E11/Podoplanin Protein Stabilization Through Inhibition of the Proteasome Promotes Osteocyte Differentiation in Murine in Vitro Models

摘要: The transmembrane glycoprotein E11 is considered critical in early osteoblast-osteocyte transitions (osteocytogenesis), however its function and regulatory mechanisms are still unknown. Using the late osteoblast MLO-A5 cell line we reveal increased protein/mRNA expression (P < 0.001) concomitant with extensive osteocyte dendrite formation matrix mineralization (P < 0.001). Transfection significantly mRNA levels (P < 0.001), but immunoblotting failed to detect any correlative increases protein levels, suggestive of post-translational degradation. We found that exogenous treatment osteocytic IDG-SW3 cells 10 μM ALLN (calpain proteasome inhibitor) stabilized induced a profound increase Treatment other calpain inhibitors promote similar osteocytogenic changes, suggesting these effects rely upon inhibitor actions. Accordingly proteasome-selective (MG132/lactacystin/ Bortezomib/Withaferin-A) produced dose-dependent primary cells. This proteasomal targeting was confirmed by immunoprecipitation ubiquitinylated proteins, which included E11, addition MG132/Bortezomib. Activation RhoA, small GTPase, be peak downstream signaling also observed formation. Our data indicate mechanism reliant blockade proteasome-mediated destabilization contributes osteocytogenesis this may involve RhoA. work adds our mechanistic understanding factors regulating bone homeostasis, lead future therapeutic approaches.