Role of AMP-activated protein kinase α1 in 17α-ethinylestradiol-induced cholestasis in rats

Arch Toxicol. 2017 Jan;91(1):481-494. doi: 10.1007/s00204-016-1697-8. Epub 2016 Apr 18.

Abstract

Estrogen-induced cholestasis occurs in many women who are susceptible due to pregnancy or hormone replacement therapy for postmenopausal syndrome. 17α-Ethinylestradiol (EE), as a synthetic estrogen, has been widely used to study the underlying mechanisms of estrogen-induced cholestasis. Recent studies have also reported that liver kinase B1 (LKB1)-mediated activation of AMP-activated protein kinase (AMPK) plays a critical role in the regulation of canalicular network formation. However, the role of AMPK in EE-induced cholestasis remains to be determined. In this study, the effects of EE (1-100 µM) on AMPK activation and the expression of farnesoid X receptor (FXR) and hepatic bile acid transporters were examined in in vitro using 3D-cultured rat primary hepatocytes and in in vivo using rat cholestasis models. We also used specific chemical agonist and antagonist of AMPK, AMPK subunit-specific antibodies and lentiviral shRNAs for AMPKα1 and AMPKα2 to delineate the role of AMPK in EE-induced cholestasis and potential cellular mechanisms. We found that EE-induced phosphorylation of AMPKα1 via extracellular signal-regulated kinases-LKB1-mediated signaling pathways and subsequent nuclear translocation accounted for the down-regulation of FXR and bile acid transporters and disruption of bile acid homeostasis. Inhibition of AMPK activation using an AMPK antagonist Compound C (2 µM) or down-regulation of AMPKα1 using gene-specific shRNA attenuated EE-induced cholestasis both in in vitro and in in vivo. In conclusion, these results revealed that activation of cAMP-ERK-LKB1-AMPKα1 signaling pathway plays a critical role in EE-mediated dysregulation of the expression of FXR and bile acid transporters. AMPKα1 may represent an important therapeutic target for estrogen-induced cholestasis.

Keywords: AMPKα1; Bile acid transporters; Cholestasis; Estrogen; FXR.

MeSH terms

  • AMP-Activated Protein Kinases / antagonists & inhibitors
  • AMP-Activated Protein Kinases / chemistry
  • AMP-Activated Protein Kinases / genetics
  • AMP-Activated Protein Kinases / metabolism*
  • ATP Binding Cassette Transporter, Subfamily B, Member 11
  • ATP-Binding Cassette Transporters / antagonists & inhibitors
  • ATP-Binding Cassette Transporters / genetics
  • ATP-Binding Cassette Transporters / metabolism
  • Active Transport, Cell Nucleus / drug effects
  • Animals
  • Bile Acids and Salts / metabolism
  • Cells, Cultured
  • Cholestasis / chemically induced*
  • Cholestasis / enzymology
  • Cholestasis / pathology
  • Cholestasis / prevention & control
  • Cyclic AMP / metabolism
  • Disease Models, Animal*
  • Enzyme Activation / drug effects
  • Estrogens / adverse effects*
  • Estrogens / chemistry
  • Ethinyl Estradiol / adverse effects*
  • Ethinyl Estradiol / antagonists & inhibitors
  • Hepatocytes / drug effects*
  • Hepatocytes / metabolism
  • Hepatocytes / pathology
  • Male
  • Phosphorylation / drug effects
  • Protein Kinase Inhibitors / pharmacology
  • Protein Kinase Inhibitors / therapeutic use
  • Protein Processing, Post-Translational / drug effects
  • RNA Interference
  • Random Allocation
  • Rats, Sprague-Dawley
  • Receptors, Cytoplasmic and Nuclear / antagonists & inhibitors
  • Receptors, Cytoplasmic and Nuclear / genetics
  • Receptors, Cytoplasmic and Nuclear / metabolism
  • Second Messenger Systems / drug effects

Substances

  • ATP Binding Cassette Transporter, Subfamily B, Member 11
  • ATP-Binding Cassette Transporters
  • Abcb11 protein, rat
  • Bile Acids and Salts
  • Estrogens
  • Protein Kinase Inhibitors
  • Receptors, Cytoplasmic and Nuclear
  • farnesoid X-activated receptor
  • Ethinyl Estradiol
  • Cyclic AMP
  • Prkaa1 protein, rat
  • AMP-Activated Protein Kinases