Vitamin D induction of antibacterial activity in gingival cells

描述（由申请人提供）：抗菌肽如2-防御素和抗菌肽为牙龈上皮提供了抵抗致病菌的第一道防线。体外和体内实验表明，这些肽在先天宿主防御中既可以作为直接的抗菌剂，也可以作为连接先天和适应性免疫反应的趋化因子。活细菌和细菌产物在牙龈上皮细胞（GEC）中增加了这两类肽的产生，这表明有可能在治疗上调节这些抗生素药物的水平。最近，2 -防御素和抗菌素LL-37的表达在髓细胞和几种细胞系中被一种毒性较小的物质，维生素D的激素活性形式，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。我们的初步结果表明，1,25(OH)2D3可以提高气道上皮细胞（AEC）和GEC中LL-37 mRNA的水平，以及气液界面（ALI）中生长的AEC气道表面液中LL-37依赖性的抗菌活性。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调查维生素D作为一种药物的潜力，以帮助先天免疫防御口腔病原体。在这一建议中，我们将重点关注在细胞系和人牙龈上皮原代培养中诱导抗菌肽基因表达和抗放线菌聚集杆菌（Aa）的活性。为了验证我们的假设，我们建议：1)量化健康个体牙龈上皮细胞原代培养中维生素d介导的抗菌肽表达和抗菌活性的诱导，并将其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）介导的信号转导通路，明确维生素D诱导牙龈细胞LL-37的机制。发现一种可以通过转录机制提高牙龈抗菌能力的无毒药物，对牙周病和口腔感染性疾病的研究具有深远的意义。我们希望这项初步研究的结果将成为深入研究125 (OH)2D3的效用以增强当前牙周病治疗的基础。

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.