Lipoatrophic diabetes in Irs1(-/-)/Irs3(-/-) double knockout mice - PubMed (original) (raw)
Lipoatrophic diabetes in Irs1(-/-)/Irs3(-/-) double knockout mice
Palle G Laustsen et al. Genes Dev. 2002.
Abstract
Based on the phenotypes of knockout mice and cell lines, as well as pathway-specific analysis, the insulin receptor substrates IRS-1, IRS-2, IRS-3, and IRS-4 have been shown to play unique roles in insulin signal transduction. To investigate possible functional complementarity within the IRS family, we generated mice with double knockout of the genes for IRS-1/IRS-3 and IRS-1/IRS-4. Mice with a combined deficiency of IRS-1 and IRS-4 showed no differences from Irs1(-/-) mice with respect to growth and glucose homeostasis. In contrast, mice with a combined deficiency of IRS-1 and IRS-3 developed early-onset severe lipoatrophy associated with marked hyperglycemia, hyperinsulinemia, and insulin resistance. However, in contrast to other models of lipoatrophic diabetes, there was no accumulation of fat in liver or muscle. Furthermore, plasma leptin levels were markedly decreased, and adenovirus-mediated expression of leptin in liver reversed the hyperglycemia and hyperinsulinemia. The results indicate that IRS-1 and IRS-3 play important complementary roles in adipogenesis and establish the Irs1(-/-)/Irs3(-/-) double knockout mouse as a novel model of lipoatrophic diabetes.
Figures
Figure 1
Irs1−/−/Irs3−/− double knockout mice are lipoatrophic. (A) Growth curves. Body weights for wild-type, Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout male mice were determined at indicated time points between the ages of 4 and 16 wk. Growth curves for female mice followed a similar pattern (data not shown). (B) Photographs of epididymal and interscapular fat pads isolated from wild-type (WT), Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout mice (left panel). Sections of white adipose tissue (WAT) are shown at 40× magnification (middle panel) and of brown adipose tissue (BAT) at 10× magnification (right panel). (C) Perigonadal fat pad weight as percentage of total body weight. Data in C_–_E are shown as means ± S.E. (D) Interscapular fat pad weight as percentage of total body weight. (E) Total body triglyceride content as percentage of total body weight. (F) Cell size distribution in perigonadal fat pads was analyzed from tissue sections as described in Materials and Methods. (G) Reduced expression of PPARγ in epididymal fat pads. Northern blot of total RNA isolated from epididymal fat pads probed with a 32P-labeled PPARγ cDNA fragment. Each lane represents a single animal.
Figure 1
Irs1−/−/Irs3−/− double knockout mice are lipoatrophic. (A) Growth curves. Body weights for wild-type, Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout male mice were determined at indicated time points between the ages of 4 and 16 wk. Growth curves for female mice followed a similar pattern (data not shown). (B) Photographs of epididymal and interscapular fat pads isolated from wild-type (WT), Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout mice (left panel). Sections of white adipose tissue (WAT) are shown at 40× magnification (middle panel) and of brown adipose tissue (BAT) at 10× magnification (right panel). (C) Perigonadal fat pad weight as percentage of total body weight. Data in C_–_E are shown as means ± S.E. (D) Interscapular fat pad weight as percentage of total body weight. (E) Total body triglyceride content as percentage of total body weight. (F) Cell size distribution in perigonadal fat pads was analyzed from tissue sections as described in Materials and Methods. (G) Reduced expression of PPARγ in epididymal fat pads. Northern blot of total RNA isolated from epididymal fat pads probed with a 32P-labeled PPARγ cDNA fragment. Each lane represents a single animal.
Figure 1
Irs1−/−/Irs3−/− double knockout mice are lipoatrophic. (A) Growth curves. Body weights for wild-type, Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout male mice were determined at indicated time points between the ages of 4 and 16 wk. Growth curves for female mice followed a similar pattern (data not shown). (B) Photographs of epididymal and interscapular fat pads isolated from wild-type (WT), Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout mice (left panel). Sections of white adipose tissue (WAT) are shown at 40× magnification (middle panel) and of brown adipose tissue (BAT) at 10× magnification (right panel). (C) Perigonadal fat pad weight as percentage of total body weight. Data in C_–_E are shown as means ± S.E. (D) Interscapular fat pad weight as percentage of total body weight. (E) Total body triglyceride content as percentage of total body weight. (F) Cell size distribution in perigonadal fat pads was analyzed from tissue sections as described in Materials and Methods. (G) Reduced expression of PPARγ in epididymal fat pads. Northern blot of total RNA isolated from epididymal fat pads probed with a 32P-labeled PPARγ cDNA fragment. Each lane represents a single animal.
