A, Male wild-type (Wt) and heterozygous TrJ mice were used for these studies. Body weight measured every other week during the study indicates a significant increase for all four groups (p<0.0001 ANOVA, n=9 mice per condition; error bars represent SEM). B, Performance of the mice on the rotarod shows a main effect of genotype (p<0.001 ANOVA; error bars represent SEM) and a difference across trials (p<0.05 ANOVA). The performance of TrJ mice after four months on the regimen significantly improved (Post hoc ANOVA, p<0.001). Statistical analysis is presented in Supplementary Table 2. C, After five months on the regimen, the IF animals demonstrate improvement in forelimb strength (***p<0.001 ANOVA, n=6). Results are presented using Tukey box-and-whisker plots, where the box represents the inter-quartile range; the horizontal line indicates median value and error bars represent maximum and minimum values.

Cross-sectional views of proximal sciatic nerves from AL and intermittently fed Wt and TrJ mice are shown. On thin sections from AL fed neuropathic animals (left, inset), poorly myelinated axons (A) and multiple layers of loose basal lamina (arrows) are visible. In mice that had undergone an IF regimen (right), axonal myelination is thicker and the multiple basal lamina layers of the Schwann cells is replaced by a single tight basal lamina (arrowheads). N:nucleus. Micron bar: 10 μm and 2 μm (insets).

A, In samples from neuropathic mice, the cross-sectional area occupied by nerve fibers is improved in response to the regimen (*p<0.05). B, The percentage of myelinated vs. total fibers within nerves of neuropathic mice on the IF regimen is higher, as compared to AL fed (**p<0.01). A, B: n=4 mice; error bars represent SEM. C, The distribution of fiber diameters in micrometer is shown (horizontal lines indicate mean values). The IF regimen increases the mean fiber diameter in nerves from both Wt and TrJ mice as compared to AL fed (***p<0.001, n=4 mice). D, The thickness of myelin sheath is increased in nerves from mice on the regimen as compared to AL fed (***p<0.001). E, Quantification of the median g-ratios (axon diameter/fiber diameter) of individual nerve fibers (n=150) indicates a significantly improved myelination in nerves from mice on the IF regimen, as compared to AL fed (***p<0.001). D and E, Results are presented using Tukey box-and-whisker plots, where the box represents the inter-quartile range; the horizontal line indicates median value and error bars represent maximum and minimum values. Statistical analysis is presented in Supplementary Table 2.

A-B, Whole sciatic nerve lysates (20 μg/lane) from Wt and TrJ mice were probed with the indicated anti-myelin protein (P0, PMP22 and MBP), or anti-heat shock protein (Hsp) antibodies. GAPDH is shown as a protein loading control. Molecular mass in kDa, on left. C, On longitudinal sections of sciatic nerves from intermittently fed neuropathic animals, MBP-positive myelin internodes are prominent in IF mice (arrows). Co-staining the sections with anti-HSC70 antibody reveals increased expression of this cytosolic chaperone within Schwann cells (arrows). Nuclei are stained with Hoechst dye (blue). Samples from Wt mice are shown in the insets for comparison. Micron bar, 20 μm.

A, Whole sciatic nerve lysates (20 μg/lane) from Wt and TrJ mice were probed with anti-p75 and pHH3 antibodies. GAPDH is shown as a protein loading control. Molecular mass in kDa, on left. B, In neuropathic nerves on the IF regimen, the number of Schwann cell nuclei per fixed area of tissue is decreased significantly (***p<0.001 ANOVA, n=3 mice).

A, On longitudinal nerve sections from Wt mice fed AL or intermittently, PMP22 is detected along internodal segments of myelin (arrowheads). PMP22-containing protein aggregates are visible within neuropathic samples (arrows). Myelin-like PMP22 immunoreactivity is visible in nerves from neuropathic mice on the regimen (TrJ-IF) (arrowheads). Nuclei are stained with Hoechst dye (blue). Micron bar, 20 μm. B, The frequency of PMP22 protein aggregates is significantly reduced in the intermittently fed TrJ mice as compared to AL fed (***p<0.001 ANOVA, n=4; error bars represent SEM). C, Western blot analyses on total nerve lysates (20 μg/lane) reveal that the dietary regimen dramatically attenuates the accumulation of poly-ubiquitinated (pUbi) proteins in neuropathic samples, while mono-ubiquitinated (mUbi) substrate levels are maintained across genotype and diet. D, The levels of the autophagy pathway markers, Atg7, LC3 and p62 are shown. The lipidation of LC3, as visualized by the presence of LC3 II, is evident in samples from TrJ mice on the regimen. E, The expression of LAMP1 and cathepsin D is influenced by the genotype and stimulated by IF regimen. C-E, GAPDH is shown as a protein loading control. Molecular mass in kDa, on left.

A, Longitudinal nerve sections from TrJ mice on the AL and IF regimen were immunostained with an anti-CD11b antibody. Nuclei are labeled with Hoechst dye (blue). Micron bar, 20 μm. B, Quantification of the number of CD11b positive cells in a fixed area of nerve tissue (***p<0.001 ANOVA, n=3). C, Western blot analysis of whole nerve lysates (20 μg/lane) with an anti-mouse IgG antibody is shown. The heavy (HC) and light chains (LC) of immunoglobulin G are shown. GAPDH serves as a protein loading control. Molecular mass in kDa, on left.