Conserved skeletal muscle transcriptomic responses to pacing strategies in Thoroughbred horses.

Conserved patterns of gene expression changes occur in the skeletal muscles of Thoroughbred horses after high-intensity exercise, regardless of different pacing strategies, with only a few genes showing distinct responses based on pacing.

Study Objective and Design

• The research aimed to determine if different pacing strategies during high-intensity exercise cause different changes in gene expression in horse skeletal muscle.
• Eight Thoroughbred horses participated in a randomized crossover study involving two treadmill exercise sessions.
• Two pacing conditions were tested:
  • Positive-pacing: 1 minute at 110% of maximal oxygen uptake (V˙O₂max), then 1 minute at 90% V˙O₂max.
  • Negative-pacing: 1 minute at 90% V˙O₂max, then 1 minute at 110% V˙O₂max (reverse order).

Gene Expression Analysis After Exercise

• Muscle biopsies were collected 4 hours post-exercise to assess changes in gene expression.
• Positive-pacing protocol results:
  • 1,989 genes showed increased expression (upregulation).
  • 840 genes showed decreased expression (downregulation).
• Negative-pacing protocol results:
  • 1,710 genes were upregulated.
  • 593 genes were downregulated.
• Statistical criteria for gene expression changes:
  • False discovery rate (FDR) less than 0.05 for significance.
  • Minimum fold change of 1.5 in expression level.

Shared Gene Expression Response

• Most genes altered by exercise were common to both pacing strategies.
• Shared pathways affected included:
  • Hypoxia signaling, which relates to the response to low oxygen during intense exercise.
  • Extracellular matrix remodeling, important for structural tissue adaptation and repair.
  • Metabolic regulation pathways, which adjust energy production and utilization.

Differences Between Pacing Protocols

• Direct comparison of the two pacing protocols revealed a small subset of genes with different expression patterns:
  • Four genes were more highly expressed after positive pacing:
    • RP1 (Retinitis pigmentosa 1 protein)
    • MORN5 (MORN repeat containing 5 gene)
    • Two unannotated equine transcripts (ENSECAG00000060378 and ENSECAG00000057614)
  • Five genes had higher expression after negative pacing:
    • OLFML2B (Olfactomedin-like 2B)
    • POSTN (Periostin)
    • ANGPTL1 (Angiopoietin-like 1)
    • MAP1A (Microtubule-associated protein 1A)
    • ZNF554 (Zinc finger protein 554)
• These genes could be involved in subtle differences in muscle adaptation or recovery related to pacing order but do not dominate the overall response.

Conclusions and Implications

• The skeletal muscle gene expression response to matched high-intensity exercise work is largely the same regardless of pacing strategy in Thoroughbred horses.
• Only a few genes showed pacing-related differences, suggesting the muscle’s transcriptional programs are robust to pacing pattern variations.
• This understanding may help in optimizing training and performance strategies in racehorses by showing pacing pattern has limited influence on muscle molecular adaptation shortly after exercise.
• Future research could explore the functional roles of the few differentially expressed genes and examine if these minor differences affect longer-term muscle adaptation or performance outcomes.