Vitamin D induction of antibacterial activity in gingival cells

描述（由申请人提供）：抗菌肽如2-防御素和凯萨林菌素为牙龈上皮提供了抵抗病原菌的第一道防线。体外和体内实验表明，这些肽在先天宿主防御中作为直接抗微生物剂和作为可以连接先天和适应性免疫应答的趋化因子的作用。这两类肽的生产增加牙龈上皮细胞（GEC）的活细菌和细菌的产品，这表明治疗调节这些抗生素的药物水平的潜力。最近，2-防御素和cathelicidin LL-37的表达在骨髓细胞和几种细胞系中由毒性较小的药剂，维生素D的神经活性形式，1，25（OH）2-维生素D3诱导。这种诱导也与这些细胞分泌的抗菌活性的增加相对应。我们的初步研究结果表明，1，25（OH）2D 3可以增加气道上皮细胞（AEC）和GEC中LL-37 mRNA的水平，以及在气液界面（ALI）中生长的AEC气道表面液中LL-37依赖的抗菌活性。已发表的数据表明维生素D受体（VDR）基因多态性与早发性牙周炎之间存在关联。基于这些结果，我们假设生理剂量的维生素D可以增加口腔上皮细胞的抗菌肽表达和抗菌活性。这项初步研究的目的是研究维生素D作为一种试剂的潜力，以帮助先天免疫防御口腔病原体。在这个建议中，我们将集中在诱导cathelicidin基因的表达和活性对Aggregatibacteractinomycetemcomitans（Aa）在细胞系和原代培养人牙龈上皮。为了验证我们的假设，我们提出：1）定量维生素D介导的健康人牙龈上皮细胞原代培养物中抗菌肽表达和抗菌活性的诱导，并与牙周炎患者细胞的反应进行比较; 2）通过检测维生素D受体元件（VDRE），明确维生素D诱导牙龈细胞中LL-37的机制。介导的信号转导途径。鉴定出一种能够通过转录机制增加牙龈抗菌能力的无毒试剂，在牙周病和口腔感染性疾病的研究中具有深远的意义。我们希望，这项试点研究的结果将成为深入研究1，25（OH）2D 3的效用以增强目前牙周病治疗的基础。 公共卫生相关性：牙周病是由细菌引起的，这些细菌粘附并定植在牙齿附近的牙龈细胞上。为了提高这种组织阻止细菌生长的能力，研究人员已经研究了具有有效抗生素活性的小蛋白质的表达，称为抗菌肽。我们已经证明，这些肽之一，LL- 37，可以增加牙龈细胞的治疗与低剂量的维生素D。该提案旨在研究我们是否可以对使用维生素D作为牙周病预防性治疗的潜力进行更深入的研究。

DESCRIPTION (provided by applicant): Antimicrobial peptides such as 2-defensins and cathelicidins provide the gingival epithelium with a first line of defense against pathogenic bacteria. In vitro and in vivo experiments have indicated a role for these peptides in the innate host defense as both direct antimicrobial agents and as chemokines which can link the innate and adaptive immune responses. Production of both classes of peptides is increased in gingival epithelial cells (GEC) by live bacteria and by bacterial products, suggesting the potential for modulating levels of these antibiotic agents therapeutically. More recently, expression of 2-defensins and the cathelicidin LL-37 were induced in myeloid cells and several cell lines by a less toxic agent, the hormonally active form of vitamin D, 1,25(OH)2-vitamin D3. This induction corresponded with an increase in antibacterial activity secreted by these cells as well. Our preliminary results demonstrate that 1,25(OH)2D3 can increase LL-37 mRNA levels in both airway epithelial cells (AEC) and GEC, and LL-37-dependent antibacterial activity in airway surface fluid of AEC grown in an air-liquid interface (ALI). Published data has demonstrated an association between polymorphisms of the vitamin D receptor (VDR) gene and early-onset periodontitis. Based upon these results, we hypothesize that physiological doses of Vitamin D can increase antimicrobial peptide expression and antibacterial activity of oral epithelial cells. The goal of this pilot study is to investigate the potential of vitamin D as an agent to aid in the innate immune defenses against oral pathogens. In this proposal we will focus on the induction of cathelicidin gene expression and activity against Aggregatibacter actinomycetemcomitans (Aa) in cell lines and in primary cultures of human gingival epithelium. To address our hypothesis, we propose to: 1) Quantify the vitamin D-mediated induction of antimicrobial peptide expression and antibacterial activity in primary cultures of gingival epithelial cells from healthy individuals, and compare the response with cells from periodontitis patients; and 2) Define the mechanism of induction of LL-37 in gingival cells by vitamin D by the examination of Vitamin D receptor element (VDRE)-mediated signal transduction pathways. Identification of a non- toxic agent that could increase antibacterial capabilities of the gingiva through transcriptional mechanisms could have long-reaching implications in the study of periodontal disease and oral infectious diseases in general. It is our hope that the results from this pilot study will be a foundation to develop an in-depth study of the utility of 1,25 (OH)2D3 to augment current therapies for periodontal disease. PUBLIC HEALTH RELEVANCE: Periodontal disease is caused by bacteria that adhere to and colonize the gingival cells adjacent to the teeth. To increase the ability of this tissue to prevent bacterial growth, investigators have examined the expression of small proteins with potent antibiotic activity, called antimicrobial peptides. We have shown that one of these peptides, LL- 37, can be increased in gingival cells by treatment with low doses of vitamin D. This proposal seeks to examine whether we can develop more in-depth studies of the potential for using vitamin D as a preventative treatment for periodontal disease.