High-resolution mapping of osteoblast metabolism and bone matrix turnover in vivo

Understanding how osteoblasts build and remodel bone matrix in vivo remains a fundamental challenge because cellular metabolism and matrix turnover are difficult to resolve across time and space within mineralized tissues. Here, we developed an integrated imaging platform combining stable isotope labeling with correlative electron microscopy and nanoscale secondary ion mass spectrometry (NanoSIMS) to visualize bone cell metabolism and matrix dynamics at nanometer resolution in vivo. This approach revealed rapid incorporation of dietary amino acids into osteoblast subcellular compartments within minutes of oral administration, followed by deposition of newly labeled extracellular matrix within hours. By linking elemental composition, isotope incorporation, and ultrastructure, we further show that cellular phosphorus signal is associated with early osteoblast amino acid incorporation. Multiday labeling revealed that newly deposited matrix is spatially associated with local osteocyte process architecture. Long-term amino acid tracing uncovered localized matrix turnover at osteocyte and osteoclast interfaces, including osteocyte-associated pericellular matrix remodeling and osteoclast association with newly formed, old, and mixed matrix regions. Finally, aging was associated with reduced osteoblast amino acid incorporation, diminished matrix deposition, and impaired osteocyte process-associated activity. Together, this work establishes a high-resolution platform for linking bone cell metabolism with matrix deposition and turnover in vivo, providing a broadly adaptable strategy to investigate skeletal aging, tissue remodeling, and metabolic dysfunction in disease.