Figure 1.
Effect of UA on the viability of 3T3-L1 adipocytes.
Mature 3T3-L1 adipocytes were incubated with different concentrations of UA for 24, 48, or 72 h, respectively. The MTT reagents were added to the medium. After 4 h of incubation, the medium was aspirated and 150 µL DMSO was added to each well. The absorbance was measured at 570 nm. Data are expressed as means ± SD (n = 3). *P<0.05, **P<0.001 and ***P<0.001 vs. the control of 0 µM UA.
Figure 2.
Effect of UA on glucose uptake in 3T3-L1 adipocyte.
Mature 3T3-L1 adipocytes were incubated with 2.5 µM, 5 µM, or 10 µM of UA for 2 h and glucose uptake was measured in the presence or absence of 1 µg/mL insulin. The fluorescence intensity of NBD-glucosewas measured at 466/550 nm on a Varioskan Flash spectral scanning multimode plate reader. Data are expressed as means ± SD (n = 3). * P<0.05 and **P<0.01 vs. the control of 0 µM UA; #P<0.05 and ##P<0.01 vs. 1 µg/mL of insulin.
Figure 3.
Effect of the PI3K inhibitor wortmannin on glucose uptake in 3T3-L1 adipocyte.
Mature 3T3-L1 adipocytes were incubated with 10 µM of UA for 2 h, or pretreated with 1 µM of wortmannin for 30 min before incubation with 10 µM of UA and 1 µM of wortmannin for 2 h. Glucose uptake was measured in the presence or absence of 1 µg/mL insulin. The fluorescence intensity of NBD-glucose was measured at 466/550 nm on a Varioskan Flash spectral scanning multimode plate reader. Data are expressed as means ± SD (n = 3). *P<0.05 and **P<0.01 vs. the control of 0 µM UA; #P<0.05, ##P<0.01 and ###P<0.001 vs. the insulin control of 1 µg/mL. Wor indicates 1 µM of wortmannin.
Figure 4.
Effect of UA on the activity of PDK, AKT, PKC and AS160, and the expression of GLUT1 and GLUT4.
Mature 3T3-L1 cells were treated with 10 µM of UA in the absence or presence of 1 µg/mL insulin for 24 h, or pretreated with 1 µM of wortmannin for 30 min before incubation 10 µM of UA and 1 µM of wortmannin for 24 h. The activity of PDK, AKT, PKC and AS160, and the expression of GLUT1 and GLUT4 were assessed by Western blotting. Data are expressed as means ± SD (n = 3). Wor indicates 1 µM of wortmannin.
Figure 5.
Effect of UA on GLUT4 expression and translocation in 3T3-L1 adipocyte.
(A) Mature 3T3-L1 cells were treated with 10 µM of UA in the absence or presence of 1 µg/mL insulin for 24 h, or pretreated with 1 µM of wortmannin for 30 min before incubation with 10 µM of UA and 1 µM of wortmannin. The expression level of GLUT4 was determined by immunofluorescence. (B) Mature 3T3-L1 cells were treated with 10 µM of UA in the absence or presence of 1 µg/mL insulin for 24 h, or pretreated with 1 µM of wortmannin for 30 min and then with 10 µM of UA and 1 µM of wortmannin. Cell membrane was obtained and the expression of GLUT4 was determined with Western blotting (n = 3). Wor indicates 1 µM of wortmannin.