Ctsk + Osteoclasts Orchestrate Condylar Morphogenesis via Hypoxic Lysosome

While the biological functions and molecular mechanisms of osteoclasts in condylar morphogenesis remain incompletely characterized, and the regulatory effects of hypoxic conditions within the deep condyle on osteoclast-mediated alveolar bone remodeling are poorly understood, this investigation systematically examines the hypoxia response mechanisms of cathepsin K–positive (ctsk + ) osteoclasts and their specialized functions in condylar tissue organization. Genetic ablation of ctsk+ cells in diphtheria toxin receptor (DTR) transgenic mice resulted in pronounced accumulation of calcified cartilage within the mandibular condylar cartilage (MCC) accompanied by impaired subchondral bone formation—phenotypes directly attributable to osteoclast deficiency. Intriguingly, DTR animals exhibited concurrent upregulation of chondroblast markers, osteoblastic factors, and bone resorption mediators, suggesting that ctsk+ cells normally function to restrain MCC proliferation and premature hypertrophy while simultaneously inhibiting subchondral osteogenesis. Conditional knockout of hypoxia-inducible factor-1α (HIF-1α) specifically in ctsk+ cells (HIF-1α ∆ctsk-cre ) replicated the cartilage accumulation phenotype observed in DTR mice during early development (≤5 wk), with comparable disorganization of the fibrocartilage layers. However, after 6 wk, HIF-1α ∆ctsk-cre mutants displayed paradoxical cartilage reduction coupled with accelerated subchondral mineralization. We propose that progressive osteoclast depletion, combined with extensive chondrocyte apoptosis and elevated matrix metalloproteinase 13 expression, drives the gradual MCC thinning in knockout animals. The sustained osteoclast deficiency appears to disrupt the physiological cartilage-to-bone transition, ultimately causing delayed but exaggerated subchondral mineralization. At the cellular level, HIF-1α deficiency in ctsk+ osteoclasts induced profound structural abnormalities, including disorganized ruffled borders and defective lysosomal biogenesis. Mechanistically, we identified the osteoclast-specific TSC2-mTORC1-TFEB axis as a critical regulator of hypoxia-responsive lysosomal formation. Collectively, these findings establish a novel dual regulatory role for ctsk+ cells: suppressing chondrocyte proliferation, premature hypertrophy, and osteogenesis through bone resorption signaling and mediating HIF-1α–dependent control of osteoclastogenesis and lysosomal function during calcified cartilage degradation.