ObjectiveWe explored the hypothesis that TGR5, the bile acid (BA) G-protein-coupled receptor highly expressed in biliary epithelial cells, protects the liver against BA overload through the regulation of biliary epithelium permeability.DesignExperiments were performed under basal and TGR5 agonist treatment. In vitro transepithelial electric resistance (TER) and FITC-dextran diffusion were measured in different cell lines. In vivo FITC-dextran was injected in the gallbladder (GB) lumen and traced in plasma. Tight junction proteins and TGR5-induced signalling were investigated in vitro and in vivo (wild-type [WT] and TGR5-KO livers and GB). WT and TGR5-KO mice were submitted to bile duct ligation or alpha-naphtylisothiocyanate intoxication under vehicle or TGR5 agonist treatment, and liver injury was studied.ResultsIn vitro TGR5 stimulation increased TER and reduced paracellular permeability for dextran. In vivo dextran diffusion after GB injection was increased in TGR5-knock-out (KO) as compared with WT mice and decreased on TGR5 stimulation. In TGR5-KO bile ducts and GB, junctional adhesion molecule A (JAM-A) was hypophosphorylated and selectively downregulated among TJP analysed. TGR5 stimulation induced JAM-A phosphorylation and stabilisation both in vitro and in vivo, associated with protein kinase C-ζ activation. TGR5 agonist-induced TER increase as well as JAM-A protein stabilisation was dependent on JAM-A Ser285 phosphorylation. TGR5 agonist-treated mice were protected from cholestasis-induced liver injury, and this protection was significantly impaired in JAM-A-KO mice.ConclusionThe BA receptor TGR5 regulates biliary epithelial barrier function in vitro and in vivo through an impact on JAM-A expression and phosphorylation, thereby protecting liver parenchyma against bile leakage.

Endothelin-1 (ET-1), platelet-derived growth factor (PDGF), and epidermal growth factor (EGF) stimulated thymidine incorporation with different efficiency (PDGF >> EGF = ET-1) in rat myometrial cells. They also stimulated ERK activation, which culminated at 5 min and then declined to reach a plateau (at 45 min: EGF > 90%, PDGF = 50%, and ET-1 < 10% of maximum). Inhibition and downregulation of PKC demonstrated that ERK activation at 5 min involved PKC delta and -zeta for ET-1 and PKC alpha plus another PKC isoform for PDGF. By contrast, the EGF response did not involve PKC. Stimulation of Ras was more important with EGF than with PDGF, with ET-1 being the weakest activator. The simultaneous incubation of the cells with EGF and ET-1 potentiated the ERK activation at 5 min and mimicked the plateau phase obtained with PDGF. Under these conditions thymidine incorporation was comparable to that induced by PDGF. Taken together, our results indicated that the kinetic profile of ERK activation and its impact on cell proliferation can be modulated by the differential involvement of PKC isoforms and the amplitude of Ras activation.