Development of a diagnostic tool for periodontal disease by utilizing the monoclonal antibody directed the specific protease from periodontopathogensis bacteria

利用针对牙周病原菌特异性蛋白酶的单克隆抗体开发牙周病诊断工具

• 批准号: 06557113
• 负责人: NAKAMURA Ryo
• 金额: $ 4.35万
• 依托单位: The University of Tokushima
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06557113/
• 关键词: Clinical diagnosis; Trypsin-like enzyme; Porphyromonas gingivalis; Monoclonal antibody; Periodontal disease; Gingival crevicular fluid; P.gingivalis

A monoclonal antibody (mAb-PC) was produced against a trypsin-like enzyme (Pase-C) from Porphyromonas gingivalis. By the neutrolizing experiment, it was revealed that the mAb-PC bound Pase-C apart from the acive site of the enzyme and ihibited its enzyme activity. Other trypsin-like enzymes (Pase-B and Pase-S) from P.gingivalis did not react with this antibody. By ELISA or dot blotting analysis, mAb-PC recognized all strains of P.gingivalis tested, but did not recognized other putative periodontopathogenic organisms including Bacteroides forsythus and Treponema denticola.Fifty samples of gingival crevicular fluid (GCF) were collected from individuals with adult periodontitis, and the levels of Pase-C and P.gingivalis antigens in GCF were measured by a quantitative ELISA,using mAb-PC and anti-P.gingivalis polyclonal antibody, respectively. By the stastistic analysis, it was found that the Pase-C level was not correlated with the P.gingivalis level. However, when "BOP negative group" which bleeding was not found on probing, was subjected to the analysis, these levels were correlated singnificantly. The Pase-C level of "BOP positive group" was higher than that of "BOP negative group" , whereas on the P.gingivalis level, there was not any difference between these groups. Since the bleeding tendency is considered to reflect the inflammatory activity in the periodontal site to some extent, it was suggested that the detection of Pase-C in GCF sample had the potential to be used for the diagnosis and the prediction of prognosis in adult periodontitis.

制备了抗牙龈卟啉单胞菌胰蛋白酶样酶（Pase-C）的单克隆抗体（mAb-PC）。酶联免疫吸附试验表明，单克隆抗体-PC与Pase-C的结合位点不在酶的活性位点，从而抑制了酶的活性。来自牙龈卟啉单胞菌的其他胰蛋白酶样酶（Pase-B和Pase-S）不与该抗体反应。用ELISA或斑点印迹法检测了50例成人牙周炎患者的龈沟液（GCF）中Pase-C抗原和牙龈卟啉单胞菌抗原的含量，并与10例牙周炎患者的GCF进行比较。分别用单克隆抗体PC和抗牙龈卟啉单胞菌多克隆抗体进行检测。经统计学分析发现，牙龈卟啉单胞菌与Pase-C水平无相关性。然而，当对探查时未发现出血的“BOP阴性组”进行分析时，这些水平显著相关。“BOP阳性组”的Pase-C水平高于“BOP阴性组”，而牙龈卟啉单胞菌水平在各组间无差异。由于出血倾向在一定程度上反映了牙周组织中的炎症活动，因此认为GCF中Pase-C的检测有可能用于成人牙周炎的诊断和预后预测。

项目成果

D.Hinode,K.Masuda,M.Yoshioka,H.Hayashi,R.Nakamura: "Purification of an extracllular soluble protease possesing BApNA hydrolytic activity from culture supernatants of Porphyromonas gingivalis" Journal of Dental Health. 44. 308-314 (1994)

D.Hinode、K.Masuda、M.Yoshioka、H.Hayashi、R.Nakamura：“从牙龈卟啉单胞菌培养物上清液中纯化具有 BApNA 水解活性的胞外可溶性蛋白酶”《牙科健康杂志》。

M.Yoshioka: "Possible relationship between the cell aggregating activity and the arginine metabolizing enzyme from Porphyromonas gingivalis" J.Dent.Health. 47. 83-88 (1997)

M.Yoshioka：“细胞聚集活性与牙龈卟啉单胞菌精氨酸代谢酶之间的可能关系”J.Dent.Health。

Hinode, D., Masuda, K., Yoshioka, M., Hayashi, H.and Nakamura, R.: "Purification of an extracellular soluble protease possessing BApNA hydrolytic activity from culture supernatants of Porphyromonas gingivalis" J.Dent.Health. 44. 308-314 (1994)

Hinode, D.、Masuda, K.、Yoshioka, M.、Hayashi, H. 和 Nakamura, R.：“从牙龈卟啉单胞菌培养物上清液中纯化具有 BapNA 水解活性的细胞外可溶性蛋白酶”J.Dent.Health。

Yoshioka, M., Otsuka, C., Tamatani, K., Masuda, K., Miki, O., Hayashi, H.and Nakamura, R.: "Possible relationship between the cell aggregating activity and the arginine metabolizing enzyme from Porphyromonas gingivalis" J.Dent.Health. 47. 83-88 (1997)

Yoshioka, M.、Otsuka, C.、Tamatani, K.、Masuda, K.、Miki, O.、Hayashi, H. 和 Nakamura, R.：“细胞聚集活性与卟啉单胞菌精氨酸代谢酶之间的可能关系

M.Yoshioka: "Possible relationship between the ecll aggregating activity and the arginine metubolizing enzymes from Porphyromonas gingivalis" J.Dental Health. 47. 83-88 (1997)

M.Yoshioka：“牙龈卟啉单胞菌的 ecl 聚集活性与精氨酸代谢酶之间的可能关系”J.Dental Health。