Establishment of Diagnostic Methods of Periodontal Disease Basing on the Alteration of Immunocompetent Cells

最近的研究表明，CD45RO+记忆T细胞通过产生各种B细胞激活细胞因子而在B细胞免疫球蛋白的产生中发挥重要作用，并通过促进黏附分子的表达而促进细胞与细胞的接触。本研究采用免疫组织化学双重染色的方法，研究记忆T细胞和活化B细胞在成人牙周炎和牙周炎病变冰冻切片中的分布和频率。我们使用相同的技术通过染色原位检测T细胞与B细胞的相互作用：T细胞产生IL-4，它增强B细胞上CD23的表达，LFA-1在T细胞上，ICAM-1在B细胞上。在牙周手术前完成牙周准备后，共采集了19份牙龈活检标本，从需要拔除正畸治疗的前磨牙部位和第三磨牙部位采集了5份牙周炎活检标本。连续6微米厚的冷冻切片取自每一次活检的中心部分。AVI…双重染色用DIN-生物素免疫过氧化物酶和亲和素-生物素-碱性磷酸酶或碱性磷酸酶抗碱性磷酸酶技术鉴定CD4+CD45RO+记忆性T细胞、IL-4+CD45RO+细胞、LFA-1+CD45RO+细胞、ICAM-1+CD19+B细胞和CD23+或CD25+、CD19+活化的B细胞。CD45RO+记忆细胞在CD4+细胞中所占比例一直较高(65.9%、61.2%、62.5%)，而CD45RA+幼稚细胞所占比例较低(分别为17.1%、14.8%、12.4%)。在牙周炎活检组织中，T细胞是主要的细胞类型，CD4+细胞以CD45RO+为主。在19例牙周组织中，有14例表达IL-4的细胞呈不同强度的簇状和/或单个细胞，但数量明显少于CD45RO+细胞。CD23+和CD25+、CD19+B细胞的百分率有明显的个体差异(CD23为0-100%，CD25为0-36.2%)，尽管临床状态相似。部分CD19+B细胞ICAM-1表达增强。上述结果提示：1)T细胞和B细胞在进入牙周炎病变前可能发生在局部淋巴结的早期激活事件；2)进入牙周炎病变的B细胞表现出不同阶段向浆细胞分化的特征。较少

Recent studies showed that CD45RO+ memory T-cells play important roles in the immunoglobulin production of B-cells by producing various B-cell activating cytokines and cell-cell contact due to enhanced expression of adhesion molecules. This study was designed to investigate frequency and distribution of memory T-cells and activated B-cells in cryostat sections of adult periodontitis and gingivitis lesions by immunohistochemical double staining. We used the same technique to examine T-B interactions in situ by staining: T-cells producing IL-4 which enhances CD23 expression on B-cells, LFA-1 on T-cells, and ICAM-1 on B-cells. Nineteen gingival biopsies were obtained at the time of periodontal surgery following the completion of initial preparation and 5 gingivitis biopsies were obtained from premolar sites requiring extraction for the orthodontic treatment and third molar sites. Serial 6 um-thick cryostat sections were obtained from the central part of each biopsy. Double staining by avi … More din-biotin- immunoperoxidase and avidin-biotin-alkaline phosphatase or alkaline phosphatase anti-alkaline phosphatase techniques were used to identify CD4+CD45RO+ memory T-cells, IL-4+CD45RO+ cells, LFA-1+CD45RO+ cells, ICAM-1+CD19+ B-cells and CD23+ or CD25+,CD19+ activated B-cells. The mean percentage of CD45RO+ memory cells among CD4+ cells was consistently high (65.9% in the crevicular one-third, 61.2% in the middle one-third, 62.5% in the oral one-third), whereas the mean percentage of CD45RA+ naive cells among CD4+ cells was low (17.1% in the crevicular one-third, 14.8% in the middle one-third and 12.4% in the oral one-third). In gingivitis biopsies, T-cells were the major cell type and most of CD4+ cells were CD45RO+. IL-4 producing cells were observed as clusters and/or single cells with various staining intensity in 14 out of 19 periodontal tissues, however, they were much lower in number compared with CD45RO+ cells. On the other hand, the percentage of CD23+ and CD25+, CD19+ B-cells displayed pronounced individual variations (0-100% for CD23, 0-36.2% for CD25) even though the clinical status was similar. Some CD19+ B-cells with enhanced expression of ICAM-1 were identified. The above findings suggest that: 1)early activation events of T-cells and B-cells may take place possibly in the regional lymphnodes before infiltrating into the periodontitis lesion; 2)B-cells infiltrating into the periodontitis lesion displayed features attributable to several different stages of differentiation pathway into plasma cells. Less