ADHERENCE OF PERIODONTAL DISEASE-ASSOCIATED BACTERIA

这项研究的长期目标是阐明分子 重要口腔细菌对牙齿的附着机制， 制定方法，用于调节突出的附着和殖民化 口腔中的牙周病原体。 建议的具体目标 项目：（1）识别、分离和表征相关粘附 与富含脯氨酸的蛋白质相互作用的放线菌1型菌毛 被认为是实验唾液膜中的受体;（2） 确定附着体与实验牙齿的粘附相关性 表面和天然牙齿在体内;（3）建立功能相关性 (i.e.,粘附抑制活性（ALA），以及对感染的调节， 体内）的特异性的遗传调节的变化 从用粘性放线菌T14 V免疫的近交系小鼠获得抗体，并研究 AIA在人类中受基因调控的可能性。 获得的数据 这个项目将有望成为未来研究的模型 研究其他细菌附着和定植的分子机制 与人类牙周病相关的口腔微生物。 在 此外，如果AIA对放线菌具有功能相关性， 殖民化，并在人类中进行遗传调节，希望 为建立这种关联而制定的原则和方法可以 也适用于主要的牙周病病原体。 这些研究应 导致制定更好的办法，防止 牙周病原体的免疫调节。 为了鉴定粘附，将进行抗体介导的吸附抑制。 在体外羟基磷灰石-细菌吸附试验中进行研究;以及 生物化学（快速蛋白质液相色谱法，差示和蔗糖- 梯度离心等）和免疫学（亲和层析 使用多克隆或单克隆抗粘附抗体）方法将 用于粘合剂的纯化。 分子机制 将在可能的情况下，通过研究 耐药菌毛缺陷型和粘附型的吸附， 粘性放线菌T14 V缺陷突变体对从提取的第三代 臼齿 功能相关性（即，AIA）基因调控 来自近交系小鼠的血清抗体特异性的变化将是 通过检测各种抗体谱型和独特型进行评价 使用等电聚焦从免疫小鼠的血清中纯化， 独特型特异性ELISA，在AIA测定中。 初步实验还将 研究AIA在人类中受基因调控的可能性， 用一组家系血清进行AIA的分离和连锁分析 这些人之前已经在5号染色体上进行了HLA分型， 11号染色体上的同种异型。 为了确定AIA的基因调控是否 影响口腔定植，产生高或低水平的近交系小鼠 的AIA将用纯化的1型菌毛相关粘附免疫 并用菌株T14 V进行攻击，以确定AIA水平是否与 牙齿上T14 V的定植水平。

The long-term objectives of this research are to elucidate molecular mechanisms of attachment for important oral bacteria to teeth, and to develop approaches for modulating attachment and colonization of prominent periodontopathogens in the oral cavity. The Specific Aims of the proposed project: (1) identify, isolate and characterize the adhesion associated with Actinomyces type 1 fimbriae which interacts with proline-rich proteins thought to be receptors in the experimental salivary pellicle; (2) establish relevance of the adhesion on attachment to experimental tooth surfaces and natural teeth in vivo; (3) establish functional relevance (i.e., adherence inhibition activity (ALA), and modulation of infection in vivo) of genetically regulated variations in specificity of serum antibodies from inbred mice immunized with A.viscosus T14V, and investigate the possibility that AIA is genetically regulated in humans. Data obtained from this project will hopefully serve as a model for future studies investigating molecular mechanisms of attachment and colonization of other oral microorganisms associated with periodontal diseases in humans. In addition, if AIA has functional relevance regarding actinomyces colonization and is genetically regulated in humans, it is hoped that principles and methods developed to establish this association can be applied to prominent periodontopathogens as well. These studies should lead to development of better approaches for preventing colonization of periodontopathogens by immune modulation. To identify the adhesion, antibody-mediated adsorption inhibition will be studied in an in vitro hydroxyapatite-bacterial adsorption assay; and biochemical (Fast Protein Liquid Chromatography, differential and sucrose- gradient centrifugation, etc.) and immunologic (affinity-chromatography using polyclonal or monoclonal anti-adhesion antibodies) methods will be used for the purification of the adhesion. Molecular mechanisms established in vitro will be confirmed in humans where possible by studying the adsorption of antibiotic-resistant fimbrial-deficient and adherence- defective mutants of A.viscosus T14V to enamel slabs cut from extracted 3rd molars. The functional relevance (i.e., AIA) of genetically regulated variations in the specificity of serum antibodies from inbred mice will be evaluated by testing various spectrotypes and idiotypes of antibodies purified from sera of immunized mice using isoelec-tric focusing and idiotype-specific ELISAs, in AIA assays. Preliminary experiments will also investigate the possibility that AIA is genetically regulated in humans by segregation and linkage analysis of AIA using sera from a group of families who have been previously typed for HLA on chromosome 5 and immunoglobulin allotype on chromosome 11. To establish whether genetic regulation of AIA influences oral colonization, inbred mice which produce high or low levels of AIA will be immunized with purified type 1 fimbriae-associated adhesion and challenged with strain T14V to determine if AIA levels correlate with the level of T14V colonization on teeth.