Lipopolysaccharide of Aggregatibacter actinomycetemcomitans up-regulates inflammatory cytokines, prostaglandin E2 synthesis and osteoclast formation in interleukin-1 receptor antagonist-deficient mice.

Background and Objective The interleukin ( IL)-1 receptor antagonist ( Ra) binds to IL-1 receptors and inhibits IL-1 activity. However, it is unclear whether the IL-1 Ra plays a protective role in periodontal disease. The purpose of this study was to compare IL-1 Ra knockout ( KO) and wild-type ( WT) mice in regard to proinflammatory cytokine production, osteoclast formation and bone resorption in response to periodontal bacterial lipopolysaccharide ( LPS). Material and Methods Peritoneal macrophages ( Mφs) were obtained from 13-wk-old IL-1 Ra KO and WT mice. Peritoneal Mφs were cultured with or without 10 μg/mL of Aggregatibacter actinomycetemcomitans LPS for 24 h. The levels of IL-1alpha ( IL-1α), IL-1beta ( IL-1β), tumor necrosis factor-α ( TNF-α) and IL-6 were measured in periotoneal Mφs supernatant fluid ( PM-SF) using an ELISA. Bone marrow cells were obtained from the mice and stimulated with PM-SF for 9 d, then stained with TRAP. The frequency of TRAP-positive multinucleated giant cell formation was calculated based on a fusion index. PM-SF-stimulated calvarial bone resorption was analyzed using micro-computed tomography, and calvarial histological analysis was performed using hematoxylin and eosin and TRAP staining. The expression of cyclooxygenase-2 ( Cox2), prostanoid receptor EP4 ( Ep4) and Rank m RNAs in bone marrow cells were measured using real-time quantitative PCR, while prostaglandin E2 ( PGE₂) production was determined by ELISA. Results The levels of IL-1α, IL-1β, TNF-α and IL-6 in IL-1 Ra KO mice PM-SF stimulated with A. actinomycetemcomitans LPS were significantly increased by approximately 4- ( p < 0.05), 5- ( p < 0.05), 1.3- ( p < 0.05) and 6- ( p < 0.05) fold, respectively, compared with the levels in WT mice. Moreover, osteoclast formation, expression of Rank, Ep4 and Cox2 m RNAs and production of PGE₂ were significantly increased by approximately 2- ( p < 0.05), 1.6- ( p < 0.05), 2.5- ( p < 0.05), 1.6- ( p < 0.05) and 1.9- ( p < 0.05) fold, respectively, in IL-1 Ra KO mice stimulated with A. actinomycetemcomitans LPS compared with WT mice. Conclusion IL-1 Ra regulates IL-1 activity and appears to reduce the levels of other inflammatory cytokines, including TNF-α and IL-6, while it also reduces expression of the EP4 receptor related to prostanoid sensitivity and osteoclast formation. These results suggest that IL-1 Ra is an important molecule for inhibition of inflammatory periodontal bone resorption. [ABSTRACT FROM AUTHOR]