Figure 4: Browning of inguinal adipose tissue and activation of a thermogenic programme by BBR.

(a) Representative haematoxylin and eosin staining from IWAT sections of db/db mice after 4 weeks of BBR treatment. Original magnification, × 200. Scale bar, 100 μm. (b) Representative electron microscopy micrographs determined in IWAT of vehicle and drug-treated mice. L, lipid droplet; M, mitochondria; Original magnification, × 7400. Scale bar, 2,000 nm. (c) MtDNA copy number of IWAT from BBR or vehicle-treated db/db mice (n=5). (d) Immunohistochemistry for UCP1 protein (brown stain) (upper panel) and TH (lower panel) in IWAT sections of db/db mice treated with vehicle and BBR. (e) BBR induces brown/brite-typical gene expression in IWAT. Real-time PCR analysis of thermogenic gene expression is shown (n=4–6). (f) Representative western blots showing the key protein changes in IWAT. (g) Thermogenic-related gene expression profile in primary IWAT adipocytes after BBR treatment. Primary adipocytes were sourced from IWAT and differentiated as described in ‘Methods’, then treated with 2.5 μM BBR for 12 h. (h) Representative western blots showing the protein expression levels in primary IWAT adipocytes treated with vehicle or BBR. Cells were treated with drug for 24 h. Values represent mean±s.e.m. Error bars represent s.e.m., and significant differences compared with vehicle controls are indicated by *P≪0.05, **P≪0.01 and ***P≪0.001 (assessed by Student's t-test).