title: Rho-kinase/myosin light chain kinase pathway plays a key role in the impairment of bile canaliculi dynamics induced by cholestatic drugs.
abstract: Intrahepatic cholestasis represents a frequent manifestation of drug-induced liver injury; however, the mechanisms underlying such injuries are poorly understood. In this study of human HepaRG and primary hepatocytes, we found that bile canaliculi (BC) underwent spontaneous contractions, which are essential for bile acid (BA) efflux and require alternations in myosin light chain (MLC2) phosphorylation/dephosphorylation. Short exposure to 6 cholestatic compounds revealed that BC constriction and dilation were associated with disruptions in the ROCK/MLCK/myosin pathway. At the studied concentrations, cyclosporine A and chlorpromazine induced early ROCK activity, resulting in permanent MLC2 phosphorylation and BC constriction. However, fasudil reduced ROCK activity and caused rapid, substantial and permanent MLC2 dephosphorylation, leading to BC dilation. The remaining compounds (1-naphthyl isothiocyanate, deoxycholic acid and bosentan) caused BC dilation without modulating ROCK activity, although they were associated with a steady decrease in MLC2 phosphorylation via MLCK. These changes were associated with a common loss of BC contractions and failure of BA clearance. These results provide the first demonstration that cholestatic drugs alter BC dynamics by targeting the ROCK/MLCK pathway; in addition, they highlight new insights into the mechanisms underlying bile flow failure and can be used to identify new predictive biomarkers of drug-induced cholestasis.
authors.affiliation: INSERM U991, Liver Metabolisms and Cancer, Rennes, France.|Rennes 1 University, Rennes, France.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Sharanek
authors.affiliation: INSERM U991, Liver Metabolisms and Cancer, Rennes, France.|Rennes 1 University, Rennes, France.
authors.forename: Audrey
authors.initials: A
authors.lastname: Burban
authors.affiliation: INSERM U991, Liver Metabolisms and Cancer, Rennes, France.|Rennes 1 University, Rennes, France.
authors.forename: Matthew
authors.initials: M
authors.lastname: Burbank
authors.affiliation: ImPACcell platform, Biosit, Rennes 1 University, Rennes, France.
authors.forename: Rémy
authors.initials: R
authors.lastname: Le Guevel
authors.affiliation: Biopredic International, St Grégoire, France.
authors.forename: Ruoya
authors.initials: R
authors.lastname: Li
authors.affiliation: INSERM U991, Liver Metabolisms and Cancer, Rennes, France.|Rennes 1 University, Rennes, France.
authors.forename: André
authors.initials: A
authors.lastname: Guillouzo
authors.affiliation: INSERM U991, Liver Metabolisms and Cancer, Rennes, France.|Rennes 1 University, Rennes, France.|Biopredic International, St Grégoire, France.
authors.forename: Christiane
authors.initials: C
authors.lastname: Guguen-Guillouzo
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D001647:Bile Acids and Salts / Q000378:metabolism; D001648:Bile Canaliculi / Q000187:drug effects* / Q000502:physiology; D024722:Cardiac Myosins / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002746:Chlorpromazine / Q000494:pharmacology*; D016572:Cyclosporine / Q000494:pharmacology*; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence; D018994:Myosin Light Chains / Q000378:metabolism*; D024462:Myosin Type II / Q000378:metabolism; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D059008:Time-Lapse Imaging; D062826:Zonula Occludens-1 Protein / Q000378:metabolism; D054460:rho-Associated Kinases / Q000378:metabolism*
chemical_list: D001647:Bile Acids and Salts; D018994:Myosin Light Chains; D062826:Zonula Occludens-1 Protein; C107184:myosin light chain 2; D016572:Cyclosporine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D054460:rho-Associated Kinases; D009219:Myosin-Light-Chain Kinase; D024722:Cardiac Myosins; D024462:Myosin Type II; C049347:fasudil; D002746:Chlorpromazine