Figure 1.
Expression of hypoxia inducible factor 1 alpha (HIF-1α, panel A) and a canonical HIF-1 regulated gene, Glut-1 (panel B) in different tissues of C57BL/6J mice with diet induced obesity (DIO) treated with HIF-1α antisense oligonucleotides (HIF-1α ASO), control ASO or observed untreated for 8 weeks.
High fat diet was administered for 12 weeks prior to the ASO treatment and continued during the treatment. BAT, brown adipose tissue; EPI, epididymal adipose tissue; ING, inguinal adipose tissue; OM, omental adipose tissue. *, † and ‡ denote p<0.05, <0.01 and <0.001, respectively, for the difference with HIF-1α ASO treated mice.
Figure 2.
Western blot for HIF-1α in the livers (panel A) and epididymal fat (panel B) of mice with diet-induced obesity (DIO) treated with HIF-1α antisense oligonucleotides (ASO), control ASO (Con) or observed untreated (U) for 8 weeks.
N = 5 per group. Representative samples shown. High fat diet was administered for 12 weeks prior to the ASO treatment and continued during the treatment.
Figure 3.
The body weight trajectory of DIO mice treated with HIF-1α ASO, control ASO or observed untreated.
* denotes p<0.001 for the difference with HIF-1α ASO treated mice.
Table 1.
Basic Characteristics of DIO mice treated with Hif-1a ASO and control ASO compared to untreated animals.
Figure 4.
O2 consumption (VO2) has been analyzed every 16 min for 6 consecutive days in DIO mice treated with HIF-1α ASO, control ASO or observed untreated.
Shaded areas represent the 12 hr dark phase (DP) and unshaded areas represent the 12 hr light phase (LP). VO2 in HIF-1α ASO treated mice significantly exceeded VO2 in mice treated with control ASO and untreated mice, p<0.001. VO2 during the dark phase significantly exceeded those during the light phase, p<0.001.
Figure 5.
Energy Expenditure (EE, panel A) and Respiratory Exchange Ratio (RER, panel B) in DIO mice treated with HIF-1α ASO, control ASO or observed untreated during the dark and light phases.
* denotes p<0.001 for the difference with HIF-1α ASO treated mice; † denotes p<0.001 for the difference with the light phase.
Figure 6.
Fasting blood glucose (A) and serum insulin (B), leptin (C), adiponectin (D), fasting serum cholesterol (E) and triglycerides (F), liver cholesterol (G) and triglycerides (H) in DIO mice treated with HIF-1α ASO, control ASO or observed untreated.
* and † denote p<0.05 and 0.01, respectively, for the difference with HIF-1α ASO treated mice.
Figure 7.
The intraperitoneal glucose tolerance test (A) and insulin tolerance test (B) were performed in DIO mice treated with HIF-1α ASO, control ASO or observed untreated.
The hyperinsulinemic euglycemic clamp was performed in HIF-1α ASO treated and untreated mice (C, D). The baseline hepatic glucose output (C) was determined as the ratio of the [3H] glucose infusion rate to the specific activity of plasma glucose prior to the clamp. The hepatic glucose output during the clamp was determined as the difference between the ratio of the [3H] glucose infusion rate to the specific activity of plasma glucose and non-radioactive glucose infusion rate during last 30 min of the clamp. The whole body insulin sensitivity (D) was measured by glucose infusion rate during last 30 min of hyperinsulinemic euglycemic clamp (glucose levels were clamped at 100–125 mg/dl). * denotes p<0.001 for the difference between untreated or control ASO treated and HIF-1α ASO treated mice; † denotes p<0.001 for the difference between untreated and control ASO treated mice; ‡ denotes p<0.05 for the difference between untreated and HIF-1α ASO treated mice.
Figure 8.
Accumulation of glycogen during HIF-1α ASO treatment.
(A) Representative images of liver sections in HIF-1α ASO treated (upper panel), control ASO treated (middle panel) and untreated mice (bottom panel). Periodic acid Schiff (PAS) staining, x 400 original magnification. HIF-1α ASO treated mice showed intense PAS positive staining, which was absent in other groups. Control ASO treated and untreated mice show macrovesicular steatosis, which was attenuated in HIF-1α ASO treated mice. (B) Glycogen levels in livers measured biochemically (see Methods). * and † denote p<0.05 and <0.01, respectively, for the difference with HIF-1α ASO treated mice. (C) Western blots showing phosphorylation of glycogen synthase kinase (GSK) α in HIF-1α ASO treated, control ASO treated and untreated mice in representative samples. GAPDH, glyceraldehydes-3-phosphate dehydrogenase.
Figure 9.
Expression of selected genes of lipid (A) and carbohydrate (B) metabolism in livers of HIF-1α ASO treated, control ASO treated and untreated mice determined by real time reverse transcriptase PCR (RT-PCR).
ACC-1, acetyl coenzyme A carboxylase; HMGCoAR, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase; LPK, liver pyruvate kinase; PEPCK, phosphoenolpyruvate carboxykinase; PPAR, peroxisome proliferator-activated receptor; SCD, stearoyl coenzyme A desaturase; SREBP, sterol regulatory element binding protein. *, † and ‡ denote p<0.05, <0.01 and <0.001, respectively, for the difference with HIF-1α ASO treated mice.