Extensive metabolic remodeling after limiting mitochondrial lipid burden is consistent with an improved metabolic health profile.

Mitochondrial lipid overload in skeletal muscle contributes to insulin resistance, and strategies limiting this lipid pressure improve glucose homeostasis; however, comprehensive cellular adaptations that occur in response to such an intervention have not been reported. Herein, mice with skeletal muscle-specific deletion of carnitine palmitoyltransferase 1b (Cpt1bM-/-), which limits mitochondrial lipid entry, were fed a moderate fat (25%) diet, and samples were subjected to a multimodal analysis merging transcriptomics, proteomics, and nontargeted metabolomics to characterize the coordinated multilevel cellular responses that occur when mitochondrial lipid burden is mitigated. Limiting mitochondrial fat entry predictably improves glucose homeostasis; however, remodeling of glucose metabolism pathways pales compared with adaptations in amino acid and lipid metabolism pathways, shifts in nucleotide metabolites, and biogenesis of mitochondria and peroxisomes. Despite impaired fat utilization, Cpt1bM-/- mice have increased acetyl-CoA (14-fold) and NADH (2-fold), indicating metabolic shifts yield sufficient precursors to meet energy demand; however, this does not translate to enhance energy status as Cpt1bM-/- mice have low ATP and high AMP levels, signifying energy deficit. Comparative analysis of transcriptomic data with disease-associated gene-sets not only predicted reduced risk of glucose metabolism disorders but was also consistent with lower risk for hepatic steatosis, cardiac hypertrophy, and premature death. Collectively, these results suggest induction of metabolic inefficiency under conditions of energy surfeit likely contributes to improvements in metabolic health when mitochondrial lipid burden is mitigated. Moreover, the breadth of disease states to which mechanisms induced by muscle-specific Cpt1b inhibition may mediate health benefits could be more extensive than previously predicted.

A low fat diet ameliorates pathology but retains beneficial effects associated with CPT1b knockout in skeletal muscle.

Inhibiting fatty acid oxidation is one approach to lowering glucose levels in diabetes. Skeletal muscle specific Carnitine Palmitoyltransferase 1b knockout mice (Cpt1bm-/-) comprise a model of impaired fat oxidation; and have decreased fat mass and enhanced glucose disposal and muscle oxidative capacity compared to controls. However, unfavorable effects occur relative to controls when Cpt1bm-/- mice are fed a 25% fat diet, including decreased activity and fat free mass and increased intramuscular lipid and serum myoglobin. In this study we explore if a low fat, high carbohydrate diet can ablate the unfavorable effects while maintaining the favorable phenotype in Cpt1bm-/- mice. Mice were fed either 10% fat (low fat) or 25% fat (chow) diet. Body composition was measured biweekly and indirect calorimetry was performed. Low fat diet abolishes the decreased activity, fat, and fat free mass seen in Cpt1bm-/- mice fed chow diet. Low fat diet also reduces serum myoglobin levels in Cpt1bm-/- mice and diminishes differences in IGF-1 seen between Cpt1bm-/- mice and control mice fed chow diet. Glucose tolerance tests reveal that glucose clearance is improved in Cpt1bm-/- mice relative to controls regardless of diet, and serum analysis shows increased levels of muscle derived FGF21. Electron microscopic analyses and measurements of mRNA transcripts show increased intramuscular lipids, FGF21, mitochondrial and oxidative capacity markers regardless of diet. The favorable metabolic phenotype of Cpt1bm-/- mice therefore remains consistent regardless of diet; and a combination of a low fat diet and pharmacological inhibition of CPT1b may offer remedies to reduce blood glucose.

Impaired Mitochondrial Fat Oxidation Induces FGF21 in Muscle.

Fatty acids are the primary fuel source for skeletal muscle during most of our daily activities, and impaired fatty acid oxidation (FAO) is associated with insulin resistance. We have developed a mouse model of impaired FAO by deleting carnitine palmitoyltransferase-1b specifically in skeletal muscle (Cpt1b(m-/-)). Cpt1b(m-/-) mice have increased glucose utilization and are resistant to diet-induced obesity. Here, we show that inhibition of mitochondrial FAO induces FGF21 expression specifically in skeletal muscle. The induction of FGF21 in Cpt1b-deficient muscle is dependent on AMPK and Akt1 signaling but independent of the stress signaling pathways. FGF21 appears to act in a paracrine manner to increase glucose uptake under low insulin conditions, but it does not contribute to the resistance to diet-induced obesity.

