Ketogenic Diet Ameliorates Dysferlinopathy Phenotype in Bla/J Mice by Promoting Mitochondrial Function

Grant Duration
03/20 – 12/23

Objective

Determine whether the induction of ketosis in pre-symptomatic and symptomatic Bla/J mice by a ketogenic diet reduces motor function loss by improving mitochondrial function.

Background

The ketogenic diet, introduced into practice in the 1920s by Russell Wilder to treat epilepsy, consists of a high-fat, low-carbohydrate diet that induces production of ketone bodies (acetoacetate and β-hydroxybutyrate) in the liver through fat metabolism. The ketogenic diet and especially β-hydroxybutyrate (β-HB) are gaining attention due to their therapeutic potential. β-HB reduces oxidative stress, inflammation and protein degradation. The ketogenic diet has been shown to preserve motor function in aged mice by activation of mitochondrial biogenesis, oxidative metabolism and antioxidant capacity. In a mouse model of dysferlinopathy (Bla/J) we found that a continuous 3-month ketogenic diet starting right before the disease onset (5-month-old mice) improves motor function and muscle strength to the levels observed in WT mice. This physical improvement correlates with an increase in mitochondrial mass, suggesting a rise in mitochondrial biogenesis and/or activity.

Our goal in this year’s project is to determine whether the beneficial effects of the ketogenic diet can be sustained over a longer time-period. Also, we want to see if a ketogenic diet can reverse muscle function loss or prevent further muscle deterioration once the clinical manifestations of dysferlinopathy appear. This answer is of the utmost importance as most people with dysferlinopathy are diagnosed after they already have symptoms. In this project we will address these questions by feeding pre-symptomatic and symptomatic Bla/J a ketogenic diet for 6 months. We will measure motor performance monthly and evaluate mitochondrial function at the end of the experiment using the Seahorse assay.

Project Results

In muscle, mitochondria play a fundamental role in maintaining bioenergetic homeostasis. Mitochondrial dysfunction has been associated with several muscle disorders and mitochondrial function has been shown to be reduced in dysferlinopathy. It is unknown whether this is the result of mitochondrial failure or a metabolically unfavorable, sugar-rich environment. Using a cell culture model of dysferlinopathy, we determined that the absence or the reduction of glucose in the culture media improved mitochondrial function. We observed similar results when the cells were cultured with β-hydroxybutyrate as the main energy substrate.

These experiments demonstrate that mitochondria are not dysfunctional in cells lacking dysferlin, but they turn off. To determine whether the activation of mitochondria by changes in the energetic substrate impact in the survival of the cells after a mechanical stress, we challenged cells cultured in different media. Interestingly, only the dysferlinopathy cells cultured in a high glucose media had a significant death rate, while cells cultured in β-hydroxybutyrate or in galactose (no glucose) survived at a similar rate to cells with dysferlin. We concluded that the activation of mitochondria, achieved by a change in food source, helps dysferlin-deficient cells cope with stress.

We also investigated whether a reduction in dietary carbohydrate improves muscle perform in a mouse model of dysferlinopathy (Bla/J). We fed pre-symptomatic mice either a ketogenic diet (low carbohydrate-high fat), an isocaloric control diet (high carbohydrate-low fat), or regular chow for three months. We then assessed muscle strength using the grip test, motor coordination using a rotarod, and endurance on a treadmill. We found that the mice fed with a ketogenic diet maintained or improved their strength or motor coordination performance to the level shown by the wild type mice. This correlates with an increase in the mitochondrial mass of different muscles, as determine by an increase of the mitochondrial marker VDAC by Western blot. Overall, we concluded that removing carbohydrate as the main energetic substrate, and replacing it with ketone bodies, improves mitochondrial function and therefore resistance to stress and muscle performance.

By J. Cesar Cardenas PhD., Universidad Mayor, Santiago, Chile|2024-07-08T19:14:55+00:00October 1st, 2023|Past Projects|Comments Off

About the Author: J. Cesar Cardenas PhD., Universidad Mayor, Santiago, Chile

J. Cesar Cardenas earned his Ph.D. from the University of Chile, where he studied the mechanism that regulated nuclear Ca2+ under Dr. Enrique Jaimovich’s mentoring. Then he moved to the University of Pennsylvania Medical School where he explored the nuclear localization 1,4,5-trisphosphate receptor (InsP3R) Ca2+channel by using high resolution electron microscopy and cryofracture as a postdoctoral fellow with Dr. Clara Franzini-Armstrong. Motivate in gain a better understanding of the physiological role of the InsP3R he joined Dr. Kevin Foskett’s lab also at UPENN, where he developed a strong interest in the regulation of cellular metabolism and bioenergetics by Ca2+. He was the first to show that basal constitutive low-level Ca2+ signaling by the InsP3R, is essential to maintain the sufficient mitochondrial NADH production to support oxidative phosphorylation in resting cells. In the absence of this calcium signaling, cells become metabolically compromised and a pro-survival AMPK-dependent mTOR-independent autophagy is turned on. He joined the Department of Anatomy and Cell Developmental Biology Program at the Institute of Biomedical Science of the University of Chile School of Medicine in March of 2012.