Analysis of signal transduction of inflammatory cytokines in bone destruction

在小鼠成骨细胞与骨髓细胞共培养的基础上，研究了脂多糖（LPS）诱导破骨细胞生成的机制。LPS刺激破骨细胞生成和前列腺素E_2（PGE_2）的产生，这两种作用均被环氧合酶-2抑制剂NS 398抑制。LPS刺激成骨细胞NF-κB配体受体激活剂（RANKL）mRNA表达，抑制成骨细胞骨保护素（OPG）mRNA表达。NS 398仅抑制LPS诱导的OPG mRNA表达下调，提示LPS刺激的PGE_2产生在OPG表达下调中起重要作用。事实上，在含有OPG敲除小鼠来源的成骨细胞的共培养物中，NS 398未能抑制LPS诱导的破骨细胞生成。钙/蛋白激酶C（PKC）抑制剂和PD 98059 [丝裂原活化蛋白激酶（MAPK）/细胞外信号调节激酶（ERK）激酶（MEK）抑制剂]抑制成骨细胞中RANKL mRNA的表达，并抑制LPS处理的共培养物中的破骨细胞生成。LPS诱导成骨细胞MEK和ERK磷酸化，这种诱导作用可被钙/PKC抑制剂抑制。此外，PD 98059和NS 398抑制共培养物中白细胞介素1（IL-1）诱导的破骨细胞生成，但只有PD 98059抑制IL-1诱导的成骨细胞RANKL mRNA表达。这些结果表明，LPS和IL-1通过两个平行事件促进破骨细胞生成：通过钙/PKC信号随后MEK/ERK信号增强RANKL表达，以及通过PGE_2介导抑制OPG产生。

The mechanism of osteoclastogenesis induced by lipopolysaccharide (LPS) was studied in cocultures of mouse osteoblasts and bone marrow cells. LPS stimulated osteoclastogenesis and prostaglandin E_2 (PGE_2) production in the cocultures, both of which were inhibited by NS398, a cyclooxygenase-2 inhibitor. LPS stimulated receptor activator of NF-κB ligand (RANKL) mRNA expression, and inhibited osteoprotegerin (OPG) mRNA expression in osteoblasts. NS398 inhibited only LPS-induced down-regulation of OPG mRNA expression, suggesting that LPS-stimulated PGE_2 production is important for the down-regulation of OPG. Indeed, NS398 failed to inhibit LPS-induced osteoclastogenesis in cocultures containing OPG knockout mouse-derived osteoblasts. Calcium/protein kinase C (PKC) inhibitors and PD98059 [mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor] suppressed RANKL mRNA expression in osteoblasts, and inhibited osteoclastogenesis in the cocultures treated with LPS. LPS induced phosphorylation of MEK and ERK in osteoblasts, and this induction was inhibited by calcium/PKC inhibitors. In addition, PD98059 and NS398 inhibited interleukin 1 (IL-1)-induced osteoclastogenesis in the cocultures, but only PD98059 suppressed RANKL mRNA expression induced by IL-1 in osteoblasts. These results suggest that LPS and IL-1 similarly promote osteoclastogenesis through two parallel events : enhancement of RANKL expression through calcium/PKC signals followed by MEK/ERK signals, and suppression of OPG production mediated by PGE_2.