Beta-hydroxy-beta-methylbutyrate (HMB) improves daily activity and whole-body protein metabolism in Duchenne muscular dystrophy dogs: a pilot study

Abstract

Duchenne muscular dystrophy (DMD) is a severe neuromuscular disease due to loss of dystrophin, leading to progressive muscle wasting and physical inactivity. In this pilot study, we studied the effect of daily supplementation of the anabolic substrate beta-hydroxy-beta-methylbutyrate (HMB) on whole body protein and amino acid kinetics using novel isotope methods and daily activity in a canine model of DMD. Six DMD dogs were administered 3 g daily of HMB or placebo for 28 days according to a randomized, placebo-controlled, double-blinded crossover design. We measured pre- and post-intervention daily activity, biochemistry markers, and whole-body amino acid kinetics. We tracked daily activity with an activity monitoring device and measured plasma creatine kinase and standard clinical biochemistry panels to monitor muscle and organ function. To calculate whole body and intracellular amino acid production, we administered in the postabsorptive state an IV stable isotope solution containing 20 amino acid tracers. We collected blood before and six times after until two hours post tracer pulse administration and measured amino acid enrichments and concentrations by LC-MS/MS, subsequently followed by (non) compartmental modeling of the decay enrichment curves. Results were expressed as mean with 95% CI. Whole body production, plasma concentrations, and intra-/extracellular compartmental analyses of various amino acids were attenuated in HMB-dosed DMD dogs. Specifically, the plasma concentration of hydroxyproline (marker of collagen breakdown) was significantly higher in the placebo group compared to the HMB group. The intra- and extracellular pool sizes and flux between the two compartments of hydroxyproline was reduced in HMB treated dogs. DMD dogs treated with HMB as compared to placebo had a respective 40% increase in exertional (play) (951 [827, 1075] versus 569 [491, 647]; p < 0.0001) and 10.5% increase in non-exertional (active) activity (15,366 [14742, 15990] versus 13,806 [13148,14466]; p = 0.0016). In addition, a 6% reduction was found in rest time after HMB supplementation compared to placebo (23,857 [23,130, 24,584], versus 25,363 [24500, 26225]; p = 0.0122). Creatine kinase was not statistically different between groups. We did not observe any adverse clinical or biochemical-related effects of HMB dosing. Daily HMB supplementation in DMD dogs can safely and positively influence protein and amino acid metabolism and improve overall daily activity.

Keywords: Dog; Duchenne; Dystrophin; Dystrophy; HMB; Muscle.

Fig. 1

Schematic of the study design. Dogs were dosed in a blinded, placebo-controlled cross-over study for 28 days, washed out for at least 2 weeks, and then crossed over to the opposite dosing regimen (HMB or placebo). Activity was monitored for the entire duration of the cross-over studies. Amino acid isotope pulsing and biochemistry panel analyses were performed at baseline (0d) and endpoint (28d) for each dosing regimen.

Fig. 2

Forrest plot of whole-body production (WBP) and interconversions of amino acids (AAs). Hydroxyproline, arginine, glycine, leucine, methionine, ornithine, proline, and valine were significantly reduced, while glutamate and isoleucine were increased in the HMB group. Data portrayed as a % difference from the placebo-treated group. Open blue circles were placebo, closed red circles were HMB. Asterisks = statistically different compared to placebo. N = 6 per group.

Fig. 3

Activity levels improved for HMB-treated DMD dogs compared to placebo. 3(A) AUC for play (high exertional) activity. 3(B) AUC analysis bar graph for play activity. 3(C) AUC for active (non-exertional) activity. 3(D) AUC analysis bar graph for active activity. 3(E) AUC for rest (not moving/sleeping). 3(F) AUC analysis bar graph for rest. Data were expressed as mean [95% CI] over a 28-day period. N = 6 per group.

Fig. 4

Creatine kinase levels in each dosing group. U/L = units per liter. N = 6 per group. Single data points are now included. Male = square; female = circles. Data were expressed as mean [95% CI]. Results were statistically not significant.