Abstract
Aims/hypothesisImpaired glucose uptake in skeletal muscle is an important contributor to glucose intolerance in type 2 diabetes. The aspartate protease, beta-site APP-cleaving enzyme 1 (BACE1), a critical regulator of amyloid precursor protein (APP) processing, modulates in vivo glucose disposal and insulin sensitivity in mice. Insulin-independent pathways to stimulate glucose uptake and GLUT4 translocation may offer alternative therapeutic avenues for the treatment of diabetes. We therefore addressed whether BACE1 activity, via APP processing, in skeletal muscle modifies glucose uptake and oxidation independently of insulin.MethodsSkeletal muscle cell lines were used to investigate the effects of BACE1 and α-secretase inhibition and BACE1 and APP overexpression on glucose uptake, GLUT4 cell surface translocation, glucose oxidation and cellular respiration.ResultsIn the absence of insulin, reduction of BACE1 activity increased glucose uptake and oxidation, GLUT4myc cell surface translocation, and basal rate of oxygen consumption. In contrast, overexpressing BACE1 in C2C12 myotubes decreased glucose uptake, glucose oxidation and oxygen consumption rate. APP overexpression increased and α-secretase inhibition decreased glucose uptake in C2C12 myotubes. The increase in glucose uptake elicited by BACE1 inhibition is dependent on phosphoinositide 3-kinase (PI3K) and mimicked by soluble APPα (sAPPα).Conclusions/interpretationInhibition of muscle BACE1 activity increases insulin-independent, PI3K-dependent glucose uptake and cell surface translocation of GLUT4. As APP overexpression raises basal glucose uptake, and direct application of sAPPα increases PI3K–protein kinase B signalling and glucose uptake in myotubes, we suggest that α-secretase-dependent shedding of sAPPα regulates insulin-independent glucose uptake in skeletal muscle.
Highlights
The increase in glucose uptake elicited by beta-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1) inhibition is dependent on phosphoinositide 3-kinase (PI3K) and mimicked by soluble APPα
Glucose uptake and GLUT4 translocation in myotubes are modulated by BACE1 activity in an insulin-independent manner We detected BACE1 and APP protein in wild-type C2C12 myoblasts and myotubes (Fig. 1a, b) and demonstrated that BACE1 was proteolytically active by the presence of sAPPβ in the incubation medium (Fig. 1c, d)
Uptake in control and M-3-treated myotubes (n=11); white bars, vehicle; black bars, insulin. (f) 2DG uptake in myotubes treated with vehicle (Veh), M-3 or BACE1 inhibitor (Inh) II (n=6–11). (g) 2DG uptake in myotubes transfected with EVor mBACE1 (n=4). (h) Total [14C]glucose incorporation in myotubes after vehicle and M-3 (n=4). (i, j) Gain in cell surface GLUT4myc±insulin (Ins; 20 and 100 nmol/l)±M-3 in myotubes (i) and myoblasts (j) (n=4). *p
Summary
Glucose uptake and GLUT4 translocation in myotubes are modulated by BACE1 activity in an insulin-independent manner We detected BACE1 and APP protein in wild-type C2C12 myoblasts and myotubes (Fig. 1a, b) and demonstrated that BACE1 was proteolytically active by the presence of sAPPβ in the incubation medium (Fig. 1c, d). Palmitate (750 μmol/l) inhibited insulin-stimulated glucose uptake in the absence and presence of M-3 (Fig. 3b), this was associated with substantially increased BACE1 and APP protein levels (Fig. 3c). Batimastat-treated myotubes showed reduced ability to switch substrate from glucose to palmitate in response to increased substrate delivery (Fig. 5h), indicating diminished metabolic flexibility These data suggest that myotube α-secretase substrate cleavage resulted in the maintenance or enhancement of basal (insulin-independent) glucose uptake but did not affect fuel oxidation. Overexpression of APP in C2C12 myotubes increased basal and insulin-stimulated (Fig. 5j, k) glucose uptake, but had little or no effect on glucose or palmitate oxidation (Fig. 5l, m). Consistent with the involvement of PI3K signalling in APP/BACE1-modulated basal glucose uptake, the gain in cell surface GLUT4myc caused by M-3, in the absence and presence of insulin, was inhibited by wortmannin (Fig. 6c). These data suggest that modification of skeletal muscle APP cleavage by α- and β-secretases results in altered sAPPα abundance, which affects PI3K– PKB signalling to alter GLUT4 translocation and glucose uptake
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