Lipopolysaccharide and high concentrations of glucose enhances zoledronate-induced increase in rankl/opg ratio by upregulating pge₂ production in osteoblasts

Preexisting diseases, such as diabetes and chronic inflammation in periodontal tissue, are risk factors associated with bisphosphonate-related osteonecrosis of the jaw. Osteoblasts produce prostaglandin (PG)E₂ via cyclooxygenases (COX), and the autocrine action of PGE₂ impacts the function of osteoblasts, including receptor activator of NF-kappa B li-gand (RANKL) and osteoprotegerin (OPG) production. This study assessed the effects of the stimulation of zoledronate in the presence of lipopolysaccharide (LPS) and high concentrations of glucose on the expression of COX-2, RANKL, and OPG, in addition to PGE₂ production in osteoblasts. MG-63 cells were cultured in medium containing 1 µg/ml LPS, 25 mM glucose (high glucose), and/or zoledronate (1×10^-8, 1×10^-7, 1×10^-6, 5×10^-6, or 1×10^-5 M). The mRNA expression of COX-2, RANKL, and OPG genes was determined by real-time polymerase chain reaction. The concentrations of RANKL and OPG protein and PGE₂ in the culture supernatant were examined by enzyme-linked immunosorbent assay. Zoledronate at a concentration of 5×10^-6 M overwhelmingly increased COX-2 mRNA expression. The expression levels of RANKL and OPG as well as PGE₂ production was significantly increased in cells stimulated with 5×10^-6 M zoledronate in the presence of LPS and high glucose than in the unstimulated cells (control). NS398, a specific inhibitor of COX-2, blocked the stimulatory effects of zoledronate (in the presence of LPS and high glucose) on PGE₂ production and the protein expression levels of RANKL and OPG. The ratio of RANKL/OPG was also increased following zolendronate stimulation. In addition, a significant difference was observed not in the stimulation with zoledronate alone, but by the stimulation of zoledronate in the presence of LPS and high glucose as compared that in controls. These results suggest that LPS and high concentrations of glucose enhances zoledronate-induced increase in RANKL/OPG ratio via the autocrine action of NS398-blocked PGE₂ in osteoblasts.