High glucose alters the secretome of mechanically stimulated osteocyte‐like cells affecting osteoclast precursor recruitment and differentiation

Abstract

Diabetes mellitus (DM) induces bone deterioration, while mechanical stimulation promotes osteocyte-driven bone formation. We aimed to evaluate the interaction of acute exposure (24 h) to high glucose (HG) with both the pro-survival effect conferred to osteocytic MLO-Y4 cells and osteoblastic MC3T3-E1 cells by mechanical stimulation and the interaction of these cells with osteoclast precursor RAW264.7 cells. We found that 24 h of HG (25 mM) pre-exposure prevented both cell survival and ERK and β-catenin nuclear translocation upon mechanical stimulation by fluid flow (FF) (10 min) in both MLO-Y4 and MC3T3-E1 cells. However, migration of RAW 264.7 cells was inhibited by MLO-Y4 cell-conditioned medium (CM), but not by MC3T3-E1 cell-CM, with HG or FF. This inhibitory effect was associated with consistent changes in VEGF, RANTES, MIP-1α, MIP-1β MCP-1, and GM-CSF in MLO-Y4 cell-CM. RAW264.7 proliferation was inhibited by MLO-Y4 CM under static or HG conditions, but it increased by FF-CM with or without HG. In addition, both FF and HG abrogated the capacity of RAW 264.7 cells to differentiate into osteoclasts, but in a different manner. Thus, HG-CM in static condition allowed formation of osteoclast-like cells, which were unable to resorb hydroxyapatite. In contrast, FF-CM prevented osteoclastogenesis even in HG condition. Moreover, HG did not affect basal RANKL or IL-6 secretion or their inhibition induced by FF in MLO-Y4 cells. In conclusion, this in vitro study demonstrates that HG exerts disparate effects on osteocyte mechanotransduction, and provides a novel mechanism by which DM disturbs skeletal metabolism through altered osteocyte-osteoclast communication.

Spatiotemporally Controlled Release of Rho‐Inhibiting C3 Toxin from a Protein–DNA Hybrid Hydrogel for Targeted Inhibition of Osteoclast Formation and Activity

“…To analyze C3‐mediated toxicity in osteoclasts, hydrogel alone or hydrogel loaded with C3‐derived recombinant fusion toxin C2IN‐C3lim‐G205C were incubated with RAW 264.7 cells in osteoclastogenic medium. The RAW 264.7 cell is a well‐characterized preosteoclastic cell line, which is widely used as a model for osteoclast formation and activity . Preliminary experiments showed that these cells were, like J774A.1 cells, susceptible to the cytotoxic effect of the recombinant fusion toxin (Figures S1 and S2, Supporting Information).…”

Pulsed electromagnetic fields regulate osteocyte apoptosis, RANKL/OPG expression, and its control of osteoclastogenesis depending on the presence of primary cilia

“…You et al () have found that fluid shear stress stimulation for 2 hr had no changes in sRANKL of MLO‐Y4 cells, but decreased osteoclast formation. The results from Maycas et al () showed that high glucose induced no significant change of RANKL expression from MLO‐Y4 cells, but induced decease of osteoclast activity. Thus, the findings from both our current study and other groups indicated that RANKL may not be the only molecule that secreted by osteocytes to control osteoclast formation and activity, and some other molecules (e.g., M‐CSF and VEGF) may also contribute to the regulation of osteoclast functions.…”

Metabolomics reveals citric acid secretion in mechanically–stimulated osteocytes is inhibited by high glucose

“…Previous studies of our group have shown that HG affects osteocyte mechanotransduction, especially osteocyte-osteoclast communication, suggesting new pathways to link the mechanisms involved in the deleterious effects of DM in the skeleton 14 . We emphasized that the analysis of the metabolic secretome may be critical to obtain new insights into this process and the role of HG levels in these effects.…”

“…No significant changes in cell viability were observed between the different experimental conditions (with or without HG and/or FF). The details of the experiment can be found in a previously published article 14 .…”

“…It has been demonstrated that a diabetic background affects osteocyte mechanotransduction by impairing several mechanisms induced through mechanical forces in both in vivo and in vitro models 11–13 . Recently, our group has shown that HG alters the secretome of mechanically stimulated osteocyte cells, affecting the manner these cells communicate with osteoclasts 14 .…”