Anti-proliferative capsular-like polysaccharide antigen from Actinobacillus actinomycetemcomitans induces apoptotic cell death in mouse osteoblastic MC3T3-E1 cells

Actinobacillus actinomycetemcomitans (A. actinomycetemcomitans) has been im plicated in the etiology of localized juvenile periodontitis (LJP), and pro duces a multiplicity of tissue-damaging products. Among those products, the capsular-like polysaccharide antigen (CPA) from A. actinomycetemcomitans i s a potent mediator of bone resorption. In fact, this CPA (serotype b) is k nown to promote osteoclast-like cell formation via interleukin (IL)-1 alpha production in mouse marrow cultures. Although osteoblasts complete bone fo rmation, there are few reports focusing on the effect of CPA in bone-formin g activity of osteoblasts in inflammatory disease sites. We hypothesized th at CPA plays a mediating role in osteoblastic cells. Therefore, the purpose of this study was to examine the effect of CPA from A. actinomycetemcomita ns ns on the mouse osteoblastic cell line MC3T3-E1 and human osteosarcoma S aOS-2 cells. A. actinomycetemcomitans serotype c resulted in a potent dose- dependent inhibition of cell proliferation of both cell lines. Characteriza tion of the antiproliferative activity in the CPA demonstrated that it was not cytotoxic for MC3T3-E1. A 20-hour incubation with CPA-c resulted in a s ignificant increase in apoptotic cell death in the cells, as evaluated by b oth cellular DNA fragmentation ELISA and FAGS analysis. In contrast to the results obtained with a cytokine mixture (tumor necrosis factor-alpha, IL-1 beta, and interferon-gamma), no inducible nitric oxide (NO) synthase gene expression or NO release could be detected in MC3T3-E1 after incubation wit h CPA-c. Further, both CPA-b and -c caused potent induction of ayoptosis-re lated modifiers, e.g., Fas mRNA, whereas bcl-2 mRNA levels were unchanged. Therefore, this study has shown that CPA from A. actinomycetemcomitans cont ains a potent antiproliferative polysaccharide whose activity is associated with apoptotic cell death in MC3T3-E1, and that CPA per se is an inducer o f apoptosis mediated by the Fas system but not by NO.