Kang Philip, Korostoff Jonathan, Volgina Alla, Grzesik Wojciech, DiRienzo Joseph M
Departments of Periodontics1, Microbiology2 and Anatomy & Cell Biology3, School of Dental Medicine, University of Pennsylvania, 240 South 40th Street, Philadelphia, PA, USA.
J Med Microbiol. 2005 Aug;54(Pt 8):785-794. doi: 10.1099/jmm.0.46077-0.
The periodontal pathogen Actinobacillus actinomycetemcomitans expresses a cytolethal distending toxin (CDT) that typically arrests the growth of eukaryotic cells at either the G0/G1 or G2/M phase of the cell cycle. It was previously found that CDT failed to arrest the growth of human periodontal ligament fibroblasts (HPLFs) when grown in pure culture. In contrast, proliferation of an oral epithelial cell line was rapidly inhibited by the toxin. In this study, the feasibility of using mixed-cell cultures and cell-specific markers to evaluate the response of oral cells, when in heterogeneous populations, to CDT was established. Proliferation of epithelial cells was rapidly inhibited and the cells were selectively eliminated in co-culture with HPLFs or cementoblasts by 24-48 h post-intoxication. Epithelial cells and HPLFs were detected and counted in co-cultures following cell-specific immunolabelling with antibodies against simian virus 40 large T antigen and the Ab-1 surface antigen, respectively. These results demonstrated that the activities of potential virulence factors, such as CDT, from periodontal pathogens can be successfully examined in mixed-cell cultures. This approach is especially relevant to infectious diseases that affect tissues with a diverse cellular composition, such as the periodontium.
牙周病原体伴放线放线杆菌表达一种细胞致死性膨胀毒素（CDT），该毒素通常会使真核细胞的生长停滞在细胞周期的G0/G1期或G2/M期。此前发现，当在纯培养物中生长时，CDT无法阻止人牙周膜成纤维细胞（HPLF）的生长。相反，毒素可迅速抑制口腔上皮细胞系的增殖。在本研究中，确定了使用混合细胞培养物和细胞特异性标志物来评估口腔细胞在异质群体中对CDT反应的可行性。中毒后24至48小时，上皮细胞的增殖被迅速抑制，并且在与HPLF或成牙骨质细胞共培养时，细胞被选择性清除。在用抗猿猴病毒40大T抗原和Ab-1表面抗原的抗体进行细胞特异性免疫标记后，在共培养物中检测并计数上皮细胞和HPLF。这些结果表明，牙周病原体潜在毒力因子（如CDT）的活性可以在混合细胞培养物中成功检测。这种方法对于影响具有多种细胞组成的组织（如牙周组织）的传染病尤为重要。
