Osteoclasts are large multinucleated cells exquisitely adapted to degrade bone matrix. Upon contact with bone, osteoclasts segregate their surface membrane into four polarised domains: 1) the sealing zone; 2) the basolateral membrane; 3) functional secretory domain (FSD) and; 4) the bone-apposed ruffled border (RB). The RB functions as the resorptive apparatus, serving as a release site for protons and osteolytic enzymes (i.e. cathepsin K) required to digest the mineral and organic phases of bone. At the same time, the RB facilitates the uptake and transcytosis of bone matrix by-products to the opposing FSD where they are expelled into the extracellular milieu. Although the crucial importance of vesicular trafficking between the RB and FSD during the functional bone resorption cycle is now well-established, the spatiotemporal dynamics of this process has not yet been appreciated in real-time. Here using confocal microscopy, we have monitored the intracellular uptake and trafficking dynamics of degraded bone matrix in osteoclasts actively engaged in bone resorption. Using fluorescently labelled bone substrates together with a panel of intracellular compartment markers we demonstrate that osteoclasts utilise multiple endo-lysosomal trafficking pathways to ingest degraded bone matrix particles at the RB. In addition, we show that osteoclasts employ, pseudopodal-like actin-rich projections to facilitate phagocytic bone uptake. Following phagocytosis, bone particles undergo further degradation and associate with both lysosomal and autophagic pathways before converging with the transcytotic pathway en route to the FSD. Overall, these studies provide new insights into the uptake, trafficking and dynamics of degraded bone matrix during the bone resorption cycle.
Disclosure: The authors declared no competing interests. This work was funded in part by grants awarded to NJP by the National Health Research and Council of Australia.