The adaptation of the skeleton to its mechanical environment is orchestrated by mechanosensitive osteocytes, largely by regulating the abundance of sclerostin, a secreted inhibitor of bone formation. We defined a microtubule-dependent mechanotransduction pathway that linked fluid shear stress to reactive oxygen species (ROS) and calcium (Ca2+) signals that led to a reduction in sclerostin abundance in cultured osteocytes. We demonstrated that microtubules stabilized by detyrosination, a reversible posttranslational modification of polymerized α-tubulin, determined the stiffness of the cytoskeleton, which set the mechanoresponsive range of cultured osteocytes to fluid shear stress. We showed that fluid shear stress through the microtubule network activated NADPH oxidase 2 (NOX2)–generated ROS that target the Ca2+ channel TRPV4 to elicit Ca2+ influx. Furthermore, tuning the abundance of detyrosinated tubulin affected cytoskeletal stiffness to define the mechanoresponsive range of cultured osteocytes to fluid shear stress. Finally, we demonstrated that NOX2-ROS elicited Ca2+ signals that activated the kinase CaMKII to decrease the abundance of sclerostin protein. Together, these discoveries may identify potentially druggable targets for regulating osteocyte mechanotransduction to affect bone quality.

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Objectives This study sought to demonstrate that short-term cardiac unloading with a left ventricular (LV) assist device (LVAD) after acute myocardial infarction (MI) can conserve calcium cycling and improve heart function. Background Heart failure secondary to MI remains a major source of morbidity and mortality. Alterations in calcium cycling are linked to cardiac dysfunction in the failing heart. Methods Adult Dorsett hybrid sheep underwent acute MI and were mechanically unloaded with an axial-flow LVAD (Impella 5.0) for 2 weeks (n = 6). Six sheep with MI only and 4 sham sheep were used as controls. All animals were followed for 12 weeks post-MI. Regional strains in the LV were measured by sonomicrometry. Major calcium-handling proteins (CHPs), including sarco-/endoplasmic reticulum calcium ATPase-2α (SERCA-2α), Na+-Ca2+ exchanger-1, and phospholamban, and Ca2+-ATPase activity were investigated. The electrophysiological calcium cycling in single isolated cardiomyocytes was measured with the patch-clamp technique. The related ultrastructures were studied with electron microscopy. Results LVAD unloading alleviated LV dilation and improved global cardiac function and regional contractility compared with the MI group. The regional myocardial strain (stretch) was minimized during the unloading period and even attenuated compared with the MI group at 12 weeks. Impaired calcium cycling was evident in the adjacent noninfarcted zone in the MI group, whereas CHP expression was normalized and Ca2+-ATPase activity was preserved in the LVAD unloading group. The electrophysiological calcium cycling was also conserved, and the ultrastructural damage was ameliorated in the unloaded animals. Conclusions Short-term LVAD unloading may conserve calcium cycling and improve heart function during the post-infarct period.

The down regulation of sclerostin in osteocytes mediates bone formation in response to mechanical cues and parathyroid hormone (PTH). To date, the regulation of sclerostin has been attributed exclusively to the transcriptional downregulation of the Sost gene hours after stimulation. Using mouse models and rodent cell lines, we describe the rapid, minutes-scale post-translational degradation of sclerostin protein by the lysosome following mechanical load and PTH. We present a model, integrating both new and established mechanically- and hormonally-activated effectors into the regulated degradation of sclerostin by lysosomes. Using a mouse forelimb mechanical loading model, we find transient inhibition of lysosomal degradation or the upstream mechano-signaling pathway controlling sclerostin abundance impairs subsequent load-induced bone formation by preventing sclerostin degradation. We also link dysfunctional lysosomes to aberrant sclerostin regulation using human Gaucher disease iPSCs. These results reveal how bone anabolic cues post-translationally regulate sclerostin abundance in osteocytes to regulate bone formation.

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