Abstract
Osteocytes comprising over 90% of the bone cell population are highly susceptible to the adverse effects of glucocorticoids (GC) administration. Here we observed that Dexamethasone (Dex) induces a robust cytoskeleton rearrangement and decreases Cx43 protein expression in osteocyte-like MLO-Y4 cells. Using a Dmp1Cre-mT/mG osteocyte ex vivo culture system, we found significant shortening of dendritic processes in primary osteocytes following Dex administration. Loss of dendritic processes is a consequence of reduced Cx43 connectivity upon Dex induced autophagy in both RFP-GFP-LC3B transfected MLO-Y4 cells and primary calvarial osteocytes from LC3GFP transgenic mice. Upon the induction of autophagy by Dex, Cx43 was internalized into autophagosome/autolysosomes and degraded by autophagy. The degradation was attenuated following lysosomal inhibition using chloroquine (CLQ) and suppression of autophagy by Atg5 silencing. Inhibition Akt-mTORC1 signaling by Dex induces autophagy subsequently resulting in Cx43 degradation. Activation of Akt phosphorylation by IGF-1 attenuated Dex induced autophagy and degradation of Cx43. Together, we demonstrated that GC impair osteocyte cell-cell connectivity via autophagy mediated degradation of Cx43 through inhibition of the Akt-mTORC1 signaling. This may account for the deleterious effect of GC-induced bone loss.
Highlights
Being entombed within the mineralized matrix of bone, osteocytes form a complicated yet sophisticated network of intercellular connections via their dendritic processes to communicate and regulate neighboring osteocytes and cells on the bone surface
The osteocyte dendritic network formed by dendritic connections linked by Connexin 43 (Cx43) between neighboring osteocytes plays a critical role in bone homeostasis [12]
MLO-Y4 cells treated with Dex demonstrated a dosedependent shortening of dendritic processes, which were accompanied by a drastic rearrangement of actin cytoskeleton characterized by increased stress fiber formation and a significant decrease in Cx43 immunoreactivity from 10-6M Dex treatment (Figure 1A, 1B and 1C)
Summary
Being entombed within the mineralized matrix of bone, osteocytes form a complicated yet sophisticated network of intercellular connections via their dendritic processes to communicate and regulate neighboring osteocytes and cells on the bone surface. Connexin 43 (Cx43), a major hemichannel protein plays an important role in maintaining dendritic connection between neighboring osteocytes [1, 2]. Chondroosteogenic lineage Cx43 deficient mice exhibit increased bone resorption and TRAP positive osteoclasts [6, 7]. Together, these data suggested that Cx43 in osteocytes plays a critical role regulating both bone resorption and formation to maintain bone hemostasis [1, 8]
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