Bone pain is one of the most prevalent and devastating complications of cancer in bone. The pathophysiology of cancer-associated bone pain (CABP) is poorly understood but likely involves complex interactions among the cancer cells, peripheral sensory nerves and bone cells. Recent studies reported that the calcitonin gene-related peptide-positive (CGRP+) sensory neurons densely innervate mineralised bone, in which numerous osteocytes are present, leading us to hypothesise that osteocytes interact with these CGRP+ sensory neurons to evoke CABP. We tested this hypothesis using an animal model in which inoculation of the JJN3 human multiple myeloma (MM) cells into tibiae induced progressive CABP. We found that JJN3 MM-colonised bone was acidic and that blockade of the acidification by the proton pump inhibitor bafilomycin A1 significantly reduced the CABP. Immunohistochemical examination demonstrated that osteocytes localised in the close proximity of CGRP+ primary afferent sensory neurons in mineralised bone. Co-culture of MLO-A5 osteocytic cells and F11 sensory neuronal cells showed that MLO-A5 cells transferred the permeable living dye calcein to F11 cells by extending dendritic processes to contact the neurites of F11 cells. The general gap junction inhibitor 18β-GA and the selective connexin43 (Cx43) blocker GAP27 and silencing Cx43 in MLO-A5 cells by shRNA all decreased the dye transfer, suggesting that the Cx43 gap junction mediates the osteocyte-sensory neuron communication. Determination of neuronal excitation by Ca2+ influx imaging assay showed that the acidic medium excited F11 sensory neuronal cells. Importantly, acid-induced excitation of F11 cells was enhanced in the presence of MLO-A5 osteocytic cells and GAP27 and silencing Cx43 abolished acid-induced F11 excitation in the co-cultures. In conclusion, our results suggest that osteocytes contribute to the pathophysiology of CABP via Cx43-mediated communications with sensory neurons innervating bone. These communications may be a novel therapeutic target in the management of CABP.
Disclosure: The authors declared no competing interests. This work was supported by the institutional start-up fund.