Figure 1.
Body weight, fasting blood glucose, glucose tolerance test (GTT) and insulin tolerance test (ITT) of ten-week old ob/ob and lean mice.
Ob/ob mice were significantly heavier (A) with significantly higher fasting blood glucose (B) than lean mice. GTT (C) and ITT (D) were performed after 12–13 hours of fasting. Compared to lean mice, ob/ob mice displayed glucose intolerance and insulin resistance. Data are shown as mean ± SEM (n = 9 mice/group; ob/ob, closed bar/closed circle; lean mice, open bar/open circle). Analyses were performed using Students t-tests for body weight and fasting blood glucose; and two-way ANOVA for GTT and ITT. **, P<0.01; ***, P<0.001.
Figure 2.
Liver tissue sections (4 µm) were stained for haemotoxylin and eosin (H&E, x20 magnification).
Insets/sections (10 µm) were stained with Oil Red O (ORO, x40 magnification). Representative slides are displayed and ob/ob livers (left, n = 9) showed severe macrovesicular steatosis on both H&E and ORO. Lean livers (right, n = 9) showed no steatosis on H&E but ORO showed some patchy mild microvesicular steatosis.
Figure 3.
Mitochondrial function analysis of ob/ob and lean livers following different durations of cold ischaemia (hours).
A: Ob/ob livers demonstrated lower Complex I (CI) oxidative phosphorylation throughout cold ischaemia and were significantly lower from lean livers at earlier time points. B: CI+Complex II (CII) oxidative phosphorylation did not differ between groups. C: After 8 hours of cold ischaemia, ob/ob and lean livers showed significantly lower CI respiration to CII respiration ratio compared to 3 hours of cold ischaemia. Ob/ob livers also showed significantly lower CI/CII ratios compared to lean livers after 16 hours of cold ischaemia. Data are shown as mean ± SEM (n = 9 mice/group; ob/ob, closed circle; lean livers, open circle). Restricted maximum likelihood with post-hoc Tukey-Kramer analysis was performed. *, P<0.05 (vs. lean); #, P<0.05 (vs. 3 hours).
Figure 4.
Mitochondrial leak respiration and respiratory control ratios (RCR) of ob/ob and lean livers following different duration of cold ischaemia (hours).
A: Ob/ob livers showed increased mitochondrial leak rates with progressive cold ischaemia and increased significantly after 16 hours of cold ischaemia, while lean liver mitochondrial leak rates remained stable. B: Ob/ob livers consistently showed lower RCRs compared to lean livers and differed significantly after 16 hours of cold ischaemia. Ob/ob livers also demonstrated decreased RCR after 16 hours of cold ischaemia relative to 1.5 hours while lean liver RCR remained stable. Data are shown as mean ± SEM (n = 9 mice/group; ob/ob, closed circle; lean livers, open circle). Restricted maximum likelihood with post-hoc Tukey-Kramer analysis was performed. *, P<0.05 (vs. lean); #, P<0.01 (vs. 1.5 hours).
Figure 5.
Net adenosine triphosphate (ATP) production rate and ATP production rate relative to flux of ob/ob and lean livers following different duration of cold ischemia (hours).
Ob/ob livers showed significantly lower net ATP production (A) and ATP production rate relative to flux (B) at all time points. Data are shown as mean ± SEM (n = 6 mice/group; ob/ob livers, closed circle; lean livers, open circle). Analyses were performed using two-way ANOVA. *, P<0.05 (vs. lean).
Table 1.
Baseline measurement of ADP and ATP contents; Complex I, Complex II, Citrate Synthase and lactate dehydrogenase levels in ob/ob and lean livers.
Figure 6.
University of Wisconsin (UW) solution and tissue pH of ob/ob and lean livers during cold ischemia; and baseline intrinsic tissue buffering capacity of ob/ob and lean livers.
A: UW solution pH (ob/ob, closed square; lean, open square) remained stable during the whole duration of cold ischemia and did not differ between groups. However, ob/ob livers demonstrated significantly lower tissue pH following 8–16 hours of cold ischemia compared to lean livers. Ob/ob and lean livers demonstrated decreased tissue pH after 8 and 12 hours of cold ischemia, respectively, relative to 1.5 hours. B: Ob/ob livers also demonstrated significantly lower baseline tissue intrinsic pH buffering capacity. Data are shown as mean ± SEM (n = 9 mice/group; ob/ob, closed circle/bar; lean, open circle/bar). Analyses were performed using two-way ANOVA for tissue pH and un-paired t-test for intrinsic tissue buffering capacity. **, P<0.01 (vs. lean); ***, P<0.001 (vs. lean); #, P<0.01 (vs. 1.5 hours).