Lysosomal Degradation Regulates Sclerostin Protein Abundance in Response to Mechanical Stress and Parathyroid Hormone Treatment in Osteocytes

Abstract

The development of low bone mass during aging and disuse increases fracture risk and represents an important health concern in many populations. Most current therapeutics slow bone loss by targeting and slowing bone resorption. Yet, they do not target a fundamental issue of a lack of de novo bone formation to reform lost bone. While the Wnt/-catenin inhibitor, sclerostin, is critical to the activation of new bone formation, its precise molecular control and the molecular basis of bone mechano-sensing has remained elusive. We recently described an osteocyte mechano-transduction cascade that requires production of reactive oxygen species (ROS) by NADPH Oxidase 2 (NOX2). Subsequent calcium influx through transient receptor potential vanilloid 4 (TRPV4) activates calcium/calmodulin-dependent kinase II (CaMKII), which is necessary for rapid sclerostin protein loss. Here, we set out to determine how fluid shear stress (FSS) rapidly controls sclerostin protein abundance and if this regulation is conserved with another bone anabolic stimulus, parathyroid hormone (PTH). We found that both FSS and PTH regulate the rapid lysosomal degradation of sclerostin protein on a minutes’ timescale. Both FSS and PTH activate CaMKII, which regulates lysosomal activity, indicating increased lysosomal flux following bone anabolic stimulation. We then evaluated the physiological relevance of lysosomal sclerostin degradation to load-induced bone formation in vivo. Using an ulnar loading model, we found that lysosomal function is necessary for load-induced sclerostin degradation and subsequent bone formation. We found that disrupted lysosomal function in the lysosomal storage disorder Gaucher Disease leads to accumulation of sclerostin in diseased iPSCs and that this accumulation is reversed when lysosomal function is restored. We additionally found that NOX2 ROS are necessary for load-induced sclerostin degradation and bone formation in vivo, spurring the analysis of a bone-specific NOX2 conditional deletion mouse model. While we are still evaluating the consequence of NOX2 conditional deletion in cells of the osteoblast/osteocyte lineage to the in vivo bone mechano-response, we find that female mice have mild osteopenia. In total, these results will expand our understanding of osteocyte mechano-sensing and fill the knowledge gaps as to how osteocytes sense and transduce mechanical signals and regulate sclerostin abundance in vivo.