontaneous gallstone formation occurs in Mdr2 (Abcb4) knockout mice that are characterized by phospholipid-deficient bile. Biliary phenotypes and cholesterol crystallization sequences in fresh gallbladder biles and non-fixed liver sections were determined by direct and polarizing light microscopy. The physical chemical nature and composition of crystals and stones were determined by sucrose density centrifugation and before mass and infrared spectroscopy. Gallbladder biles of Mdr2-/- mice precipitate needle-like cholesterol crystals at 12 weeks of age on chow. After 15 weeks, more than 50% of Mdr2-/- mice develop gallbladder stones, with female mice displaying a markedly higher gallstone-susceptibility. Although gallbladder biles of Mdr2-/- mice contain only traces (≤ 1.1 mM) of phospholipid and cholesterol, they become supersaturated with cholesterol and plot in the left 2-phase zone of the ternary phase diagram, consistent with "anhydrous" cholesterol crystallization. Furthermore, more than 40% of adult female Mdr2-/- mice show intra- and extrahepatic bile duct stones. In conclusion, spontaneous gallstone formation is a new consistent feature of the Mdr2-/- phenotype. The Mdr2-/- mouse is therefore a model for low phospholipid-associated cholelithiasis recently described in humans with a dysfunctional mutation in the orthologous ABCB4 gene. The mouse model supports the concept that this gene is a monogenic risk factor for cholesterol gallstones and a target for novel therapeutic strategies.