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

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.

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.

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.

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.