Analysis of antigen receptor genes of T cells specific for the periodontal disease

我们以前的RT-PCR-SSCP分析表明，T细胞在牙周炎病变中的克隆性很高，与类风湿关节炎等自身免疫性疾病的炎症病变中的T细胞克隆性相似。考虑到牙周袋中复杂的微生物区系，推测T细胞识别与宿主成分同源的细菌抗原。由于热休克蛋白60s具有显著的免疫原性，尽管它们在进化上高度保守，但已有报道在各种炎症条件下对Hsp60的T细胞和抗体反应。为了阐明hsp60S在牙周炎T细胞反应中的作用，我们检测了牙周炎患者和对照组外周血单个核细胞(PBMC)在重组人hsp60O和牙龈卟啉单胞菌(P.gigivalis)GroEL刺激下的增殖反应以及T细胞的细胞因子谱和克隆性。此外，NU…在T细胞受体β链的CDR3区域中，比较了HSP60反应的PBMC和牙周炎病变浸润性T细胞中更多的Clebit序列。在牙周炎患者中，PBMC对人HSP60的增殖反应明显高于对照组，而对牙龈假单胞菌的增殖反应不明显。对CDR3区核苷酸序列的分析清楚地表明，Hsp60反应性T细胞克隆和牙周炎病变浸润性T细胞表达相同的受体，提示Hsp60反应性T细胞在牙周炎病变中积聚。细胞因子分析表明，在牙周炎患者中，Hsp60反应的PBMC产生显著水平的干扰素-γ，而牙龈假单胞菌并不诱导向类型1或类型2倾斜。在对照组中，Hsp60或P.gigivalis GroEL均未诱导出明显的干扰素-γ或IL-4的表达。这些结果提示，在牙周炎患者中，感染牙周病原菌可能诱导具有1型细胞因子的自身HSP60反应性T细胞的激活，从而刺激巨噬细胞产生促炎细胞因子，从而导致组织破坏。较少

Our previous studies using RT-PCR-SSCP analysis demonstrated the high clonality of T cells infiltrating into periodontitis lesion similar to that in inflammatory lesion of autoimmune diseases such as rheumatoid arthritis. Considering that the complex microbial flora in the periodontal pocket, it was speculated that T cells are recognizing antigens of bacteria which are homologous to host components. Because heat shock protein 6Os are remarkably immunogenic, despite their high degree of evolutionary conservation, both T cell and antibody responses to hsp60 have been reported in a variety of inflammatory conditions. In order to clarify the role of hsp60s on the T cell response in periodontitis, we examined the proliferative response of peripheral blood mononuclear cells (PBMC) as well as cytokine profile and clonality of T cells upon stimulation with recombinant human hsp6O and Porphyromonas gingivalis (P.gingivalis) GroEL from periodontitis patients and control subjects. Furthermore, nu … More cleotide sequences within the CDR3 region of the T cell receptor β-chain was compared between hsp60-reactive PBMC and periodontitis lesion-infiltrating T cells. In periodontitis patients significantly higher proliferative response of PBMC to human hsp60 but not to P.gingivalis GroEL than in control subjects. Analysis of the nucleotide sequences within the CDR3 region clearly demonstrated that hsp60-reactive T cell clones and periodontitis lesion-infiltrating T cells expressed the same receptors suggesting that hsp60-reactive T cells accumulated in periodontitis lesion. The analysis of cytokine profile demonstrated that hsp60-reactive PBMC produced significant level of IFN-γ in periodontitis patients, whereas P.gingivalis GroEL did not induce any skewing towards type1 or type2. In control subjects no significant expression of IFN-γ or IL-4 was induced upon stimulation with either hsp60 or P.gingivalis GroEL.These results suggest that in periodontitis patients infection with periodontopathic bacteria may induce the activation of self hsp60-reactive T cells with type1 cytokine profile resulting the stimulation of macrophage to produce proinflammatory cytokines leading to the tissue destruction. Less