Inhibition of NPC1 repressed insulin-stimulated glucose uptake.

<p>(<b>a</b>) 3T3-L1 adipocytes were treated with U18666a for indicated periods of time before radiolabelled 2-deoxy-D-glucose uptake assays were performed on basal and insulin-stimulated cells. Data are mean ±S.E.M, n = 4 independent experiments. (<b>b</b>) 3T3-L1 adipocytes electroporated with scrambled (Scr) or siRNA directed to NPC1 (NPC1) were subjected to radiolabelled 2-deoxy-D-glucose uptake assays under basal and insulin-stimulated conditions. Insulin doses as indicated. Data are mean ±S.E.M, n = 3 independent experiments. (<b>c</b>) Plasma membrane (PM) levels of IRAP, GLUT4 and Caveolin 1 in unstimulated and insulin-stimulated (100 nM, 20 min) adipocytes were determined by immunoblotting. (<b>d</b>) PM levels of HA-GLUT4 was measured using a fluorescence based assay in 3T3-L1 adipocytes overexpressing HA-GLUT4 treated with U18666a for indicated periods of time. Data are mean ±S.E.M, n = 3 independent experiments. Significance calculated by two-way ANOVA. Comparisons to unstimulated cells (a, b, d) indicated by n.s = non-significant, * = p<0.05 ** = p<0.01 *** = p<0.001 **** = p<0.0001. Comparisons to insulin-stimulated cells (a, b, d) indicated by n.s = non-significant, # = p<0.05 ## = p<0.01 ### = p<0.001 #### = p<0.0001.