Unique Inhibition of Bile Salt-Induced Apoptosis by Lecithins and Cytoprotective Bile Salts in Immortalized Mouse Cholangiocytes

Abstract

Bile duct epithelium is physiologically exposed to high concentrations of bile salts, suggesting the presence of a cytoprotective mechanism(s). The aim of this study was to clarify whether bile salts cause bile duct cell damage and to elucidate the mechanism(s) providing protection against such an action of bile salts. Immortalized mouse cholangiocytes were incubated with taurocholate, taurochenodeoxycholate, glycochenodeoxycholate (GCDC), taurodeoxycholate, and tauroursodeoxycholate (TUDC), followed by flow-cytometric analysis and caspase activity assay to evaluate the induction of apoptosis. GCDC time-dependently induced caspase 3 (3.4-fold)- and caspase 9 (1.4-fold)-mediated apoptosis of cholangiocytes, but this was inhibited by lecithins and TUDC. Further, expression of cholangiocyte bile salt transporters (apical sodium-dependent bile salt transporter [Asbt] and multidrug resistance protein 3 [Mrp3]) was examined by RT-PCR and western blotting, and cholangiocyte bile salt uptake was determined using radiolabeled bile salts. Expression of cholangiocyte Asbt and Mrp3 was increased by bile salts, whereas lecithins interestingly reduced bile salt uptake to inhibit cholangiocyte apoptosis. In conclusion, bile salts themselves cause cholangiocyte apoptosis when absorbed by and retained inside the cell, but this is inhibited by washing out cytotoxic bile salts according to Mrp3, a rescue exporting molecule. Biliary lecithin is seemingly another cytoprotective player against cytotoxic bile salts, reducing their uptake, and this is associated with a reduced expression of Mrp3.

Taurocholate Potentiates Ethanol-Induced NF-κB Activation and Inhibits Caspase-3 Activity in Cultured Rat Gastric Mucosal Cells

“…At larger TC concentrations, the activation of plain caspase-3 enzyme was decreased, which might be caused by formation of micelles by TC. TC has been shown to decrease caspase activation in rat cholangiocyte after vagotomy [18] and in mouse cholangiocytes exposed to tauroursodeoxycholate [19] as well as in human cholangiocytes exposed to glycoursodeoxycholic acid [20]. In the case of ursodeoxycholic acids, these effects on caspase cascade are mainly due to downregulation of caspase precursors.…”

“…The cells were lysed by adding lysing buffer (pH 7.6) containing 150 mM NaCl, 10 mM TRIS-HCl, 1 mM EDTA, 1 mM EGTA, 1% TritonX-100, 0.5% NP-40, 19 Complete TM protease inhibitor cocktail (Roche, Mannheim, Germany) and 1 mM Pefabloc SC (Roche, Mannheim, Germany) and incubating them for 1 h at 0°C followed by short vortexing. Protein concentrations of the samples were determined by colorimetric Bradford assay and equal amounts of cytoplasmic protein extracts (20 lg) were diluted in Laemmli sample buffer with 5% mercaptoethanol.…”

A Biliary HCO3 − Umbrella Constitutes A Protective Mechanism Against Bile Acid–Induced Injury in Human Cholangiocytes

“…9 Uncontrolled, carrier-independent membrane traffic and cell invasion of bile salts is determined by their polarity and degree of protonation. 10 Glycine conjugates account for the majority of bile salts in human bile, have a pK a of $4, 11 and, at a physiologic pH of $7.4, are partially protonated, apolar, and thus cell permeable at micromolar amounts.…”

Identification of bile biomarkers of biliary tract cancer through a liquid chromatography/mass spectrometry-based metabolomic method

“…It is synthesized in the hepatocytes and is subsequently transported into the biliary canaliculus by flippase multidrug-resistant protein 3 (39). Phosphatidylcholine is cytoprotective towards the biliary epithelium and reduces the cellular toxicity of bile acids (40,41). The reduction of phosphatidylcholine in the bile exposes the biliary epithelium to 'toxic' bile and predisposes it to biliary malignancies (42).…”