Establishment of new system to prevent oral infectious diseases relating to oral biofilm

变形链球菌表面蛋白抗原（Pac）中富含丙氨酸的重复区（a区）作为龋齿疫苗的抗原成分受到广泛关注。a区Pac（残基361-386）肽具有与多种HLA-DR分子的多重结合基序（L—V-K-A）和被S. mutans抑制抗体识别的b细胞核心表位（Y—L—Y）。在本研究中，我们研究了Pac（361-386）肽是否是诱导人体内抑制抗体的有效抗原，以及它与唾液中抗体诱导的关系。采集151名健康受试者（年龄：36.6±12.6）的刺激唾液样本。采用人外周血单个核细胞（PBMC）移植NOD-scid小鼠，检测Pac（361-386）肽的免疫原性。采用酶联免疫吸附法检测人唾液和移植小鼠血清中Pac（361-386）肽的Hu- IgA和IgG抗体滴度。人唾液中抗pac（361-386）肽IgA抗体滴度(a)呈年龄依赖性显著降低。高抗体组（3<a） MS和MS / tS比低于无抗体组（0.1>a）和中等抗体组（3≧>a）。DRB1^*1501和DRB1^*0406与抗体的高诱导率显著相关，也倾向于减少乳酸杆菌和变形链球菌。此外，在表达各种DRB1基因型的NOD-scid小鼠中证实了肽的免疫原性。肽免疫诱导的单克隆抗体（SH2、KH5）可抑制变形链球菌在大鼠牙表面的定植。因此，Pac（361-386）肽具有较强的免疫原性，可在人口腔内诱导产生抑制抗体，其诱导受衰老、HLA-DRB1基因型和细胞因子的调节。这些结果可能有助于利用牙药delivary系统（3DS）和人单克隆抗体去除口腔生物膜，预防口腔疾病。

Alanine-rich repeating region (A-region) in surface protein antigen (Pac) of Streptococcus mutans has been given much attention as an antigenic component for dental caries vaccines. The Pac (residue 361-386) peptide in A-region possesses a multiple binding motif (L--V-K-A) to various HLA-DR molecules and the B-cell core epitope (Y---L--Y) recognizing by the inhibiting antibody to S. mutans. In this study, we investigated whether Pac (361-386) peptide was effective antigen for induction of the inhibiting antibody in human, and what correlate with the antibody induction in saliva. Stimulated saliva samples were collected from 151 healthy human subjects (age : 36.6±12.6). NOD-scid mice grafted with human peripheral blood mononuclear cells (PBMC)were used to examine immunogenicity of Pac (361-386) peptide. Hu- IgA and IgG antibody titers to Pac (361-386) peptide in the human saliva and sera from the grafted mice were analyzed by enzyme-linked immuno sorbent assay. Anti-Pac(361-386) peptide IgA antibody titer (a) in human saliva decreased significantly in age-dependent mannaer. MS and mS/ tS ratio were lower in high antibody group (3<a) than no antibody (0.1>a) and moderate group (3≧a>1). Allel DRB1^*1501 and DRB1^*0406 were significantly correlated with high induction of the antibodies and also tended to reduce Lactobacilli and S. mutans. Moreover, the peptide immunogenicities were confirmed in the grafted NOD-scid mice expressing various DRB1 genotypes. The monoclonal antibody induced by the peptide immunization (SH2, KH5) inhibited S. mutans colonization on the tooth surfaces in rats. Therefore, the Pac(361-386) peptide has strong immunogenicity and induce the inhibiting antibody in human oral cavity, and the induction are regulated by aging, HLA-DRB1 genotype and cytokine. These results may be helpful in removing system of oral biofilm using Dental Drug delivary system(3DS) with the human monoclonal antibody to prevent dental diseases.