In situ insights into spatial and temporal expressions of mesenchymal stem cell markers during human skeletal...

Bone formation is a complex process involving multiple cell types and molecular signals that regulate osteogenesis from three embryological origins. Depending on the anatomical site, bone develops through either intramembranous or endochondral ossification. Although skeletal development has been extensively studied, the spatial and temporal expression of proteins during endochondral ossification, particularly according to anatomical site and cellular compartment, remains incompletely understood. In this descriptive study, we evaluated the in-situ expression of cell surface proteins previously described as mesenchymal stem cell markers-other than those proposed by the International Society for Cellular Therapy (CD105, CD90, and CD73)-during human skeletal development. Protein expression was analyzed in a spatiotemporal manner during endochondral ossification in axial and appendicular skeletal elements from human embryos, fetuses, and post-term individuals across the three trimesters of gestation. Our findings identified strong expression of CD166, CD271, and Podoplanin in chondroblasts of vertebral bodies and mesenchymal cells of the autopod during the first trimester. In contrast, CD146, CD29, CD44, and STRO-1 were not detected in the first trimester but appeared after week 13, predominantly within the extracellular matrix; therefore, not associated with early osteochondroprogenitors. Overall, this study provides novel insights into the anatomical and developmental distribution of seven cell surface markers during human skeletogenesis. Notably, these proteins were detected in cellular compartments beyond the plasma membrane, including the extracellular matrix, and exhibited distinct spatial and temporal expression patterns during gestation.