Elucidation of the caries-inducing mechanism by S.mutans and development of a diagnostic method to estimate caries-risk.

最近，我们构建了分别表达编码GTF-I、-SI和-S酶的变形s.m ileri转化子KSB8、KSC43和NH5，并建立了能产生三种gtf单特异性单克隆抗体的杂杂瘤。在本研究中，我们比较了三种转化体的体内和体外蛀牙性，以阐明变形链球菌诱导蛀牙的机制。此外，利用单特异性单克隆抗体，建立了一种高灵敏度的Gross-dot法测定三种gtf。实验结果如下：1。从感染KSC43.2的大鼠口腔中重新分离出更稳定的表达gtfc的转化KSC43-RT。通过定殖试验检测了转化子的体外龋齿性。结果表明，只有KSC43-RT细胞能在玻璃表面定植。用大鼠模型系统测试了转化子的体内致病性。结果表明，KSC43-RT细胞可诱导颊部和颊部龋齿，KSB8细胞可诱导近端龋齿，而NH5细胞则不能。建立并应用高灵敏度粗点法评价变形链球菌的致病性。用该方法分析了不同临床分离株的细胞相关GTFs。结果表明，3种gtf的含量及其比值在不同菌株间存在显著差异。这些结果表明，gtfC产物对突变s.t ans引起的龋齿发育特别重要，高灵敏度的Gross-dot法可能有助于准确估计突变s.t ans单个菌株的致病性。

Recentry, we have constructed S.milleri transformants KSB8, KSC43 and NH5 which individually express S.mutans gtfB,gtfC and gtfD coding GTF-I,-SI and -S enzymes, and established hybridoma which produce monoclonal antibodies mono-specific to the three GTFs. In this study, in vitro and in vivo cariogenicities of the three transformants were compared to elucicate the caries-inducing mechanism by S.mutans. Furthermore, using the mono-specific MAbs, a high-sensitive Gross-dot method for determination of the three GTFs was developed. The results obtained were as follows :1. More stable gtfC-expressing transformant KSC43-RT was reisolated from an oral cavity of rat infected with KSC43.2. In vitro cariogenicity of the transformants was examined by a colonization test. The results showed that only KSC43-RT cells could makedly colonized on glass surfaces.3. In vivo cariogenicity of the transformants was tested by a rat model system. The results showed that the KSC43-RT cells could induced the buccal and sulcal caries, and the KSB8 cells could induced the proximal caries, but the NH5 cells did not.4. High-sensitive Gross-dot assay method was established and applied in order to evaluate the cariogenicity of S.mutans strains.5. Cell-associated GTFs from various clinical isolates of S.mutans were analyzed by the method. The results showed that the levels of three GTFs and those ratio were markedly different among the isolates.These results suggest that the gtfC product was especially important for caries development caused by S.mutans, and that the high-sensitive Gross-dot method may be useful to estimate the exact cariogenicity of individual strain of S.mutans.