Hepatic Mitochondrial Defects in a Nonalcoholic Fatty Liver Disease Mouse Model Are Associated with Increased Degradation of Oxidative Phosphorylation Subunits

Abstract

Nonalcoholic fatty liver disease (NAFLD) is associated with hepatic mitochondrial dysfunction characterized by reduced ATP synthesis. We applied the ²H₂O-metabolic labeling approach to test the hypothesis that the reduced stability of oxidative phosphorylation proteins contributes to mitochondrial dysfunction in a diet-induced mouse model of NAFLD. A high fat diet containing cholesterol (a so-called Western diet (WD)) led to hepatic oxidative stress, steatosis, inflammation and mild fibrosis, all markers of NAFLD, in low density cholesterol (LDL) receptor deficient (LDLR^-/-) mice. In addition, compared with controls (LDLR^-/- mice on normal diet), livers from NAFLD mice had reduced citrate synthase activity and ATP content, suggesting mitochondrial impairment. Proteome dynamics study revealed that mitochondrial defects are associated with reduced average half-lives of mitochondrial proteins in NAFLD mice (5.41 ± 0.46 versus 5.15 ± 0.49 day, p < 0.05). In particular, the WD reduced stability of oxidative phosphorylation subunits, including cytochrome b-c1 complex subunit 1 (5.9 ± 0.1 versus 3.4 ± 0.8 day), ATP synthase subunit α (6.3 ± 0.4 versus 5.5 ± 0.4 day) and ATP synthase F(0) complex subunit B1 of complex V (8.5 ± 0.6 versus 6.5 ± 0.2 day) (p < 0.05). These changes were associated with impaired complex III and F0F1-ATP synthase activities. Markers of mitophagy were increased, but proteasomal degradation activity were reduced in NAFLD mice liver, suggesting that ATP deficiency because of reduced stability of oxidative phosphorylation complex subunits contributed to inhibition of ubiquitin-proteasome and activation of mitophagy. In conclusion, the ²H₂O-metabolic labeling approach shows that increased degradation of hepatic oxidative phosphorylation subunits contributed to mitochondrial impairment in NAFLD mice.

Keywords: Energy metabolism; Mass Spectrometry; Mitochondria function or biology; Oxidative stress; Protein Degradation; Protein Turnover; heavy water; metabolic labeling; oxidative phosphorylation.