Epithelial Cell TLRs in Disease Susceptibility

描述(由申请人提供)：人类会患上牙周炎和牙周炎，以应对微生物牙菌斑产生的挑战。不同的人对牙周炎(浅表性炎症)和牙周炎(广泛的炎症会导致骨质和牙齿脱落)的易感性不同。牙周病的病因尚不完全清楚，然而，人们普遍认为，宿主对微生物菌斑的反应与宿主的先天、炎症或免疫防御系统有关。单核苷酸多态(SNPs)存在于人类基因中，编码许多与宿主-微生物相互作用相关的分子。牙周炎(牙周炎和牙周炎)始于细菌通过包括Toll样受体(TLRs)在内的受体对上皮细胞的干扰。我们的初步数据表明，对细菌和细胞因子扰动的反应产生的促炎细胞因子和趋化因子在个体之间存在差异，这些差异可能与携带特定的TLR4基因有关。我们的一般假设是，牙龈上皮细胞对外来挑战的个体反应特征与其细胞内TLR基因有关，TLR基因影响牙周炎和牙周炎的易感性。因此，我们将利用多个原代人类牙龈上皮细胞(HGECs)、临床研究以及基因和表型分析来解决以下目标。第一个目的是利用定量聚合酶链式反应、基因表达谱分析、蛋白质分析和免疫反应性来表征HGEC中TLR基因表达的患者间差异，并将其与TLR基因型别相关联。其次，对转基因HGEC的功能分析(使用siRNA敲除和过表达)将阐明TLR4、Asp299Gly和其他SNP在上皮内毒素低反应中的作用。第三，我们的目标是通过临床确定TLR4基因多态和低反应性是否与实验性牙周炎的严重程度相关，以及它们是否可以区分牙周炎病例和对照，从而翻译这项工作。最后，我们的目标是挑战HGECs和其他类型的细胞、单核细胞、肠道上皮细胞和内皮细胞，以评估与TLR4相关疾病(如炎症性肠病、动脉粥样硬化和哮喘)的病因相关的内毒素反应性。反应差异的表征和相关SNPs的测定可能导致一系列传染病和炎症性疾病的新疗法或诊断。

DESCRIPTION (provided by applicant): Humans develop gingivitis and periodontitis in response to the challenges produced by microbial dental plaque. Individuals vary in their susceptibility to both gingivitis (superficial inflammation) and periodontitis (extensive inflammation causing bone and tooth loss). The etiology of periodontal disease is not fully understood; however, it is accepted that variance in the human host response to microbial plaque will relate to the host's innate, inflammatory or immune defense systems. Single nucleotide polymorphisms (SNPs) exist in the human genes encoding many molecules pertinent to the host-microbe interaction. Periodontal inflammation (gingivitis and periodontitis) begins with the perturbation of the epithelial cells by bacteria via receptors, including Toll-like Receptors (TLRs). Our preliminary data indicates that the pro-inflammatory cytokines and chemokines produced in response to bacterial and cytokine perturbations differ between individuals and these differences may relate to carriage of a specific TLR4 polymorphism. Our general hypothesis is that individual response characteristics of gingival epithelial cells to foreign challenges, relate to their cellular TLR genotype which influences susceptibility to gingivitis and periodontitis. Thus we will address the following aims utilizing multiple primary human gingival epithelial cells (HGECs), clinical studies and genotype and phenotype analyses. The first aim is to characterize molecularly the inter-patient differences in TLR expression in HGECs using qPCR, gene expression profiling, protein assays and immunoreactivity and to correlate this to the TLR genotype. Secondly, functional analysis of genetically- modified HGECs (using siRNA knock-downs and over-expression) will elucidate the role of TLR4 and the Asp299Gly and other SNPs in epithelial LPS hypo-responsiveness. Thirdly, we aim to translate this work by determining clinically if TLR4 polymorphisms and hypo-responsiveness correlate with the severity of experimental gingivitis and if they can differentiate periodontitis cases and controls. Finally, we aim to challenge HGECs and other cell types, monocytes, gut epithelia and endothelial cells to assess LPS responsiveness relevant to the etiopathology of TLR4 associated diseases such as inflammatory bowel disease, atheroma and asthma. Characterization of response differences and determination of related SNPs may lead to new therapeutics or diagnostics in a range of infectious and inflammatory diseases.