Synergistic Effect of Stauntonia hexaphylla (Thunb.) Decne Fruit and Leaf on RAW 264.7 Osteoclast and MC3T3-E1 Osteoblast Differentiation

Stauntonia hexaphylla (Thunb.) Decne (SH) is known for its anti-inflammatory, analgesic, antioxidant, and anti-osteoporotic properties. This study investigated the composition of SH leaves and fruits and assessed their individual and combined effects in an in vitro osteoporosis model. Extracts with varying leaf-to-fruit ratios (SH82, SH55, SH28) were evaluated in MC3T3-E1 and RAW264.7 cells to examine osteogenesis and bone resorption biomarkers. SH leaves were rich in chlorogenic acids (CGAs) and flavonoids, while fruits contained phenolic acids with minimal flavonoids. Leaf extracts exhibited superior antioxidant activity and ROS suppression. Both leaf and fruit extracts enhanced ALP activity, calcium deposition, and collagen formation in MC3T3-E1 cells, with leaf extracts demonstrating greater efficacy. Additionally, osteoblastogenesis-related genes were upregulated, while TRAP activity and osteoclast-related gene expression were significantly inhibited. The combined extract exerted a synergistic effect, with SH28 showing the most pronounced osteogenic enhancement and TRAP inhibition. Key components, including neo-CGA, CGA, rutin, and luteolin-7-O-glucoside, positively influenced ALP and TRAP activities. These findings highlight the potential of SH, particularly at a high leaf-to-fruit ratio, as a promising natural agent for osteoporosis prevention.