Figure 3. FBF1 level is inversely correlated with the activation of the beiging program.

(A and B) Preadipocytes isolated from vWAT and differentiated. At day 10, (A) bright-field images were taken. Scale bar, 35 µm; (B) cells were fixed, stained with BIDIPY. Red: lipids, blue: nuclei. Scale bar, 50 µm.

(C) Bar graph illustrating the quantification of a brown-like lipid index, determined by calculating the ratio of total area for small (<1,000 µm²) versus large (>1,000 µm²) lipid droplets.

(D) Quantification of changes in lipid morphology is depicted as fraction of total lipid area per lipid droplet size.

(E) Immunoblotting on UCP-1 in preadipocytes from vWAT.

(F) Preadipocytes from vWAT were cultured and differentiated for 7 days, then labeled with mitochondria-GFP. At day 8, cells were fixed, stained, and imaged by fluorescence microscopy. Green: mitochondria, blue: nuclei. Scale bar, 200 µm.

(G–J) Preadipocytes from vWAT were cultured and induced to brown-like lipid morphology.

(G) At day 7, cells were fixed, stained with BIDIPY. Red: lipids, blue: nuclei. Scale bar, 50 µm.

(H) Bar graph illustrating the quantification of a brown-like lipid index, determined by calculating the ratio of total area for small (<1,000 µm²) versus large (>1,000 µm²) lipid droplets.

(I) Quantification of changes in lipid morphology is depicted as fraction of total lipid area per lipid droplet size.

(J) Cells were fixed, stained, and imaged. Green: mitochondria, red: actin, blue: nuclei. Scale bar, 50 µm.

(K) Photograph of vWAT collected from Fbf1^tm/tm/WT control exposed to 23°C or 4°C for 2 days.

(L) Photograph or WATs collected from Fbf1^tm/tm/WT control treated with vehicle or CL-316,243.

(M and N) H&E staining (M) and (N) Ucp1 mRNA level of WATs collected from mice treated with vehicle or CL-316,243.