Pathirana Rishi D, O'Brien-Simpson Neil M, Visvanathan Kumar, Hamilton John A, Reynolds Eric C
Cooperative Research Centre for Oral Health Science, School of Dental Science, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, 720 Swanston Street, Victoria 3010, Australia.
Cooperative Research Centre for Chronic Inflammatory Diseases, Department of Medicine, The University of Melbourne, Melbourne, Victoria, Australia.
Microbiology (Reading). 2008 Oct;154(Pt 10):2904-2911. doi: 10.1099/mic.0.2008/019943-0.
Porphyromonas gingivalis strains W50 and ATCC 33277 were shown to bind to cultured human fibroblast (MRC-5) cells using flow cytometry. As the concentration of P. gingivalis strain W50 cells was increased relative to the concentration of MRC-5 cells, the number of W50 cells bound per MRC-5 cell increased, as did the percentage of MRC-5 cells with bacteria bound. However, this relationship was only seen for P. gingivalis strain ATCC 33277 at low cell concentrations: at high bacterial cell concentrations strain ATCC 33277 auto-aggregated and binding to the MRC-5 cells decreased. Strain W50 was therefore chosen to study the role of the surface proteinase-adhesin complexes (RgpA-Kgp complexes) in binding to MRC-5 cells. P. gingivalis W50 cells treated with an inhibitor of the RgpA-Kgp complexes exhibited reduced binding to MRC-5 cells. The purified active and proteinase-inactive RgpA-Kgp complexes competitively inhibited binding of W50 to MRC-5 cells, and isogenic mutants of W50 lacking RgpA/B and Kgp displayed reduced binding. P. gingivalis W50 mutant cells lacking Kgp exhibited the lowest binding to MRC-5 cells, suggesting an important role for this proteinase and its associated adhesins in binding to fibroblasts.
使用流式细胞术显示牙龈卟啉单胞菌菌株W50和ATCC 33277可与培养的人成纤维细胞（MRC-5）结合。随着牙龈卟啉单胞菌菌株W50细胞浓度相对于MRC-5细胞浓度的增加，每个MRC-5细胞结合的W50细胞数量增加，结合细菌的MRC-5细胞百分比也增加。然而，这种关系仅在低细胞浓度下的牙龈卟啉单胞菌菌株ATCC 33277中观察到：在高细菌细胞浓度下，ATCC 33277菌株会自动聚集，与MRC-5细胞的结合减少。因此选择菌株W50来研究表面蛋白酶-粘附素复合物（RgpA-Kgp复合物）在与MRC-5细胞结合中的作用。用RgpA-Kgp复合物抑制剂处理的牙龈卟啉单胞菌W50细胞与MRC-5细胞的结合减少。纯化的活性和蛋白酶失活的RgpA-Kgp复合物竞争性抑制W50与MRC-5细胞的结合，缺乏RgpA/B和Kgp的W50同基因突变体显示结合减少。缺乏Kgp的牙龈卟啉单胞菌W50突变细胞与MRC-5细胞的结合最低，表明该蛋白酶及其相关粘附素在与成纤维细胞结合中起重要作用。