Figure 2
Reduced leptin, free fatty acid, and triglyceride plasma levels in Irs1−/−/Irs3−/− double knockout mice. (A) Plasma leptin levels were measured in random-fed mice (n = 3–5). *, P < 0.05 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. #, P < 0.05 for Irs1−/− versus WT mice. (B) Plasma leptin levels in mice after overnight fast (n = 2–5). *, P < 0.05 for Irs1−/−/Irs3−/− versus Irs1−/− mice. (C) Plasma triglyceride (TG) levels after overnight fast (n = 6–14). *, P < 0.05 for Irs1−/−/Irs3−/− versus WT and Irs3−/− mice. #, P < 0.01 for Irs1−/− versus WT mice. †, P < 0.01 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. (D) Plasma free fatty acid levels after overnight fast (n = 3–5). *, P < 0.05 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. #, P < 0.05 for Irs1−/− versus WT mice. †, P < 0.01 for Irs1−/−/Irs3−/− versus WT mice. In A_–_D, data are shown as means ± S.E. for the indicated number of animals. All data were from 2-month-old mice. (E) Liver and skeletal muscle triglyceride content. The total glycerol content of liver and skeletal muscle was determined after incubation of tissue homogenates with lipoprotein lipase. Values were converted into milligrams of triglyceride per gram of tissue (wet weight) after comparison with a glycerol standard (Sigma). Liver and skeletal muscle were removed from 4-month-old, random-fed male mice (n = 6 in each group). Data are shown as means ± S.E.
Figure 3
Irs1−/−/Irs3−/− double knockout mice are hyperglycemic and hyperinsulinemic. (A) Blood glucose levels in 2-month-old random-fed mice (n = 6–11 in each group). **, P < 0.001 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. *, P < 0.05 for Irs1−/− versus WT and Irs3−/− mice. (B) Blood glucose in 2-month-old mice after overnight fast (n = 6–14). **, P < 0.001 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. *, P < 0.05 for Irs1−/−/Irs3−/− versus WT, Irs3−/− mice. #, P < 0.05 for Irs1−/− versus WT and Irs3−/− mice. (C) Plasma insulin levels in 2-month-old random-fed mice (n = 6–14). **, P < 0.001 for Irs1−/−/Irs3−/− versus WT and Irs3−/− mice. P < 0.05 for Irs1−/−/Irs3−/− versus Irs1−/− mice. #, P < 0.01 for Irs1−/− versus WT and Irs3−/− mice. (D) Plasma insulin levels in 2-month-old mice after overnight fast (n = 6–14). *, P < 0.05 for Irs1−/−/Irs3−/− versus WT and Irs3−/− mice; for male mice, also versus Irs1−/− mice. In A_–_D, data are shown as means ± S.E. for the indicated number of mice.
Figure 4
Irs1−/−/Irs3−/− double knockout mice are insulin-resistant and glucose-intolerant. (A,B) Glucose tolerance tests. Glucose levels were determined in 6-week-old male (A) and female (B) mice immediately before and at the indicated time points after intraperitoneal injection of glucose (2 g/kg body weight). Each point represents the mean ± S.E. for 5–8 animals. (C,D) Insulin tolerance tests. Glucose levels were determined in 6-week-old male (C) and female (D) mice immediately before and at the indicated time points after intraperitoneal injection of human insulin (1 U/kg body weight). Each point represents the mean ± S.E. for 6–8 animals.
Figure 5
Irs1−/−/Irs4−/− mice and Irs1−/− mice show similar responses to glucose tolerance and insulin tolerance tests. (A,B) Glucose tolerance tests. Glucose levels were determined in 9–12-week-old male (A) and female (B) mice immediately before and at the indicated time points after intraperitoneal injection of glucose (2 g/kg body weight). Each point represents the mean ± S.E. for seven animals. (C,D) Insulin tolerance tests. Glucose levels were determined in 9–12-week-old male (C) and female (D) mice immediately before and at the indicated time points after intraperitoneal injection of human insulin (0.75 U/kg body weight). Each point represents the mean ± S.E. for 6–8 animals.
Figure 6
Irs1−/−/Irs3−/− double knockout mice show β-cell hyperplasia. (A) β-cell mass was determined as described in Materials and Methods. For each group, n = 4–5 males, except for the Irs1−/−/Irs3−/− group, in which n = 4 (2 males and 2 females). Data are shown as means ± SE. *, P < 0.05 for Irs1−/−/Irs3−/− versus WT, Irs1−/−, and Irs3−/− mice. **, P < 0.001 for Irs1−/− versus WT and Irs3−/− mice. (B) Representative sections from pancreas from 2-month-old WT, Irs1−/−, Irs3−/−, and Irs1−/−/Irs3−/− double knockout male mice. Sections were stained with a cocktail of antibodies against the non-β-cells and counterstained with hematoxylin as described in Materials and Methods. Magnification, 10×. Bar, 50 μm.
Figure 7
Reversal of diabetes by leptin in Irs1−/−/Irs3−/− double knockout mice. (A,B) Blood glucose levels (A) and plasma insulin levels (B) were determined in wild-type, Irs1−/−/Irs3−/− double knockout, and ob/ob mice 6 d after injection of 5 × 108 PFU/g body weight of adenovirus carrying cDNA coding for either leptin (Ad-Leptin) or β-galactosidase (Ad-LacZ). Each bar represents the mean ± S.E. for four animals, except for the bar representing ob/ob mice injected with Ad-LacZ, in which n = 3. *, P < 0.05 Ad-Leptin-treated mice versus corresponding Ad-LacZ-treated mice.
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