Objetivo

Bone tissue can adapt to environmental stimuli by altering its morphology and metabolism. Different bone cells communicate with each other through communicating junctions (CJs). Connexin 43 (Cx43) is the most abundant CJ protein with key functions in signal transduction and in response to hormonal and mechanical stimuli. Another mechanosensor element of osteocytes is the primary cilium, formed by microtubules and which develops in the cell cycle’s G0 phase.

Our study aims to determine Cx43 and primary cilium involvement in osteocytic activity, to analyze the possible interaction between these two mechanosensors and to assess the role they play in the detection and response of osteocytes to mechanical stimuli and stimulation of the parathormone type 1 receptor (PTH1R) by its ligand, the parathormone-related protein (PTHrP) (1-36).

Material and methods

The control MLO-Y4 (Cx43 +/+) osteocyte cell line was compared to Cx43-deficient MLO-Y4 (Cx43 -/-). The expression analysis of intraflagellar transport protein 88 (IFT88), Cx43 and phosphorylation of the extracellular signal regulatory kinase (P-ERK) was determined by Western blot. To characterize the possible colocalization between the primary cilium and Cx43, an immunofluorescence was carried out. To simulate mechanical stimulation in vitro, cells were subjected to mechanical stress of 10 dynes/cm2 by fluid flow for 10 minutes.

Results

The results obtained show that the number of cells with primary cilium does not vary due to the expression of Cx43 (p = 0.089). In cells with Cx43 presence, mechanical stimulation by fluid flow and PTHrP increase the phosphorylation of extracellular signal-regulated kinases (ERK) compared to unstimulated cells (p = 0.049 and p = 0.011, respectively).

Conclusions

The primary cilium and Cx43 act as mechanosensing elements for osteocytes. Deficiency in Cx43 does not influence ciliogenesis or activation by mechanical stimulation of pro-survival signaling pathways in osteocytes.

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