Journal: Molecular Metabolism

Article Title: β-catenin regulates muscle glucose transport via actin remodelling and M-cadherin binding

doi: 10.1016/j.molmet.2020.101091

Figure Lengend Snippet: Depletion of β-catenin in L6-G4-myc cells impairs insulin-stimulated glucose transport and actin remodelling independent of Wnt-mediated transcription. Cyclin D1 protein expression in gastrocnemius muscle of BCAT-mKO and WT muscle was assessed by immunoblot (A–B). Glucose uptake and analysis of cyclin D1 mRNA expression in L6-G4-myc myoblasts treated with iCRT5 (50 μM) for 1 h (C–D). L6-G4-myc myoblasts were treated with β-catenin specific siRNA prior to assessment of glucose uptake (E–F) and insulin-signalling pathway (G–K). Results are mean ± SE, with individual animals or biological replicates shown as data points in figures or in figure legend. Significance was determined using two-tailed Student's t-test (D, K) or two-way ANOVA with LSD post-hoc analysis (B, C, E–J). ∗p < 0.05, ∗∗∗p < 0.001 significance within groups. #p < 0.05 significance between groups.

Article Snippet: L6-G4-myc myoblasts (Kerafast, Boston, MA, USA) were grown in low-glucose Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% foetal bovine serum and 1% penicillin/streptomycin.

Techniques: Expressing, Western Blot, Two Tailed Test

Journal: Molecular Metabolism

Article Title: β-catenin regulates muscle glucose transport via actin remodelling and M-cadherin binding

doi: 10.1016/j.molmet.2020.101091

Figure Lengend Snippet: Insulin phosphorylates β-catenin S552 downstream of proximal insulin signalling, and β-catenin S552 phosphorylation is required for insulin-stimulated glucose transport. Immunoblotting analysis of P-β-catenin S552 , β-catenin, P-Akt S473 Akt, and β-actin in C57Bl/6j mice treated with either glucose or insulin prior to tissue collection (A–B). Immunoblotting analysis of P-β-catenin S552 , β-catenin, P-Akt S473 , Akt and β-actin in L6-G4-myc cells incubated with either BYL719 or AKTi-1/2 prior to insulin stimulation and (C–E). Insulin-stimulated glucose uptake and GLUT4 translocation to the membrane in L6-G4-myc cells transfected with S552A-β-catenin (F–H). Results are mean ± SE, with individual animals or biological replicates shown as individual data points in figures or in figure legend. Significance was determined using, one (B) or two-way ANOVA with LSD post-hoc analysis (D, E and G, H). ∗p < 0.05, ∗∗p < 0.01 significance within groups. #p < 0.05 significance between groups.

Article Snippet: L6-G4-myc myoblasts (Kerafast, Boston, MA, USA) were grown in low-glucose Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% foetal bovine serum and 1% penicillin/streptomycin.

Techniques: Western Blot, Incubation, Translocation Assay, Transfection

Journal: Molecular Metabolism

Article Title: β-catenin regulates muscle glucose transport via actin remodelling and M-cadherin binding

doi: 10.1016/j.molmet.2020.101091

Figure Lengend Snippet: Insulin promotes β-catenin/M-cadherin interaction via β-catenin S552 phosphorylation. L6-G4-myc myotubes were treated with either insulin or PBS (control) prior to lysis and immunoprecipitation with β-catenin-specific antibody (A). L6-G4-myc myoblasts were treated with M-cadherin-specific siRNA prior to determination of F-actin % (B) and glucose uptake (C) following insulin stimulation. F actin % in response to insulin (D) and PDGF-mediated glucose uptake (E) and immunoblotting of M-cadherin following immunoprecipitation with anti-β-catenin antibody (F–G) in S552A-β-catenin expressing L6-G4-myc myoblasts. Results are mean ± SE, with biological replicates shown as individual data points in figures or in figure legend. Significance was determined using the sign-rank test (A) or two-way ANOVA with LSD post-hoc analysis (B–G). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 significance within groups. #p < 0.05 significance between groups.

