Tartrate-resistant acid phosphatase/ACP5 in bone development and maintenance : age- and sex-specific skeletal phenotypes in mice

<p dir="ltr">The skeletal system is a dynamic organ network, integrating mechanical, hormonal, and molecular cues to continuously balance bone formation and resorption. Central to this homeostatic process is the interplay between osteoblasts, osteoclasts, and osteocytes, which orchestrate modelling and remodelling in response to developmental, metabolic, and mechanical stimuli. Among the osteoclast-derived factors contributing to skeletal regulation, tartrate-resistant acid phosphatase (TRAP/ACP5) has long been used as a histochemical marker of bone resorption. However, its broader physiological roles remain underexplored. Previous studies have identified TRAP as a marker of osteoclast activity and implicated it in bone resorption and immune-skeletal interactions. Building on this foundation, this doctoral thesis characterizes the multifaceted role of TRAP in bone biology using a murine knockout model, contributing significantly to the growing knowledge of its critical involvement in skeletal growth, structural maintenance, and mechanoadaptation across age, sex, and anatomical site.</p><p dir="ltr">Through cross-sectional analyses combining high-resolution micro-computed tomography (uCT), histomorphometry, in vivo mechanical loading, and in vitro functional assays, we show that TRAP deficiency (TRAP-/-) leads to a spectrum of skeletal abnormalities. In young adult mice, TRAP deletion resulted in significantly shorter tibiae and altered cortical and trabecular architecture. These defects were sex- and site-specific: male TRAP-/- mice exhibited increased trabecular bone mass, thicker cortices, and wider metaphyseal zones, whereas female TRAP-/- mice showed pronounced disruption in growth plate organization but relatively modest changes in bone mass. Importantly, the growth plate of TRAP-/- mice demonstrated increased hypertrophic zone height and reduced chondrocyte column alignment, indicating impaired endochondral ossification.</p><p dir="ltr">The role of TRAP in facilitating load-induced bone formation was assessed using strain-matched in vivo axial tibial loading in 16-week-old male mice. Wild-type animals displayed robust anabolic responses including periosteal expansion, cortical thickening, increased trabecular volume, and enhanced growth plate bridging. These load-responsive adaptations were absent in TRAP-/- mice, suggesting a critical requirement for TRAP in converting mechanical signals into bone-forming stimuli. Mechanical pre-loading confirmed that TRAP-/- tibiae were stiffer and required greater compressive force to reach target strain, yet the downstream skeletal response remained blunted. Serum bone turnover markers such as alkaline phosphatase (ALP) and procollagen type 1 N-terminal propeptide (PINP) were unchanged, suggesting that TRAP's mechanoadaptive role is not due to global osteoblast dysfunction but may involve osteocyte signalling, matrix interactions, or osteoclast-osteoblast coupling.</p><p dir="ltr">Extending these findings to aged mice, 18-month-old male and female TRAP-/- mice displayed persistent reductions in bone length, a legacy of developmental impairment. Nonetheless, sex-specific cortical and trabecular phenotypes were evident: aged males exhibited higher cortical and trabecular bone, while females showed higher cortical density and narrowed marrow cavities compared to wildtype mice. The adaptive skeletal response to mechanical loading was abolished in both sexes. uCT analyses revealed no significant loading-induced gain in any bone compartment in aged TRAP-/- mice of both sexes, establishing TRAP's indispensable role in skeletal mechanosensitivity throughout life.</p><p dir="ltr">Complementary in vitro assays further demonstrated that TRAP-deficient osteoclast progenitors exhibited reduced osteoclastogenesis in response to mechanical stimulation and aberrant ATP signalling particularly in males. These results suggest that TRAP may mediate key mechanosensitive processes through purinergic signalling pathways or redox modulation of the extracellular matrix. Notably, reduced growth plate bridging in loaded young TRAP-/- mice implicates TRAP in cartilage-to-bone transition and epiphyseal integration under mechanical challenge. In contrast, no loading-induced changes in bony bridging were observed in aged mice, suggesting an age-dependent mechanoadaptive role of TRAP.</p><p dir="ltr">Taken together, these data expand our understanding of TRAP as a multifaceted regulator of bone biology, highlighting previously underappreciated roles in skeletal growth, remodelling, and mechanoadaptation. Beyond its canonical role in mineralized matrix degradation, TRAP is required for orchestrating structural adaptation to mechanical load, maintaining growth plate integrity, and coordinating site- and sex-specific skeletal responses during growth and aging. This work suggests TRAPs role as a mechanotransductive enzyme that links osteoclast function with broader skeletal plasticity, revealing new dimensions in the regulation of bone remodelling. The findings presented in this thesis argue for the reconsideration of TRAP's role in bone beyond resorption, with implications for therapeutic targeting in disorders such as osteoporosis, osteopetrosis, and age-related bone fragility, where mechanical responsiveness and bone turnover are impaired.</p><h3>List of scientific papers</h3><p dir="ltr">1) <b>Bhavik Rathod</b> , Suchita Desai , Hasmik Jasmine Samvelyan , Laura Bock , Jianyao Wu , Claes Ohlsson, Anders Palmquist , Jessica J. Alm , Phillip T. Newton , Goran Andersson, Sara H. Windahl. Tartrate-resistant acid phosphatase (TRAP/ACP5) promotes bone length, regulates cortical and trabecular bone mass, and maintains growth plate architecture and width in a sex- and site-specific manner in mice. Bone. 2024;188:117223. <a href="https://doi.org/10.1016/j.bone.2024.117223">https://doi.org/10.1016/j.bone.2024.117223</a></p><p dir="ltr">2) <b>Bhavik Rathod</b>, Hasmik Jasmine Samvelyan, Suchita Desai, Laura Bock, Nicole Gustafsson, Jianyao Wu, Claes Ohlsson, Per Magnusson, Göran Andersson, Sara H Windahl. Tartrate-resistant acid phosphatase (TRAP/ACP5) augments the bone anabolic response to mechanical loading in male mice. JBMR Plus. 2025;9(7):ziaf073. Published 2025 Apr 23. <a href="https://doi.org/10.1093/jbmrpl/ziaf073">https://doi.org/10.1093/jbmrpl/ziaf073</a></p><p dir="ltr">3) <b>Bhavik Rathod</b>, Jasmine Samvelyan, Nicole Gustafsson, Aneta Liszka, Narelle McGregor, Jianyao Wu, Claes Ohlsson, Anna Fahlgren, Natalie Sims, Jonas Fuxe, Goran Andersson, Jessica J Alm*, Sara H Windahl *. Tartrate acid phosphatase (TRAP/ACP5) sex-specifically regulates trabecular bone maintenance, and the bone anabolic effects of mechanical loading in old mice. [Manuscript]</p>