S100A9 Induces RANKL Expression in Osteocytes Utilizing IL-6/sIL-6R Combination and gp130 Signaling Pathways

Calprotectin, a heterodimer of S100A8 and S100A9, is predominantly expressed in neutrophils, monocytes, and chondrocytes. Clinical studies demonstrate that elevated circulating calprotectin levels correlate with diseases followed by inflammatory bone loss, including rheumatoid arthritis, osteoarthritis, and periodontitis. We previously reported that S100A9 directly increased RANKL and IL-6 expression in an osteocyte-like cell line. While RANKL is well-established as an essential factor for the formation and activation of osteoclasts, the role of S100A9-induced IL-6 in bone resorption remains unclear. On the other hand, osteocytes require sIL-6R to activate the gp130 signaling pathway. Therefore, we hypothesized that S100A9-induced IL-6 cooperates with soluble IL-6 receptor (sIL-6R) to increase RANKL through gp130-mediated signaling. Here, we show that S100A9 increases the bone resorption area and TRAP-positive cell number in the murine calvaria. In addition, Rankl and Il6 mRNA levels in osteocytes were upregulated by S100A9 treatment. In vitro results demonstrated that S100A9 increased not only Rankl and Il6 expression in osteocytes, but also sIL-6R secretion in macrophages. Since osteocytes strongly express gp130, the IL-6 complexed with sIL-6R enhanced gp130 downstream signaling, including JAK1 and STAT3 activation in osteocytes. Blockade of either Il6 or gp130 suppressed S100A-induced effects. Notably, gp130 antagonist treatment inhibited S100A9-induced bone resorption in vivo. Collectively, these findings suggest that S100A9 promotes RANKL-mediated osteoclastogenesis through IL-6/sIL-6/gp130 signaling in osteocytes.