Host responses against heat shock proteins from periodontopathogenic bacteria

口腔微生物的代谢在很大程度上受到环境变化的影响，环境变化也会导致热休克蛋白(HSPs)等应激蛋白的合成。热休克蛋白属于一些高度保守的同源家族。我们将重点放在牙周病和作为牙周病原体的HSPs上，因为这些蛋白是免疫优势抗原。我们的结果表明，牙周病原体的GroEL样蛋白属于HSP60家族，与人类来源的GroEL蛋白具有表位交叉反应。利用与直肠弯曲菌GroEL样蛋白相对应的PCR产物直接测定核苷酸序列，并进行分析。将推导出的氨基酸序列与人HSP60的氨基酸序列进行比对，证实了其全长的相似性。这可能是由于同一家族内的热休克蛋白及其共同表位与宿主分子的高度同源性而导致自身免疫应答。该类GroEL蛋白通过人牙龈成纤维细胞培养上清液中的融合单层细胞刺激IL-6和IL-8的分泌。在22h培养过程中，IL-6和IL-8的含量分别增加了5.4和3.5倍。伴放线放线杆菌GroEL样蛋白在较高浓度下对体外培养的牙周膜上皮细胞和HaCaT上皮细胞均有明显的细胞毒作用。另一方面，我们利用免疫电子显微镜确定了伴放线放线杆菌GroEL样蛋白与特异性抗体的外部定位。这些结果表明，伴生放线菌GroEL-like蛋白和直肠放线菌GroEL-like蛋白对宿主细胞具有生物活性，可能在牙周病中发挥重要作用。

The metabolism of oral microorganisms is largely influenced by the changing environment which also lead to the synthesis of stress proteins such as heat shock proteins (HSPs). HSPs belong to some families of highly conserved homology. We focused on the periodontal disease and HSPs as periodontopathogen because these proteins were immunodominant antigens. Our results indicated that the GroEL-like protein, belonging to HSP60 family, from periodontopathogens possessed epitope cross-reactive with that from human origin. The nucleotide sequence was determined directly using the PCR product corresponding to the Campylobacter rectus GroEL-like protein, and analyzed. By the comparison of the alignment of the deducted amino acid sequence with those of human HSP60, the similarity throughout its length was confirmed. It is possible that high homology of HSPs within the same family and their common epitopes with host molecule lead to autoimmune responses.The native GroEL-like protein from C.rectus and Actinobacillus actinomycetemcomitans possessed biological activities. The C.rectus GroEL-like protein stimulated both interleukin-6 (IL-6) and IL-8 secretion by a confluent monolayer of human gingival fibroblast in their culture supernatant. During the 22h-incubation, the amounts of IL-6 and IL-8 were increased by 5.4- and 3.5-fold, respectively. A.actinomycetemcomitans GroEL-like protein had a significant cytotoxic effect on both periodontal ligament epithelial cells and HaCaT epithelial cells grown in vitro at higher concentrations. On the other hand, we determined the external localization of A.actinomycetemcomitans GroEL-like protein with specific antibodies using immunoelectron microscopy. These results demonstrated that the A.actinomycetemcomitans GroEL-like protein and the C.rectus GroEL-like protein possessed biological activities influencing on the host cells, therefore, these proteins could play important roles in periodontal disease.