Figure 1.

sLZIP inhibits adipogenesis via suppression of PPARγ₂ transcriptional activity. (a) 293T cells were transfected with PPARγ₂, β-galactosidase, FABP4-Luc, and various amounts of sLZIP (0.25 and 0.5 μg), and stimulated with or without 10 μM of rosiglitazone (Rosi) for 24 h. Promoter activity was determined using a luciferase assay. *P<0.01, **P<0.005. (b) 293T cells were transfected with various amounts of si-sLZIP (50 and 100 nM), and treated with or without 10 μM Rosi for 24 h. *P<0.01. (c) C3H10T1/2 cells stably expressing the FABP4 reporter gene were infected with sLZIP-expressing adenovirus, followed by differentiation into adipocytes for 4 days. *P<0.05. (d, e) 293T cells were transfected with PPARγ₂, β-galactosidase, FABP4-Luc, and various amounts of sLZIP (0.1, 0.25, and 0.5 μg) and RXR (0.1, 0.25, and 0.5 μg). Following transfection, cells were stimulated with or without 10 μM Rosi for 24 h. Luciferase activity was normalized to β-galactosidase activity and presented as the relative activity. (f) Pre-adipocyte cells were isolated from epididymal fat pads of WT and sLZIP TG mice. Cells were then transfected with FABP4-Luc and PPARγ₂, and treated with or without 10 μM Rosi for 24 h. Promoter activity was measured by a luciferase assay. *P<0.01. (g) C3H10T1/2 cells were infected with sLZIP-expressing adenovirus and differentiated into adipocytes. After 8 days from post-induction, cells were stained with Oil Red O. Scale bar, 50 μm. (h) Pre-adipocyte cells were isolated from epididymal fat pads of WT and sLZIP TG mice, and differentiated into adipocytes. After 8 days from post-induction, cells were stained with Oil Red O, and the expression level of FABP4 was determined using RT-PCR and Western blot analysis. Scale bar, 50 μm. (i) MEF cells isolated from WT and sLZIP TG mice were differentiated into adipocytes. Cells were stained with Oil Red O. Scale bar, 50 μm. (j) The white adipose tissue (fat) mass of WT and sLZIP TG mice. *P<0.05. (k) Total RNA was extracted and the mRNA level of PPARγ₂ target genes (FABP4, C.EBP1, and LPL) were determined using real-time PCR after 8 days from post-induction. GAPDH was used as an internal control. *P<0.005, **P<0.001. All experiments were performed in triplicate and the bar represents the mean±S.D.