Pelizaeus-Merzbacher disease (PMD) is an untreatable and fatal leukodystrophy. In a model of PMD with perturbed blood-brain barrier integrity, cholesterol supplementation promotes myelin membrane growth. Here, we show that in contrast to the mouse model, dietary cholesterol in two PMD patients did not lead to a major advancement of hypomyelination, potentially because the intact blood-brain barrier precludes its entry into the CNS. We therefore turned to a PMD mouse model with preserved blood-brain barrier integrity and show that a high-fat/low-carbohydrate ketogenic diet restored oligodendrocyte integrity and increased CNS myelination. This dietary intervention also ameliorated axonal degeneration and normalized motor functions. Moreover, in a paradigm of adult remyelination, ketogenic diet facilitated repair and attenuated axon damage. We suggest that a therapy with lipids such as ketone bodies, that readily enter the brain, can circumvent the requirement of a disrupted blood-brain barrier in the treatment of myelin disease.
Axonal degeneration; Ketogenic diet; Mitochondria; Myelin; Pelizaeus–Merzbacher disease; Remyelination.
Conflict of interest statement
The authors have nothing to declare.
Increased ketone body uptake and ketolysis is induced in KD fed animals.
a Treatment paradigm, Plp1tgB mice were fed ketogenic diet (KD) or standard chow (SD) between 2 and 12 weeks of age. b Mean blood levels of the ketone body beta-hydroxybutyrate (βHB) ± SEM. Significance was evaluated by 2way ANOVA with Tukey’s post test [ N = 4 (wild type, WT), N = 8 (SD fed Plp1tgB), N = 9 (KD fed Plp1tgB)]. c Representative immunofluorescence detecting MCT1 on spinal cord sections of SD or KD fed Plp1tgB mice. Isolectin served to visualize blood vessels. The dashed line marks the tissue outline and the boxed area is enlarged on the right. d Western Blot with quantification of OXCT1 in wild type mice ( N = 3), Plp1tgB mice fed SD ( N = 3) or KD ( N = 4). Equal protein loading was confirmed by staining of actin. Significance was tested using 1way ANOVA with Tukey’s multiple comparison test. e Immunolabeling of OXCT1 and neurofilament heavy chain (NF200) in the corticospinal tract of the spinal cord in Plp1tgB mice fed SD or KD. Dashed line marks the border between gray matter (GM) and white matter (WM) in dorsal spinal cord. The boxed area is enlarged on the right. Indicated are only significant differences between SD and KD fed Plp1tgB mice (* P < 0.05; *** P < 0.001). Scale bars 10 µm
KD ameliorates PMD pathology in Plp1tgB animals.
a Olig2 and CAII (arrowheads) immunolabeling of wild type and Plp1tgB mice fed SD and KD with quantification of cell numbers in dorsal white matter of the spinal cord on the right ( N = 4–5 (WT), N = 7–8 (Plp1tgB fed SD), N = 8–9 (Plp1tgB fed KD), 1way ANOVA with Tukey’s post test). b Western Blot with quantification of ATF6 in lumbar spinal cord of wild type mice ( N = 4), Plp1tgB mice fed SD ( N = 3) or KD ( N = 4). Equal protein loading was confirmed by reprobing for actin (1way ANOVA with Tukey’s post test). c Quantitative RT-PCR determining Plp1 in spinal cord of Plp1tgB mice fed SD or KD ( N = 8, 1way ANOVA with Sidak’s post test) normalized to wild type controls ( N = 5, set to 1). d Quantification of myelination in the corticospinal tract from wild type mice, and Plp1tgB mice fed SD or KD ( N = 7), showing g-ratio analysis as scatter plot (left panel) and the mean g ratio (right panel, 1way ANOVA with Tukey’s post test). e Relative frequency of sufficiently myelinated fibers (g ratio < 0.8), hypomyelinated fibers ( g ratio > 0.8) or unmyelinated fibers ( g-ratio = 1) in the CST of Plp1tgB fed SD or KD ( N = 7, two-sided Student’s t test of each group). f Rotarod analysis and g elevated beam test performance at 6 to 12 weeks of age ( N = 7–8; 2way ANOVA with Sidak’s post test). Indicated are only significant differences between SD and KD fed Plp1tgB mice (* P < 0.05; ** P < 0.01; *** P < 0.001). Scale bars 20 µm
KD rescues mitochondria enlargement and ameliorates impulse conduction in Plp1tgB mice.
