Biofilm Elimination and Caries Prevention using Multifunctional Nanocatalysts

尽管与生物膜有关的口腔疾病如龋齿的发病率很高，但临床上没有 破坏有毒生物膜的有效疗法，每年在美国造成400亿美元的支出。 有效控制致龋性生物膜是出了名的挑战，因为细菌被困在一种 富含胞外多糖(EPS)的保护性细胞外基质。此外，EPS缠绕的细菌会产生 促进牙釉质酸溶的高酸性微环境，导致龋齿的发生 龋齿。目前的抗菌剂不能破坏EPS基质或影响EPS的物理性能。 龋齿的化学方面，往往不能有效地杀死生物膜内的微生物，导致有限的 体内药效。为了克服这些显著的障碍，我们开发了一种令人兴奋的治疗策略 使用具有催化活性和pH响应特性的生物兼容氧化铁纳米颗粒(IO-NP)， 展示防生物膜和防龋齿的作用。IO-NP在酸性pH值下表现出类似过氧化物酶的活性 在原位快速激活过氧化氢，同时降解保护性生物膜EPS基质 并在5分钟内以非凡的效率杀死嵌入的细菌(细胞活力降低5个对数)。此外， 在酸性条件下，IO-NP还可以减少磷灰石的脱矿。我们假设IO-NP与 过氧化氢通过纳米催化增强体内抗生物被膜作用和预防龋齿的发生 在酸性pH条件下，增强抗菌、EPS降解和脱矿阻断剂的原位生产。 这项工作的意义在于开发一种可行的、优越的抗生物被膜和龋病预防方法。 与目前的化学形态相比。为了验证我们的假设，我们将优化IO-NP/H_2O_2的功效 进一步改善抗生物被膜和除盐阻挡活动(目标1)。我们将提高催化活性。 通过在纳米粒子中加入特定的金属盐来制备IO-NP，并探讨了不同的葡聚糖对IO-NP的影响。 基于涂层，以提高IO-NP在生物膜结构中的局部化。此外，我们还将把 将磷酸钙转化为IO-NP，以增强其脱矿效果。然后，我们将评估该药的疗效。 使用混合菌种、致龋性生物膜模型，增强IO-NP/H_2O_2的体外生物膜控制作用(目标2)。我们 将使用延时共聚焦和生物物理方法进一步阐明IO-NP/H_2O_2的生物学作用 检测EPS基质的时空降解、细菌杀灭和完整生物膜内的凝聚力。 使用显微硬度和显微CT来评估其对牙釉质脱矿的影响。在《目标3》中，我们将 IO-NP/H_2O_2治疗龋病生物膜的生物相容性和有效性评价 以及使用与临床相关的局部治疗的啮齿动物龋齿模型在体内的龋损发病情况 养生法。成功完成这些目标将为进一步制定方案提供一个框架 和临床疗效研究。重要的是，IO-NP可以低成本地大规模合成，而过氧化氢是 容易获得，这可能导致一种可行的新的局部使用的抗生物被膜/防龋病治疗平台。

Despite the high prevalence of biofilm-related oral diseases such as dental caries, there are no clinically effective therapies to disrupt virulent biofilms, resulting in >$40 billion expenditures annually in the US. Effective control of cariogenic biofilms is notoriously challenging because the bacteria are enmeshed in a protective extracellular matrix rich in exopolysaccharides (EPS). Furthermore, EPS-enmeshed bacteria create highly acidic microenvironments that promote acid-dissolution of tooth enamel, leading to the onset of dental caries. Current antimicrobial agents are incapable of disrupting the EPS matrix or affecting the physico- chemical aspects of caries and often fail to efficiently kill the microbes within biofilms, resulting in limited efficacy in vivo. To overcome these remarkable hurdles, we have developed an exciting therapeutic strategy using biocompatible iron oxide nanoparticles (IO-NP) with catalytic activity and pH-responsive properties that display both anti-biofilm and anti-caries actions. IO-NP exhibit peroxidase-like activity at acidic pH values that rapidly activates hydrogen peroxide (H2O2) in situ to simultaneously degrade the protective biofilm EPS-matrix and kill embedded bacteria with exceptional efficacy (>5-log reduction of cell viability) in 5 minutes. Moreover, IO-NP also reduce apatite demineralization in acidic conditions. We hypothesize that IO-NP synergizes with H2O2 to amplify anti-biofilm effects and prevent the onset of dental caries in vivo via nanocatalysis and enhanced in situ production of antibacterial, EPS-degrading and demineralization-blocking agents at acidic pH. The significance of this work is to develop a feasible and superior anti-biofilm and caries preventive approach compared to current chemical modalities. To test our hypothesis, we will optimize the efficacy of IO-NP/H2O2 to further improve anti-biofilm and demineralizing-blocking activities (Aim 1). We will enhance the catalytic activity of IO-NP by inclusion of specific metal salts into the nanoparticles, and explore the effects of various dextran- based coatings to increase IO-NP localization within biofilm structure. Furthermore, we will incorporate calcium-phosphate into IO-NP to enhance its effects on demineralization. Then, we will evaluate the efficacy of enhanced IO-NP/H2O2 for biofilm control in vitro using a mixed-species, cariogenic biofilm model (Aim 2). We will further elucidate the biological actions of IO-NP/H2O2 using time-lapsed confocal and biophysical methods to examine spatiotemporal degradation of EPS-matrix, bacterial killing and cohesiveness within intact biofilms. The effects on enamel demineralization will be assessed using micro-hardness and micro-CT. In Aim 3, we will evaluate the biocompatibility and efficacy of the developed IO-NP/H2O2 therapy in hindering cariogenic biofilms and the onset of carious lesions in vivo using a rodent caries model with a clinically-relevant topical treatment regimen. Successful completion of these aims will provide a framework for further formulation development and clinical efficacy studies. Importantly, IO-NP can be synthesized with low cost at large scale while H2O2 is readily available, which could lead to a feasible new anti-biofilm/anti-caries therapeutic platform for topical use.