Table 1.
The effect of bezafibrate treatment in mouse models of disease.
Figure 1.
Systemic effects of bezafibrate on Mut and WT mice.
(A) Body weight of mice from 2 to 14 month-old (n = 11–26/group for 2 to 11 months; 2–6/group for 12 to 14 months). The difference in body weight between WTSD and MutSD mice is statistically significant from 3 to 14 months, between WTSD and WTBD mice is significant at every time point and the difference between MutSD and MutBD is significant from 3 to 14 months. Measurement of total (B) bone mineral density (BMD), (C) bone mineral content (BMC) (D), body area (cm2), lean mass (g), total body fat (g) and percent (%) fat of 10 month-old mice (n = 5–7/group). (E) Percent survival of mice (n = 11–15/group). *, P<0.05; **, P<0.01, Student’s t-test. Error bars represent the SEM.
Figure 2.
Bezafibrate delayed hair loss and restored the skin structure of Mut mice.
(A) Pictures of mice at 7 month-old and 10 month-old showing their coat phenotype (n = 6/group). Squares highlight area of coat being described. (B) Dorsal skin sections from 10 month-old mice showing hematoxylin and eosin (H&E) staining to depict structural changes (black arrow indicates break in the epidermis layer of MutSD mice), Verhoeff’s Van Geison (EVG) staining for elastic fibers shown in black/dark brown (yellow arrow) and Masson’s trichrome staining showing collagen in blue (n = 2/group). (C) Western blot showing total Smad3 and glyceralgehyde 3-phosphate (GAPDH) protein levels in total skin homogenate from 10 month-old mice and quantification of total Smad3 band intensity normalized to GAPDH (n = 4/group). Error bars represent the SEM.
Figure 3.
Bezafibrate improved spleen size and structure of Mut mice.
(A) Spleen weight of 10 month-old mice and (B) picture of spleen from Mut mice (n = 4–6/group). (C) Quantification of cleaved caspase-3 immunostaining in paraffin sections from the spleen of 10 month-old mice (n = 3–4/group). *, P<0.05, one-way analysis of variance followed by Bonferroni’s multiple comparison test. (D) H&E staining of the spleen of 10 month-old mice showing the organization of white pulp (purple) and red pulp (pink) (n = 3/group). (E) Results from complete blood cell count in 10 month-old mice showing RBC (red blood cells, x106µl ) (n = 5–6/group) (F) PGC-1α and PPARγ mRNA levels in the spleen of 10 month-old mice normalized to actin. (G) Quantification of western blot showing mitochondrial protein levels in total homogenate from the spleen of 10 month-old mice normalized to actin. NADH dehydrogenase (ubiquinone) 1β subcomplex subunit 8 (NDUFB8; subunit of complex I), succinate dehydrogenase subunit B (SDHB; subunits of complex II), ubiquinol-cytochrome c reductase core protein 2 (UQCRC2; subunit of complex III), and ATP synthase subunit 5α (ATP5A; subunit of complex V). *, P<0.05; **, P<0.01, Student’s t-test. Error bars represent the SEM.
Figure 4.
The effect of bezafibrate on the skeletal muscle of Mut and WT mice.
(A) Western blot of PGC-1α and loading control actin in total quadricep homogenate from 10 month-old mice and quantification of PGC-1α band intensity normalized to actin (n = 4/group). (B) Gene expression of PGC-1β and PPARs in the quadricep of 10 month-old mice normalized to actin (n = 4/group). (C) Citrate synthase activity in the total quadricep homogenate from 10 month-old mice (n = 4/group). (D) Quantification of western blot of mitochondrial proteins in the total homogenate from the quadricep of 10 month-old mice (n = 4/group). NADH dehydrogenase (ubiquinone) 1β subcomplex subunit 8 (NDUFB8; subunit of complex I), succinate dehydrogenase subunit B (SDHB; subunits of complex II), ubiquinol-cytochrome c reductase core protein 2 (UQCRC2; subunit of complex III), mitochondrial cytochrome c oxidase subunit 1 (MTCO1; subunit of complex IV) and ATP synthase subunit 5α (ATP5A; subunit of complex V) (E) Quantification of mtDNA levels in the quadriceps of 10 month-old mice based on ND1 (subunit of complex I) levels normalized to glyceraldehyde 3-phosphate (GAPDH) (n = 4/group). (F) Gene expression of markers of fatty acid oxidation, acyl-coenzymeA oxidase 1 (ACOX), cluster of differentiation 36 (CD36), carnitine palmitoyl transferase (CPT1) and short-chain-acyl-coenzymeA dehydrogenase (SCAD) in the quadriceps of 10 month-old mice normalized to actin (n = 4/group) (G) Skeletal muscle weight of 10 month-old mice (n = 4–6/group). (H) The number of falls of mice when put to run on a treadmill for 3 minutes at 9 meters/minute (n = 5–10/group). *, P<0.05; **, P<0.01, ***P<0.001, Student’s t-test. Error bars represent the SEM.
Figure 5.
Bezafibrate induces hepatomegaly and fatty acid oxidation in Mut and WT mice.
(A) Liver weight of 10 month-old mice (n = 4–6/group). (B) H&E staining of the liver of 10 month-old mice showing hepatocytes and central vein (n = 4/group). (C) The level of liver enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the blood of 10 month-old mice (n = 6/group). (D) Gene expression of PGC-1 coactivators and PPARs in the liver of 10 month-old mice normalized to actin (n = 4/group). (E) The mRNA level of markers of fatty acid oxidation in the liver of 10 month-old mice normalized to actin (n = 4/group). Acyl-coenzymeA oxidase 1 (ACOX), cluster of differentiation 36 (CD36), carnitine palmitoyl transferase (CPT1) and short-chain-acyl-coenzymeA dehydrogenase (SCAD). *, P<0.05; **, P<0.01, ***P<0.001, Student’s t-test. Error bars represent the SEM.