Vitamin B12 supports skeletal muscle oxidative phosphorylation capacity in male mice

Abstract:

Background
Vitamin B12 is a cofactor in folate-mediated one-carbon metabolism (FOCM), which generates nucleotides (thymidylate (dTMP) and purines) and methionine. Depressed de novo thymidylate (dTMP) synthesis leads to uracil accumulation in DNA.

Objective
The purpose of this study was to determine how B12 availability affects mitochondrial DNA (mtDNA) integrity and mitochondrial function in skeletal muscle. B12 deficiency was modeled in young-adult mice. Intramuscular B12 injection in aged mice assessed the role of B12 supplementation in age-related changes in skeletal muscle.

Methods
Male methionine synthase knockdown (Mtr+/-) and wild-type littermates (Mtr+/+) were weaned to either an AIN93G-based control (C) diet containing 25 μg/kg vitamin B12 (Mtr+/+, n=8; Mtr+/-, n=9) or a B12-deficient (-B12) diet containing 0 μg/kg vitamin B12 (n=9 per genotype) for seven weeks. Aged (20-22mo) male C57BL/6N mice were acclimated to an AIN93G control diet four weeks, then received either weekly injections of saline (vehicle control [30 uL 0.9% NaCl], n=5) or B12 (0.65 μg per 30uL 0.9% NaCl; n=6) in each of two hindleg muscles [1.25 μg B12 total]) for eight weeks. Outcomes measured included maximal oxygen consumption rate (OCR), uracil in mtDNA (a biomarker of mtDNA integrity), mtDNA copy number, and mitochondrial mass. Data were analyzed using a two-way ANOVA in the Mtr+/- mouse model exposed to B12-deficient diets and by a student’s t-test for B12 supplementation in aged mice.

Results
The tibialis anterior (TA) muscle from Mtr+/- mice exhibited 50% lower (p=0.01) maximal respiratory capacity of the electron transport chain than did TA from Mtr+/+ mice. Exposure to the -B12 diet lowered maximal capacity of complex I in mitochondrially rich muscle (soleus and mitochondria-rich portions of quadriceps and gastrocnemius) by 25% (p=0.02). Uracil in mitochondrial DNA (mtDNA) in red muscle and gastrocnemius was elevated ∼10 fold with exposure to -B12 diet (p=0.04 and p<0.001, respectively). In aged mice, gastrocnemius complex IV activity was increased 2-fold with intramuscular B12 supplementation (p=0.04).

Conclusions
Exposure to a B12-deficient diet led to uracil accumulation in mtDNA and impaired maximal oxidative capacity in skeletal muscle. B12 supplementation improved complex IV maximal capacity in gastrocnemius from aged mice, a model of age-related skeletal muscle decline.