Indirubin and NAD + prevent mitochondrial ischaemia/reperfusion damage in fatty livers

Eur J Clin Invest. 2018 Jun;48(6):e12932. doi: 10.1111/eci.12932. Epub 2018 Apr 16.

Abstract

Background: Fatty livers are considerably more susceptible to acute stressors, such as ischaemia/reperfusion (I/R). As the incidence of I/R is high due to surgical events and some pathologies, there is an urgent need to find strategies against I/R injury (I/RI) in fatty livers. We postulate that an acute pretreatment with indirubin-3'-oxime (Ind) or NAD+ prevents mitochondrial dysfunction associated with warm I/RI in fatty livers.

Materials and methods: Zucker fatty rats were subjected to warm ischaemia and 12 hours of reperfusion. Ind or NAD+ was administered in the hepatic artery 30 minutes before ischaemia. Hepatic mitochondrial isolation was performed, and functional assays as well as molecular analysis were performed.

Results: Pretreatment decreased markers of liver injury while preserving mitochondrial cytochrome c content, which is related to the prevention of calcium-induced mitochondrial permeability transition (mPT), the decline in mitochondrial respiratory state 3 and ATP content. The generation of reactive oxygen species (ROS) was also diminished. Inhibition of GSK-3ß by Ind resulted in the prevention of cyclophilin-D (CypD) phosphorylation, unabling it to bind to the adenine nucleotide translocator (ANT), thus, preventing mPT induction. Furthermore, deacetylation of CypD at Lys residue by sirtuin 3 (SIRT3) caused its dissociation from ANT, contributing to an increase in mPT threshold in NAD+ -pretreated animals.

Conclusions: Pretreatment with Ind or NAD+ protects fatty livers by maintaining mitochondrial calcium homoeostasis, thus, preserving mitochondrial function and energetic balance. As such, CypD might be a new protective target against I/RI in fatty livers.

Keywords: NAD+; bioenergetics; hepatic steatosis; indirubin-3′-oxime; mitochondrial dysfunction; permeability transition.

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Animals
  • Antibiotics, Antineoplastic / pharmacology*
  • Calcium / metabolism
  • Cyclophilin D
  • Cyclophilins / drug effects
  • Cyclophilins / metabolism
  • Cytochromes c / drug effects
  • Cytochromes c / metabolism
  • Fatty Liver / metabolism*
  • Fatty Liver / pathology
  • Glycogen Synthase Kinase 3 beta / antagonists & inhibitors
  • Hepatic Artery
  • Indoles / pharmacology*
  • Liver / drug effects*
  • Liver / metabolism
  • Liver / pathology
  • Mitochondria, Liver / drug effects*
  • Mitochondria, Liver / metabolism
  • Mitochondrial ADP, ATP Translocases / metabolism
  • NAD / pharmacology*
  • Oximes / pharmacology*
  • Rats
  • Rats, Zucker
  • Reactive Oxygen Species / metabolism
  • Reperfusion Injury / metabolism*
  • Sirtuins / metabolism
  • Warm Ischemia*

Substances

  • Antibiotics, Antineoplastic
  • Cyclophilin D
  • Indoles
  • Oximes
  • Reactive Oxygen Species
  • SIRT3 protein, rat
  • indirubin-3'-monoxime
  • NAD
  • Adenosine Triphosphate
  • Cytochromes c
  • Mitochondrial ADP, ATP Translocases
  • Glycogen Synthase Kinase 3 beta
  • Sirtuins
  • Cyclophilins
  • Calcium
  • indirubin