Fig. 4

Testosterone-induced glucose metabolism is mediated by AMPK. a Western blot showing the phosphorylation levels of AMPK at residue Thr172 in response to 100 nM testosterone stimulation for 0, 3, 6, 12, 24 and 48 h. Ratio of p-AMPK to total AMPK protein as determined by densitometry. b ECAR was measured using a Seahorse XF analyzer and values were normalized using the total protein per well. ECAR kinetics in cells treated with 2 µM CC and 100 nM testosterone for 24 h. c Glycolysis and maximal glycolytic capacity parameters increased by testosterone were blocked by pretreatment with CC. d Cardiomyocytes were stimulated with 100 nM testosterone for 24 h and were then incubated with 2-NBDG for 20 min. Intracellular fluorescence was measured in the presence or absence of CC (2 µM). AMPK inhibition blocked the increase in glucose uptake induced by testosterone. e Cells were incubated for 30 min with 2 µM CC and then treated with 100 nM testosterone for 10 h. mRNA levels of Hk2 were assessed by RT-qPCR and normalized to 18S mRNA. Data are expressed as mean ± SEM of at least three independent experiments. P-values were determined using ANOVA followed by Tukey’s post hoc test; *P < 0.05, **P < 0.01, ***P < 0.001 vs. control; #P < 0.05, ##P < 0.01 vs. testosterone treated cells