Diet-induced obesity and kidney disease - In search of a susceptible mouse model.

Obesity and metabolic syndrome are independent risk factors for chronic kidney disease, even without diabetes or hyperglycemia. Here, we compare two mouse models that are susceptible to diet-induced obesity: the relatively renal injury resistant C57BL/6J strain and the DBA2/J strain which is more sensitive to renal injury. Our studies focused on characterizing the effects of high fat diet feeding on renal oxidative stress, albuminuria, fibrosis and podocyte loss/insulin resistance. While the C57BL/6J strain does not develop significant pathological changes in the kidney, at least on lard based diets within the time frame investigated, it does show increased renal iNOS and nitrotyrosine levels and elevated mitochondrial respiration which may be indicative of mitochondrial lipid overfueling. Restricting the high fat diet to decrease adiposity decreased the levels of cellular oxidative stress markers, indicating that adiposity-related proinflammatory changes such as increased iNOS levels may trigger similar responses in the kidney. Mitochondrial respiration remained higher, suggesting that eating excess lipids, despite normal adiposity may still lead to renal mitochondrial overfueling. In comparison, DBA/2J mice developed albuminuria on similar diets, signs of fibrosis, oxidative stress, early signs of podocyte loss (evaluated by the markers podocin and WT-1) and podocyte insulin resistance (unable to phosphorylate their glomerular Akt when insulin was given). To summarize, while the C57BL/6J strain is not particularly susceptible to renal disease, changes in its mitochondrial lipid handling combined with the easy availability of transgenic technology may be an advantage to design new knockout models related to mitochondrial lipid metabolism. The DBA/2J model could serve as a basis for studying podocyte insulin resistance and identifying early renal markers in obesity before more severe kidney disease develops. Based on our observations, we encourage further critical evaluation of mouse models for obesity related chronic kidney disease.

Impaired mitochondrial fat oxidation induces adaptive remodeling of muscle metabolism.

The correlations between intramyocellular lipid (IMCL), decreased fatty acid oxidation (FAO), and insulin resistance have led to the hypothesis that impaired FAO causes accumulation of lipotoxic intermediates that inhibit muscle insulin signaling. Using a skeletal muscle-specific carnitine palmitoyltransferase-1 KO model, we show that prolonged and severe mitochondrial FAO inhibition results in increased carbohydrate utilization, along with reduced physical activity; increased circulating nonesterified fatty acids; and increased IMCLs, diacylglycerols, and ceramides. Perhaps more importantly, inhibition of mitochondrial FAO also initiates a local, adaptive response in muscle that invokes mitochondrial biogenesis, compensatory peroxisomal fat oxidation, and amino acid catabolism. Loss of its major fuel source (lipid) induces an energy deprivation response in muscle coordinated by signaling through AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) to maintain energy supply for locomotion and survival. At the whole-body level, these adaptations result in resistance to obesity.

Effect of intrastriatal mesenchymal stromal cell injection on progression of a murine model of Krabbe disease.

One of a family of devastating lysosomal storage disorders, Krabbe disease is characterized by demyelination, psychosine accumulation, and inflammation. Affected infants rarely survive longer than 2 years. Using the twitcher mouse model of the disease, this study evaluated the potential of intrastriatal injection of adipose or bone marrow-derived mesenchymal stromal cells (MSCs) as a treatment option. Neonatal pups were injected with MSCs at 3-4 days of age and subjected to a battery of behavioral tests beginning at 15 days. While MSC injection failed to increase lifespan of twitchers, improvements in rotarod performance and twitching severity were observed at 27-38 days of age using MSCs derived from bone marrow. This study tested several different tasks developed in adult mice for evaluation of disease progression in immature twitchers. Rotarod was both reliable and extremely sensitive. Automated gait analysis using the Treadscan program was also useful for early evaluation of differences prior to overt gait dysfunction. Finally, this study represents the first use of the Stone T-maze in immature mice. Validation of rotarod and automated gait analysis for detection of subtle differences in disease progression is important for early stage efforts to develop treatments for juvenile disorders.