Figure 4. Bezafibrates reverse the brown fat mitochondrial dysfunction of mutant mice and induce beiging of subcutaneous fat depots in the regions of fat accumulation in participants with MMA.

(A) Representative sagittal, (B) coronal, and (C) transaxial micro-PET images of mice after injection with ¹⁸F-FDG. Mice were fed high-protein chow with 0.5% bezafibrate (BZF) for 2 months. White arrowheads point to the interscapular brown fat, while the red arrow in the coronal section (B) points to a markedly enhanced uptake of radioactive label in the shoulder region of the mutant animals. This area appears superimposed to the normal location of BAT ¹⁸FDG uptake (red arrow in C). Yellow arrows (C) point to the myocardium of a heterozygote and mutant animal, respectively. (D) ¹⁸FDG biodistribution in various tissues shows no significant differences between control and mutant mice treated with BZF. (E) Quantitative RT-PCR of Ucp1 mRNA expression in BAT (interscapular fat) and WAT (inguinal fat) showed no difference between mutant and control animals in BAT but a significant increase of Ucp1 expression in the subcutaneous WAT of the inguinal region in the mice fed high protein and 0.5% BZF (P < 0.0001 compared with heterozygotes, 1-way ANOVA with Tukey’s correction for multiple comparisons). (F) Immunohistochemistry staining for Ucp1 in subcutaneous adipose tissue of the shoulder region in a mutant mouse fed high protein and 0.5% BZF. Red arrows show multilocular lipid droplets staining densely for Ucp1 in the subcutaneous WAT resembling BAT (characteristic properties of beige fat). Blue arrow points to a hair follicle. Original magnification, 20×. (G) Ultrastructural studies demonstrated multinodular lipid droplets and numerous mitochondria with normal cristae in BAT from both heterozygous and mutant animals (scale bars: 2 μm). (H) Mmut^–/– Tg^INS-Alb-Mmut fed high-protein chow with 0.5% BZF show improved survival compared with high-protein chow only (Mantel-Cox survival curve comparison, P = 0.009). (I) Glomerular filtration rate (GFR) was measured with the FITC-inulin plasma decay method. Mutant mice fed high-protein chow showed a significantly reduced GFR (42.14% ± 3.35% compared with heterozygous mice on the same diet). GFR was improved to 61.29% ± 7.24% with BZF treatment (P = 0.02, compared with mutant mice on high protein only). *P < 0.05, **P < 0.01, ***P < 0.001.