Lack of phospholipids in bile enhances cholesterol cholelithogenesis in the ATP‐binding cassette transporter B4 (Abcb4) knockout mice

Abstract

BackgroundBiliary phospholipids play a key role in transporting cholesterol (Ch) by forming unilamellar and multilamellar vesicles that display a significantly higher ability to solubilize Ch in bile compared to simple and mixed micelles. The ATP‐binding cassette transporter B4 (ABCB4) is responsible for hepatic phospholipid secretion and its deletion in the liver causes a lack of phospholipids in bile of mice. Our aim is to explore whether targeted deletion of the Abcb4 gene in the liver leads to more rapid Ch crystallization and gallstone formation in Abcb4 knockout (KO) mice fed even a low Ch diet compared to wild‐type (WT) mice.MethodsThe critical role of biliary phospholipids in Ch crystallization and gallstone formation was investigated by physical‐chemical methods in male Abcb4 KO vs WT mice fed a chow diet (on day 0) and a special diet containing 0.1% Ch, 15% butterfat and 0.25% cholic acid for 4 wk.ResultsOn the chow diet (day 0), biliary phospholipids are 18.1±0.3 mM in pooled gallbladder bile of WT mice. However, no phospholipids are detected in Abcb4 KO mice fed either the chow or the special diet. After 1 wk of feeding the special diet, phase diagram analysis finds that lipid composition of pooled gallbladder bile of Abcb4 KO mice is located in crystallization region A. Moreover, many needle‐like Ch crystals, typical Ch monohydrate crystals, sandy stones and small gallstones (0.1–0.2 mm in diameter), but not liquid crystals, are detected in bile by phase contrast and polarizing light microscopy. This shows that lack of phospholipids in bile produces anhydrous crystalline metastable intermediates that rapidly evolve to Ch monohydrate crystals from supersaturated bile. In contrast, lipid composition of pooled gallbladder bile of WT mice is still within the micellar zone, indicating that bile is unsaturated with Ch. After 4 wk of feeding the special diet, lipid composition of pooled gallbladder bile of Abcb4 KO mice still stays in crystallization region A, with many larger gallstones (0.3–0.6 mm in diameter) being formed in the gallbladder. Notably, lipid composition of pooled gallbladder bile of WT mice enters crystallization region C, as analyzed by phase diagram. Thus, many liquid crystals (i.e., unilamellar and multilamellar vesicles) and classical solid plate‐like Ch monohydrate crystals, but not gallstones, are found in the gallbladder of WT mice. These results indicate that solid Ch crystals are formed through the liquid crystalline pathway in WT mice.ConclusionsLack of biliary phospholipids caused by the Abcb4 deletion in the mouse liver is a critical risk factor for Ch gallstone disease by significantly reducing Ch solubility in bile through disrupting the liquid crystalline pathway, whereas it leads to more rapid Ch crystallization via the anhydrous crystalline pathway. Our findings provide novel insight into the pathophysiological mechanisms of why Ch gallstones are rapidly formed in patients with the ABCB4 mutations, thereby leading to low phospholipid‐associated cholelithiasis, a rare biliary disease caused by a single‐gene mutation.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.