Figure 1.
Targeting of hOGG1 to mitochondria in L6 myotubes protected against palmitate-induced mitochondrial protein carbonylation.
MTS-hOGG1 or GFP transduced L6 myotubes were exposed to control medium (C, 2% BSA) or medium containing 1 mM palmitate (P). (A). Mitochondrial fractions were isolated and analyzed by Western blot with the indicated antibodies. Lamin A was used as a marker for nuclear proteins, and subunit IV of mitochondrial complex IV as used as mitochondrial marker. No nuclear contamination was detected in the skeletal muscle cell mitochondria. The values for densitometry are presented as fold induction over corresponding control data and are the means ± SE. (B). (n > 3). *p < 0.05 vs all other groups.
Figure 2.
Mitochondrial hOGG1 protected against ER stress.
Treatment of L6 myotubes with 1 mM of palmitate (P) or 5 μg/ml of tunicamycin (T) for 24 h induced phosphorylation of both PERK and EIF2α (A). Treatment of L6 myotubes with tunicamycin (T) or palmitate (P) reduced insulin-stimulated phosphorylation of Akt (Ser473) (B). For both A and B, control medium (C, 2% BSA) was used. Targeting of hOGG1 to mitochondria in L6 myotubes ameliorated palmitate-mediated activation of ER stress markers. MTS-hOGG1 or GFP transduced L6 myotubes were exposed to control medium (C, 2% BSA) or medium containing 1 mM palmitate (P) for 24 h (C and D). Total cell lysates were isolated and analyzed by Western blot analysis with the indicated antibodies. Representative blots (C) and values of densitometry data (D) from at least three independent experiments are shown. The values for densitometry are presented as fold induction over corresponding control data normalized to the actin level. In addition, for phosphorylated proteins, the values from densitometry from three independent experiments for each phosphorylated protein were normalized to the level of corresponding total protein, and expressed as fold induction over corresponding control data normalized to the actin level. (n > 3). *p < 0.05 vs all other groups.
Figure 3.
Targeting of hOGG1 to mitochondria in L6 myotubes ameliorated palmitate-mediated activation of autophagy.
Representative blots (A) and values of densitometry (B) data ± SE are shown. The values for densitometry are presented as fold induction over corresponding control data normalized to the actin level. In addition, for phosphorylated proteins, the values from densitometry for each phosphorylated protein were normalized to the level of corresponding total protein, and expressed as fold induction over corresponding control data normalized to the actin level. (n > 3). *p < 0.05 vs all other groups.
Figure 4.
A schematic model of the proposed links between palmitate-induced mtDNA damage, mitochondrial dysfunction and ROS production, ER stress, autophagy and IR in L6 myotubes.