title: Metformin activates AMP-activated protein kinase in primary human hepatocytes by decreasing cellular energy status.
abstract: OBJECTIVE
  The glucose-lowering drug metformin has been shown to activate hepatic AMP-activated protein kinase (AMPK), a master kinase regulating cellular energy homeostasis. However, the underlying mechanisms remain controversial and have never been investigated in primary human hepatocytes.
  
  
  METHODS
  Hepatocytes isolated from rat, mouse and human livers were treated with various concentrations of metformin. Isoform-specific AMPKα abundance and activity, as well as intracellular adenine nucleotide levels and mitochondrial oxygen consumption rates were determined at different time points.
  
  
  RESULTS
  Metformin dose- and time-dependently increased AMPK activity in rat and human hepatocytes, an effect associated with a significant rise in cellular AMP:ATP ratio. Surprisingly, we found that AMPKα2 activity was undetectable in human compared with rat hepatocytes, while AMPKα1 activities were comparable. Accordingly, metformin only increased AMPKα1 activity in human hepatocytes, although both AMPKα isoforms were activated in rat hepatocytes. Analysis of mRNA expression and protein levels confirmed that only AMPKα1 is present in human hepatocytes; it also showed that the distribution of β and γ regulatory subunits differed between species. Finally, we demonstrated that the increase in AMP:ATP ratio in hepatocytes from liver-specific Ampkα1/2 (also known as Prkaa1/2) knockout mice and humans is due to a similar and specific inhibition of the mitochondrial respiratory-chain complex 1 by metformin.
  
  
  CONCLUSIONS
  Activation of hepatic AMPK by metformin results from a decrease in cellular energy status owing to metformin's AMPK-independent inhibition of the mitochondrial respiratory-chain complex 1. The unique profile of AMPK subunits found in human hepatocytes should be considered when developing new pharmacological agents to target the kinase.
authors.affiliation: Laboratory of Paediatric Hepatology and Cell Therapy, Université catholique de Louvain and Cliniques St Luc, Brussels, Belgium.
authors.forename: X
authors.initials: X
authors.lastname: Stephenne
authors.forename: M
authors.initials: M
authors.lastname: Foretz
authors.forename: N
authors.initials: N
authors.lastname: Taleux
authors.forename: G C
authors.initials: GC
authors.lastname: van der Zon
authors.forename: E
authors.initials: E
authors.lastname: Sokal
authors.forename: L
authors.initials: L
authors.lastname: Hue
authors.forename: B
authors.initials: B
authors.lastname: Viollet
authors.forename: B
authors.initials: B
authors.lastname: Guigas
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000032:analysis / Q000187:drug effects*; D000227:Adenine Nucleotides / Q000032:analysis; D000818:Animals; D002478:Cells, Cultured; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology; D006801:Humans; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D008687:Metformin / Q000494:pharmacology*; D051379:Mice; D018345:Mice, Knockout; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects; D051381:Rats
chemical_list: D000227:Adenine Nucleotides; D007004:Hypoglycemic Agents; D007527:Isoenzymes; D008687:Metformin; D055372:AMP-Activated Protein Kinases