Macrolide Accumulation by Host Cells in the Gingiva

描述（由申请人提供）：侵袭性和复发性牙周炎与放线菌和牙龈假单胞菌感染有关。这些病原体侵入上皮细胞和吞噬细胞的能力使它们难以通过常规的刮除和刨根来消除。然而，结合刮治和刨根术使用全身抗生素可显著提高临床和微生物治疗效果。虽然没有被牙周病医生广泛使用，但阿奇霉素和克拉霉素可能是有用的，因为它们被宿主细胞吸收并具有良好的抗菌活性。这两种药物在牙龈中的含量都可能高于血清。初步研究表明，主动运输在这一过程中起着重要作用。培养的牙龈成纤维细胞、上皮细胞和pmn样HL-60细胞似乎通过集中和饱和的运输机制吸收克拉霉素。成纤维细胞对克拉霉素的摄取是温度依赖的，并表现出Michaelis-Menten动力学，表明参与了能量依赖的运输系统。我们假设牙龈成纤维细胞、上皮细胞和pmn具有活跃的转运蛋白，可以积累大环内酯类物质。这些转运蛋白通过提高牙周袋软组织壁、吞噬细胞和龈沟液中的大环内酯水平发挥有益作用。由于成纤维细胞是牙龈中一个相对较大的细胞区，大环内酯转运体可以使这些细胞发挥药物储存库的作用，增强和维持牙龈结缔组织和龈沟液中大环内酯的水平。在上皮细胞和pmn中，这些转运体可以通过增强细胞内大环内酯类物质的积累来促进消除侵袭性病原体。本研究的目的是研究大环内酯类药物通过宿主细胞在牙龈内转运和浓缩的机制。具体目的1是表征牙龈成纤维细胞、上皮细胞和pmn接受这些药物的运输系统，使用放射性标记的大环内酯来测定运输活性。[具体目的2]是评估宿主细胞内大环内酯积累的药理学益处（包括对药物分布的影响）。特异性目的3是评估大环内酯在培养的pmn和牙龈上皮细胞中对细胞内放线菌和牙龈假单胞菌的杀伤作用。本研究的主要目的是为选择最有效的牙周抗菌化疗药物提供合理的依据。最终，这项工作可能有助于减少牙齿脱落和治疗牙周炎的费用负担。

DESCRIPTION (provided by applicant): Aggressive and recurrent forms of periodontitis are associated with infections by A. actinomycetemomitans and P. gingivalis. The ability of these pathogens to invade epithelial cells and phagocytes makes them difficult to eliminate by conventional scaling and root planing alone. However, the use of systemic antibiotics in conjunction with scaling and root planing significantly enhances clinical and microbiological treatment outcomes. While not widely prescribed by periodontists, azithromycin and clarithromycin are potentially useful because they are taken up by host cells and have favorable antimicrobial activity. Both agents can potentially attain higher levels in gingiva than in serum. Preliminary studies suggest that active transport plays an important role in this process. Cultured gingival fibroblasts, epithelial cells and PMN-like HL-60 cells appear to take up clarithromycin by a concentrative and saturable transport mechanism. Clarithromycin uptake by fibroblasts is temperature-dependent and exhibits Michaelis-Menten kinetics, suggesting the involvement of an energy-dependent transport system. We hypothesize that gingival fibroblasts, epithelial cells and PMNs possess active transporters that accumulate macrolides. These transporters play a beneficial role by enhancing macrolide levels in the soft tissue wall of the periodontal pocket, in phagocytes and in gingival crevicular fluid. Since fibroblasts are a relatively large cellular compartment of the gingiva, macrolide transporters could permit these cells to function as drug reservoirs that enhance and sustain macrolide levels in gingival connective tissue and in gingival crevicular fluid. In epithelial cells and PMNs, these transporters could facilitate elimination of invasive pathogens by enhancing intracellular macrolide accumulation. The objective of this proposal is to study mechanisms by which macrolides are transported and concentrated by host cells in the gingiva. Specific aim 1 is to characterize transport systems by which gingival fibroblasts, epithelial cells and PMNs take up these agents, using radiolabeled macrolides to assay transport activity. [Specific aim 2 is to assess the pharmacological benefits of intracellular macrolide accumulation by host cells (including effects on drug distribution). Specific Aim 3 is to assess the killing of intracellular A. actinomycetemcomitans and P. gingivalis by macrolides in cultured PMNs and gingival epithelial cells]. The broad goal of this work is to provide a rational basis for selecting the most effective agents for periodontal antimicrobial chemotherapy. Ultimately, this work could contribute to reductions in tooth loss and the cost burden of treating periodontitis. PUBLIC HEALTH RELEVANCE: This proposal will study mechanisms by which macrolide antibiotics are transported and concentrated by cells in human gingiva. Exploiting cellular accumulation of macrolides could enhance the elimination of invasive periodontal pathogens that are difficult to treat with conventional scaling and root planing. By providing a rational basis for selecting an antibiotic for periodontal antimicrobial chemotherapy, this work could ultimately contribute to reductions in tooth loss and the cost burden of treating destructive periodontitis.