Ninjin’yoeito Kampo medicine enhances zebrafish endurance under forced-swimming conditions via muscle hypertrophy

Maintaining skeletal muscle mass and function is essential for extending healthy life expectancy in aging populations. While exercise and nutritional support are primary strategies to counteract sarcopenia and frailty, their effectiveness is often limited in the elderly. Thus, pharmacological and dietary interventions that directly enhance muscle formation have attracted increasing attention. Ninjin’yoeito (NYT), a traditional Japanese Kampo medicine composed of 12 medicinal herbs, has recently been highlighted for its antifrailty potential. In this study, we examined the effects of NYT dietary supplementation on muscle hypertrophy and endurance in zebrafish. NYT administration did not alter food intake or spontaneous locomotion under standard conditions, but it markedly improved endurance performance in a forced swimming test. NYT-fed zebrafish displayed prolonged upstream swimming times and delayed fatigue onset. Histological analysis revealed hypertrophy of fast-twitch muscle fibers, and gene expression profiling demonstrated upregulation of myhz2, actn3, and myod, which are key regulators of fast muscle development and differentiation. In contrast, genes associated with slow-twitch fibers and canonical protein synthesis pathways were unaffected. Furthermore, ldh, a pivotal enzyme in anaerobic glycolysis, was significantly upregulated in NYT-treated fish following exercise, suggesting enhanced energy metabolism. Collectively, these findings indicate that NYT promotes fast-twitch muscle hypertrophy and enhances physical performance, supporting its potential as a functional intervention to prevent or attenuate age-related decline in muscle health.