S100 Calcium-Binding Protein A16 Suppresses the Osteogenic Differentiation of Rat Bone Marrow Mesenchymal Stem Cells via Inhibiting SMAD Family Member 4 Signaling
Osteogenesis is a complex process of bone formation regulated by various factors, yet its underlying molecular mechanisms remain incompletely understood. This study aimed to investigate the role of S100A16, a novel member of the S100 protein family, in the osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs) and uncover a novel Smad4-mitogen-activated protein kinase (MAPK)/Jun N-terminal kinase (JNK) signaling axis. We herein evaluated the expression level of S100A16 in bone tissues and BMSCs from ovariectomized rats and then examined the impact of S100A16 silencing on osteogenic differentiation. Increased S100A16 expression was observed in bone tissues and BMSCs from ovariectomized rats, and S100A16 silencing promoted osteogenic differentiation. Further transcriptomic sequencing revealed that the Smad4 pathway was involved in S100A16 silencing-induced osteogenesis. The results of our Western blot analysis showed that S100A16 overexpression not only downregulated Smad4 but also activated MAPK/JNK signaling, which was validated by treatment with MAPK and JNK inhibitors U0126 and SP600125. Overall, this study elucidated the novel regulatory factors influencing osteogenic differentiation and provided mechanistic insights that could aid in the development of targeted therapeutic strategies for osteoporosis patients. Overall design: To investigate the function S100A16 in the regulation of osteogenesis, we isolated primary bone marrow stem cells from rats. After transfection of S100A16-specific siRNA during osteogenic differentiation, we  performed gene expression profiling analysis using data obtained from RNA-seq of S100A16 downregulated cells or control cells.