Figure 5.

TAG turnover is required for enhanced glucose-stimulated insulin release from LXR-activated INS-1 cells. A, Impact of orlistat on insulin release. INS-1 cells were incubated for 48 h in 16.7 mm glucose (Gluc) ± T0901317 (TO; 10 μm). Insulin release (60 min) in response to 2 or 20 mm glucose was then assessed in the presence or absence of orlistat (50 μm). Values are mean ± sem for six independent experiments. *, P < 0.001 when orlistat-treated cells are compared with control cells. B, Impact of orlistat on turnover of de novo-derived TAG, DAG, and FFA. INS-1 cells were cultured for 48 h in 16.7 mm glucose ± T0901317 (10 μm). During the last 6 h, cells were incubated with [2-¹⁴C]acetic acid (t = 0), after which cells were subjected to conditions for an acute insulin release study: 1 h incubation in 2 mm glucose followed by a 1-h incubation in 2 or 20 mm glucose. Total lipids were extracted and analyzed as described in Fig. 3A. Values are mean ± sem for three independent experiments. Data are presented relative to ¹⁴C-labeling at t = 0. PhosphoImager intensity values at t = 0 for TAG are 482,621 ± 59,573, for DAG are 39,461 ± 4,451 and for FFA are 37,267 ± 3,691. C, Impact of orlistat (50 μm) on de novo FA synthesis from glucose. INS-1 cells were incubated for 48 h in 16.7 mm glucose ± T0901317, and subjected to a insulin release assay with 2 or 20 mm glucose containing 4 or 40 μCi of [U-¹⁴C]glucose, respectively. ¹⁴C-labeled FAs were quantified as described in Materials and Methods. Values are mean ± sem of three independent experiments. *, P < 0.001 orlistat-treated cells are compared with control cells.