Oxidative stress stimulates skeletal muscle glucose uptake through a phosphatidylinositol 3-kinase-dependent pathway - PubMed (original) (raw)
Oxidative stress stimulates skeletal muscle glucose uptake through a phosphatidylinositol 3-kinase-dependent pathway
Yasuki Higaki et al. Am J Physiol Endocrinol Metab. 2008 May.
Abstract
We determined the acute effects of oxidative stress on glucose uptake and intracellular signaling in skeletal muscle by incubating muscles with reactive oxygen species (ROS). Xanthine oxidase (XO) is a superoxide-generating enzyme that increases ROS. Exposure of isolated rat extensor digitorum longus (EDL) muscles to Hx/XO (Hx/XO) for 20 min resulted in a dose-dependent increase in glucose uptake. To determine whether the mechanism leading to Hx/XO-stimulated glucose uptake is associated with the production of H2O2, EDL muscles from rats were preincubated with the H2O2 scavenger catalase or the superoxide scavenger superoxide dismutase (SOD) prior to incubation with Hx/XO. Catalase treatment, but not SOD, completely inhibited the increase in Hx/XO-stimulated 2-deoxyglucose (2-DG) uptake, suggesting that H2O2 is an intermediary leading to Hx/XO-stimulated glucose uptake with incubation. Direct H2O2 also resulted in a dose-dependent increase in 2-DG uptake in isolated EDL muscles, and the maximal increase was threefold over basal levels at a concentration of 600 micromol/l H2O2. H2O2-stimulated 2-DG uptake was completely inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin, but not the nitric oxide inhibitor NG-monomethyl-l-arginine. H2O2 stimulated the phosphorylation of Akt Ser473 (7-fold) and Thr308 (2-fold) in isolated EDL muscles. H2O2 at 600 micromol/l had no effect on ATP concentrations and did not increase the activities of either the alpha1 or alpha2 catalytic isoforms of AMP-activated protein kinase. These results demonstrate that acute exposure of muscle to ROS is a potent stimulator of skeletal muscle glucose uptake and that this occurs through a PI3K-dependent mechanism.
Figures
Fig. 1
Hypoxanthine/xanthine oxidase (Hx/XO)-stimulated 2-deoxyglucose (2-DG) uptake (A) and effects of catalase or superoxide dismutase (SOD) on Hx/XO-stimulated 2-DG uptake (B) in isolated extensor digitorum longus (EDL) muscles. A: isolated EDL muscles were stimulated in the absence or presence of 0.2–200 mU/ml XO with 1 mM Hx for 20 min followed by measurement of 2-DG uptake, as described in
RESEARCH AND METHODS
; n = 5–7 per group. B: isolated EDL muscles were preincubated in KRB buffer containing 3,000 U/ml catalase or 300 U/ml SOD for 30 min and then stimulated in 40 mU/ml XO with 1 mM Hx for 20 min followed by the measurement of 2-DG uptake. Data are means ± SE; n = 5 per group. *P < 0.05 vs. basal condition.
Fig. 2
Effects of H2O2 (0.1–24 mM) on 2-DG uptake in isolated EDL muscles. Isolated EDL muscles were stimulated in the absence or presence of 0.1–24 mM H2O2 for 20 min followed by the measurement of 2-DG uptake, as described in
RESEARCH AND METHODS
. Data are means ± SE; n = 5–7 per group.
Fig. 3
Effects of H2O2 on 5-aminoimidazole-4-carboxamide 1-β-
D
-ribonucleoside (AICAR)-stimulated 2-DG uptake in isolated EDL muscles. Isolated EDL muscles were incubated in KRB buffer in the absence or presence of 2 mmol/l AICAR for 50 min. H2O2 incubation was at a concentration (600 μmol/l) that elicited maximal increase in glucose uptake, as indicated in Fig. 2, and was added during the last 20 min the of 50-min incubation period. Data are means ± SE; n = 6–8 per group.
Fig. 4
Effects of H2O2 on AMPKα1 and -α2 activity (A) and effects of H2O2 on phosphorylation level of AMPKα (Thr172)(B) in isolated EDL muscles. Isolated EDL muscles were incubated in KRB buffer in the presence of H2O2 at a concentration (600 μmol/l) that elicited maximal increase in glucose uptake, as indicated in Fig. 2. Data are means ± SE; n = 6 per group.
Fig. 5
Effects of wortmannin on H2O2-stimulated 2-DG uptake (A) and effects of H2O2 on Akt Ser473 and Thr308 phosphorylation (B) in isolated EDL muscles. A: isolated EDL muscles were preincubated in KRB buffer in the presence or absence wortmannin (100 nM) followed by treatment with H2O2 at a concentration (600 μmol/l) that elicited maximal increase in H2O2-stimulated glucose uptake, as indicated in Fig. 2. Since wortmannin was dissolved in DMSO, which may act as a scavenger of superoxide anion, 2-DG uptake was measured in the presence of 0.2% DMSO for another control; n = 5–6 per group. B: isolated EDL muscles were incubated in KRB buffer in the presence or absence of H2O2 (600 μmol/l); n = 6 per group. Data are means ± SE. **P < 0.01 vs. control.
Fig. 6
Effects of H2O2 on insulin-stimulated 2-DG uptake in isolated EDL muscles. Isolated EDL muscles were preincubated in KRB buffer in the presence or absence of insulin at a concentration (300 nM) that induced maximal increase in insulin-stimulated glucose uptake. H2O2 incubation was at a concentration (600 μmol/l) that elicited maximal increase in glucose uptake, as indicated in Fig. 2, and was added during last 20 min of the 50-min incubation period. Data are means ± SE; n = 7–8 per group.
Fig. 7
Effects of _NG_-monomethyl-
L
-arginine (
L
-NMMA) on H2O2-stimulated 2-DG uptake in isolated EDL muscles. Isolated EDL muscles were preincubated in KRB buffer in the presence or absence of
L
-NMMA (100 nmol/l) for 30 min, followed with treatment with H2O2 at a concentration (600 μmol/l) that induces maximal increase in H2O2-stimulated glucose uptake, as indicated in Fig. 2. Data are means ± SE; n = 5–6 per group.
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