Deletion of Kinin B2 Receptor Alters Muscle Metabolism and Exercise Performance.

Metabolic syndrome is a cluster of metabolic risk factors such as obesity, diabetes and cardiovascular diseases. Mitochondria is the main site of ATP production and its dysfunction leads to decreased oxidative phosphorylation, resulting in lipid accumulation and insulin resistance. Our group has demonstrated that kinins can modulate glucose and lipid metabolism as well as skeletal muscle mass. By using B2 receptor knockout mice (B2R-/-) we investigated whether kinin action affects weight gain and physical performance of the animals. Our results show that B2R-/- mice are resistant to high fat diet-induced obesity, have higher glucose tolerance as well as increased mitochondrial mass. These features are accompanied by higher energy expenditure and a lower feed efficiency associated with an increase in the proportion of type I fibers and intermediary fibers characterized by higher mitochondrial content and increased expression of genes related to oxidative metabolism. Additionally, the increased percentage of oxidative skeletal muscle fibers and mitochondrial apparatus in B2R-/- mice is coupled with a higher aerobic exercise performance. Taken together, our data give support to the involvement of kinins in skeletal muscle fiber type distribution and muscle metabolism, which ultimately protects against fat-induced obesity and improves aerobic exercise performance

Indirect calorimetry and mitochondrial activity.

<p><b>(A)</b> Oxygen consumption and <b>(B)</b> CO₂ production in WT or B2R^-/- mice fed an HFD for 8 weeks (n = 5 each). <b>(C)</b> Respiratory quotient ratio in WT or B2R^-/- mice fed an HFD for 8 weeks. (n = 5 each). <b>(D)</b> Histochemical analysis of SDH and COX staining in skeletal muscle of 3-month-old WT or B2R^-/- mice (n = 5) Scale bar = 50μm. <b>(E)</b> Dihydroethidium (DHE) and 4’,6-diamidino-2-phenylindole dihydrochloride (DAPI) colocalization. Representative images of gastrocnemius muscle in 3-month-old mouse showing fluorescent labeling of DAPI (blue) in and DHE (red) at the corresponding location and focal plane, with merged pictures, showing colocalization (pink). Scale bar = 20μm. <i>(*P < 0.05;</i><i>**P<0.005)</i><i>Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p

B2R^-/- is protected from obesity.

<p><b>(A)</b> Total body weight of wild type (WT) or B2 knockout mice (B2R^-/-) under control diet (Control) or high-fat diet (HFD) (<i>n</i> = 5 each). Obesity was induced after 8 weeks of HFD treatment. <b>(B)</b> Feed efficiency of WT and B2R^-/- mice during 8 weeks of HFD treatment. Body composition analysis showing <b>(C)</b> fat mass, <b>(D)</b> lean mass and percent body fat <b>(E)</b> of WT and mice after 8 weeks of HFD treatment. <b>(F)</b> Epididmal and <b>(G)</b> perirenal fat pad depots and <b>(H)</b> total mass after 8 weeks of HFD treatment. (<i>n</i> = 5 for each group) <i>(*P<0.05;</i>^<i>#</i><i>P<0.05</i>, ^<i>##</i><i>P<0.01) Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p

Fiber type characterization in skeletal muscle.

<p><b>(A</b>) Example of transverse soleus sections from 3-month-old WT and B2R^-/- mice, with histochemical staining for myosin ATPase, preincubated at pH 10.3. Type I fibers; type IIA fibers and intermediary fiber are indicated. <b>(B)</b> Soleus fiber type percentage. <b>(C)</b> Media cross-sectional area (CSA) of soleus muscle from WT and B2R^-/-. (<i>n</i> = 5 each). <i>(*P < 0</i>.<i>05) Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p

Exercise performance.

<p><b>(A)</b> Maximum VO₂ and <b>(B)</b> time to exhaustion of WT or B2R^-/- mice on a metabolic treadmill. <b>(C)</b> Time to fatigue after swimming test and <b>(D)</b> percentage of swimmers in 3 month-old WT and B2R^-/- mice (n = 5 each). (<i>**P<0</i>.<i>005) Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p

Leptin and Insulin levels, GTT and ITT and skeletal muscle glucose uptake.

<p>Serum leptin <b>(A)</b> and insulin <b>(B)</b> levels measured after 8 weeks of HFD treatment in WT and B2R^-/- mice under control (Control) or high-fat diet (HFD). <b>(C)</b> Glucose tolerance test of WT and B2R^-/- under control diet and <b>(D)</b> high-fat diet <b>(E)</b> Insulin tolerance test of WT and B2R^-/- under control diet and <b>(F)</b> high-fat diet <b>(G)</b> Glucose uptake and <b>(H)</b> glycogen synthesis in skeletal muscle of 3-month-old WT and B2R^-/- mice incubated without (filled bars) or with (open bars) 10,000 μU/mL of insulin for 60 min. (<i>n</i> = 5 for each group). <i>(*P < 0</i>.<i>05; **P<0</i>.<i>005</i>, <i>***P<0</i>.<i>001) Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p

Mitochondrial biogenesis.

<p><b>(A)</b> Mitochondrial DNA quantification in skeletal muscle of 3-month-old WT or B2R^-/- mice (n = 5 each). <b>(B)</b> Expression of genes related to mitochondrial biogenesis (PGC1b; PGC1a; LCAD; SCD1; UCP3; PPARg) and OxPhos (CPT) in skeletal muscle of 3-month-old WT or B2R^-/- mice (<i>n</i> = 7 each). <i>(*P < 0</i>.<i>05; **P<0</i>.<i>005) Data are presented as mean ± S</i>.<i>E</i>.<i>M</i>.</p