a APP staining of axonal swellings/spheroids (arrowheads) in wild type and Plp1tgB mice fed SD and KD with quantification of cells per 30,000 µm 2 corticospinal tract of the spinal cord ( N = 6–8; 1way ANOVA with Tukey’s post test). Scale bar 20 µm. b Electron micrographs showing an example of enlarged axonal mitochondria (colored in yellow) in Plp1tgB mice fed SD and a normally sized mitochondrium in KD fed Plp1tgB mice. Scale bar 0.5 μm. c Mean area of axonal mitochondrial in cross-sectioned corticospinal tract of wild type mice, and Plp1tgB animals fed SD or KD (at least 60 mitochondria per animal were counted, N = 6–7 animals; 1way-ANOVA with Tukey’s post test). d Stratification of mitochondrial sizes shown in c with respect to myelinated axons ( g-ratio < 1) and unmyelinated axons ( g-ratio = 1) (two-sided Student’s t test of each group). e 3D modeling of ACO2 immunolabeled mitochondria in deconvolved confocal images from Plp1tgB mice fed SD or KD with quantification of mitochondrial volumes (SD, 3178 or KD, 2817 mitochondria; 4 animals per condition). Volume data are presented as median with interquartile range (Kolmogorov–Smirnov test). f Density of myelinated axons in the optic nerve of SD and KD fed Plp1tgB animals (mean per 100 square µm ± SEM of N = 4–5 animals counting 13 images per animal; two-sided Student’s t test). g Evaluation of mitochondrial area in axons of the optic nerve analogous to the analysis shown in c ( N = 5 animals, 1way-ANOVA with Tukey’s post test). h– n Electrophysiological evaluation of ( N = 9–11) optic nerves from SD or KD fed Plp1tgB mice, showing h mean CAP traces ± SEM, I- V curves of i the CAP amplitude and j the CAP area (2way ANOVA with Sidak’s post test), and k nerve conduction velocity (NCV, two-sided Student’s t test). l Mean CAP area ± SEM after challenging optic nerves with 5 Hz stimulation in the presence of 3 mM lactate for 75 min, followed by recovery in the presence of 3 mM lactate and 10 mM glucose ( N = 7–8 optic nerves from Plp1tgB animals and 2 WT nerves). m Mean traces extracted from l (position marked by arrowheads) at baseline, at the end of the stimulation challenge, and after recovery. n Quantification of recovery showing mean CAP amplitude and CAP area with individual data points in relation to baseline (two-sided Student’s t test for each analysis) (* P < 0.05; ** P < 0.01; *** P < 0.001)
KD supports repair after cuprizone mediated demyelination.
a Treatment paradigms. In an acute paradigm cuprizone (CUP) in SD was fed for 4 weeks, followed by feeding SD or KD for 1 week. In the chronic paradigm cuprizone was fed for 12 weeks, followed by KD or SD for up to 6 weeks. Histochemical quantification of b oligodendroglial cells (Olig2) and c mature oligodendrocytes (CAII) in the corpus callosum (two-sided Student‘s t test of each treatment cohort with N = 4–5 mice). d Relative gene expression of Olig2, Car2 and Plp1 in dissected corpus callosum of mice after 12 + 2 weeks paradigm, normalized to untreated controls (set to 1, N = 4). Significance was evaluated by 1way ANOVA with Tukey’s post test, indicated are only significant differences between cuprizone groups. e Myelin abundance (Gallyas silver impregnation, N = 4–5) and f number of myelinated axons per square 100 µm in the corpus callosum ( N = 5–6). Data are expressed as mean with individual values (two-sided Student‘s t test of each treatment cohort). g Representative electron micrographs of the corpus callosum with boxed detail below (mitochondria colored in yellow, axonal outline colored in blue). Scale bar 2 µm. h Mean number of APP spheroids with individual values as a readout of axonal damage (two-sided Student‘s t test of each treatment cohort). i Mean size of axonal mitochondria in the corpus callosum ( N = 5 animals, 1way ANOVA with Tukey’s post test). j Beam test to measure motor performance ( N = 4–5), showing mean slips with individual data points (4 + 1, two-sided Student‘s t test) or mean ± SEM (12w-12 + 2 weeks, 2way ANOVA with Sidak’s post test) (* P < 0.05; ** P < 0.01; *** P < 0.001)
Therapy of Pelizaeus-Merzbacher Disease in Mice by Feeding a Cholesterol-Enriched Diet
G Saher et al.
Nat Med 18 (7), 1130-5.
Duplication of PLP1 (proteolipid protein gene 1) and the subsequent overexpression of the myelin protein PLP (also known as DM20) in oligodendrocytes is the most frequent …
Modulation of the Innate Immune Response by Human Neural Precursors Prevails Over Oligodendrocyte Progenitor Remyelination to Rescue a Severe Model of Pelizaeus-Merzbacher Disease
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Stem Cells 34 (4), 984-96.
Pelizaeus-Merzbacher disease (PMD) results from an X-linked misexpression of proteolipid protein 1 (PLP1). This leukodystrophy causes severe hypomyelination with progress …
Neurogenetics of Pelizaeus-Merzbacher Disease
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Handb Clin Neurol 148, 701-722.
Pelizaeus-Merzbacher disease (PMD) is an X-linked disorder caused by mutations in the PLP1 gene, which encodes the proteolipid protein of myelinating oligodendroglia. PMD …
Correction To: Ketogenic Diet Ameliorates Axonal Defects and Promotes Myelination in Pelizaeus-Merzbacher Disease
SK Stumpf et al.
Acta Neuropathol 138 (4), 673-674.
The original article was published.
Pelizaeus-Merzbacher Disease: Cellular Pathogenesis and Pharmacologic Therapy
T Torii et al.
Pediatr Int 56 (5), 659-66.
Pelizaeus-Merzbacher disease (PMD) is a rare leukodystrophy that causes severe dysmyelination in the central nervous system in infancy and early childhood. Many previous …
PubMed Central articles
Myelin Sheath Structure and Regeneration in Peripheral Nerve Injury Repair
B Liu et al.
Proc Natl Acad Sci U S A 116 (44), 22347-22352.
Observing the structure and regeneration of the myelin sheath in peripheral nerves following injury and during repair would help in understanding the pathogenesis and tre …
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