Article Snippet: L6-G4-myc myoblasts (Kerafast, Boston, MA, USA) were grown in low-glucose Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% foetal bovine serum and 1% penicillin/streptomycin.

Techniques: Lysis, Immunoprecipitation, Western Blot, Expressing

Journal: Diabetologia

Article Title: Treatment with a β-2-adrenoceptor agonist stimulates glucose uptake in skeletal muscle and improves glucose homeostasis, insulin resistance and hepatic steatosis in mice with diet-induced obesity

doi: 10.1007/s00125-020-05171-y

Figure Lengend Snippet: In vitro and acute in vivo effects of clenbuterol. ( a ) Clenbuterol stimulation of L6 myotubes induced glucose uptake in a dose-dependent manner; n = 5–8; data were analysed by one-way ANOVA with Dunnett’s multiple comparison test. ( b , c ) Stimulation of L6 myotubes stably expressing GLUT4myc with 1 μmol/l clenbuterol induces GLUT4 translocation as quantified in ( c ); n = 3; data were analysed with unpaired two-tailed Student’s t test. Scale bars, 50 μm. ( d ) In vivo glucose uptake in gastrocnemius muscle of chow-fed mice treated with 1 mg/kg clenbuterol for 1 h; n = 5–6; data were analysed with unpaired two-tailed Student’s t test. ( e , f ) Acute effects of clenbuterol on blood glucose ( e ) and plasma insulin ( f ); n = 6 (diet-induced obesity was developed in C57Bl/6N mice maintained at 30°C and on HFD for 7 months; mice were fasted in the morning for 6 h prior to clenbuterol, glucose or saline i.p. injection); data were analysed by two-way ANOVA with Dunnett’s multiple comparison test. In all graphs: * p < 0.05, ** p < 0.01, *** p < 0.001 vs vehicle (in e and f vs vehicle at the same time point)

Article Snippet: L6 rat myoblasts and L6 myoblasts stably expressing GLUT4myc were purchased from KeraFast (ESK201 and ESK202), where they were tested for mycoplasma.

Techniques: In Vitro, In Vivo, Stable Transfection, Expressing, Translocation Assay, Two Tailed Test, Injection

Journal: Diabetologia

Article Title: Treatment with a β-2-adrenoceptor agonist stimulates glucose uptake in skeletal muscle and improves glucose homeostasis, insulin resistance and hepatic steatosis in mice with diet-induced obesity

doi: 10.1007/s00125-020-05171-y

Figure Lengend Snippet: In vitro and acute in vivo effects of clenbuterol. ( a ) Clenbuterol stimulation of L6 myotubes induced glucose uptake in a dose-dependent manner; n = 5–8; data were analysed by one-way ANOVA with Dunnett’s multiple comparison test. ( b , c ) Stimulation of L6 myotubes stably expressing GLUT4myc with 1 μmol/l clenbuterol induces GLUT4 translocation as quantified in ( c ); n = 3; data were analysed with unpaired two-tailed Student’s t test. Scale bars, 50 μm. ( d ) In vivo glucose uptake in gastrocnemius muscle of chow-fed mice treated with 1 mg/kg clenbuterol for 1 h; n = 5–6; data were analysed with unpaired two-tailed Student’s t test. ( e , f ) Acute effects of clenbuterol on blood glucose ( e ) and plasma insulin ( f ); n = 6 (diet-induced obesity was developed in C57Bl/6N mice maintained at 30°C and on HFD for 7 months; mice were fasted in the morning for 6 h prior to clenbuterol, glucose or saline i.p. injection); data were analysed by two-way ANOVA with Dunnett’s multiple comparison test. In all graphs: * p < 0.05, ** p < 0.01, *** p < 0.001 vs vehicle (in e and f vs vehicle at the same time point)

Article Snippet: L6 rat myoblasts and L6 myoblasts stably expressing GLUT4myc were purchased from KeraFast (ESK201 and ESK202), where they were tested for mycoplasma.

Techniques: In Vitro, In Vivo, Stable Transfection, Expressing, Translocation Assay, Two Tailed Test, Injection