Simulated Microgravity Reduces Growth but Preserves Bone-Relevant Bioactivity of Medicago truncatula

Microgravity exposure during spaceflight leads to profound physiological adaptations, including accelerated skeletal loss driven by imbalances in bone remodeling. Plants grown in altered gravity environments exhibit changes in growth, development, and secondary metabolism that may influence their potential as nutritional or biological countermeasures. In this study, Medicago truncatula seedlings were grown for 14 days under stationary 1g or simulated microgravity using a clinostat. Plant growth parameters were quantified, total flavonoid and isoflavonoid concentrations determined, and ethanolic extracts evaluated for bioactivity using murine osteoclast precursor (RAW264.7) and osteoblast-like (MC3T3-E1) cell models. Simulated microgravity significantly reduced root length, shoot length, lateral root number, and fresh mass relative to 1g controls while increasing total flavonoid and isoflavonoid concentrations by 45% and 64%, respectively. Extracts from both gravity conditions significantly inhibited RAW264.7 cell proliferation with no statistically significant differences between microgravity and 1g-derived extracts. MC3T3-E1 cell proliferation was not affected by extract exposure, while alkaline phosphatase activity showed modest induction relative to solvent controls without a consistent dose-dependent or gravity-specific trend. These findings demonstrate that simulated microgravity alters plant growth but does not confer gravity-specific differences in extract bioactivity on bone cell models, supporting selective effects on osteoclast-related pathways.