Augmenting Innate Immunity to Protect the Oral Mucosa

描述（由申请人提供）：在尤里卡项目（RFA-GM-10-009）中，我们将加速知识的发展，以找到一种临床方法来阻止与牙周炎和念珠菌病相关的侵入性口腔病原体。口腔角质形成细胞似乎使用两个区室化的细胞内抗微生物效应子系统来保护和抑制侵入性病原体：胞质溶胶中的钙卫蛋白和内体中的LL-37。通过靶向这些效应物，我们计划使用两种方法来增强角质形成细胞对侵入性病原体的细胞内抵抗力。我们假设角质形成细胞内对侵入性微生物病原体的抗性可以通过瞬时递送特异性抗微生物效应物mRNA（例如，S100 A8和S100 A9）或特异性激动剂（例如，维生素D类似物以上调LL-37）。为了避免使用转基因，我们将使用将钙卫蛋白和/或LL-37 mRNA引入角质形成细胞的不寻常方法。为了比较，在一些实验中，我们将用维生素D类似物刺激细胞以上调LL-37。我们将了解是否可以选择性地增强对入侵微生物的内体和细胞质抗性。为了了解这些特定的mRNA或维生素A类似物应用于完整组织是否有效，并考虑细胞毒性，促炎免疫和致癌副作用，我们将同时在人体扁桃体外植体体外模拟该系统。总的来说，这些实验将表征维生素D类似物和抗微生物效应物mRNA的转移，用于增强口腔角质形成细胞对侵入性口腔病原体的细胞内先天抗性的机制和有效性。我们的策略可能被证明是对大多数常见口腔感染的疫苗和抗生素的宝贵补充，这些感染不会危及生命，发病率相对较低。对于这些感染，疫苗具有不可接受的风险效益比。抗生素增加了微生物耐药性和宿主过敏的风险。我们的方法可以避免这些担忧。

DESCRIPTION (provided by applicant): In this EUREKA project (RFA-GM-10-009), we will accelerate the development of knowledge towards a clinical approach to thwart invasive oral pathogens associated with periodontitis and candidiasis. Oral keratinocytes appear to protect against and suppress invasive pathogens using two compartmentalized, intracellular antimicrobial effector systems: calprotectin in the cytosol and LL-37 in endosomes. By targeting these effectors, we plan to use two approaches to augment keratinocyte intracellular resistance to invasive pathogens. We hypothesize that intra-keratinocyte resistance against invasive microbial pathogens can be increased by transient delivery of specific antimicrobial effector mRNAs (e.g., S100A8 and S100A9) or specific agonists (e.g., vitamin D analogues to upregulate LL-37). Avoiding the use of transgenes, we will use the unusual approach of introducing calprotectin and/or LL-37 mRNAs into the keratinocyte. For comparison, in some experiments we will stimulate cells with vitamin D analogues to upregulate LL-37. We will learn whether endosomal and cytoplasmic resistance against invasive microbes can be selectively augmented. To learn whether the application of these specific mRNAs or vitamin A analogues to intact tissues will prove efficacious, and to consider cytotoxic, proinflammatory immune, and carcinogenic side effects, we will concurrently model this system in human tonsil explants ex vivo. Collectively, the experiments will characterize vitamin D analogues and transfer of antimicrobial effector mRNAs for the mechanism and effectiveness in augmenting intracellular innate resistance to invasive oral pathogens in oral keratinocytes. Our strategy may prove to be a valuable compliment to vaccines and antibiotics for most common oral infections, which are not life threatening and have relatively low morbidity. For these infections, vaccines have an unacceptable risk-benefit ratio. Antibiotics promote the risk of microbial resistance and host allergy. Our approach would circumvent these concerns.