Molecular Mechanisms of Squamous Metaplasia in Dry Eye

描述(申请人提供)：在系统性自身免疫性疾病中，如Sj“gren‘s综合征(SS)，泪腺外分泌病导致缺水的干眼，这是最常见和令人虚弱的临床表现之一。随着干眼病的发展，它启动了一个多步骤的、免疫介导的过程，包括两个病理特征：(1)将非角化性的眼粘膜表面转分化为角化性的“皮肤样”上皮；(2)眼表和杯状细胞粘蛋白的破坏。这一过程称为鳞状化生(SQM)，是干眼病的破坏性终末期后果，可导致相当大的发病率，因为晚期角化与上皮下纤维化结合导致角膜混浊和失明。虽然免疫介导的炎症与SQM的发病机制有关，但对确切的免疫致病机制知之甚少，留给我们的治疗策略是有限的、昂贵的，而且主要是姑息治疗。我们研究的目标是破译自身免疫介导的炎症如何引发威胁视力的SQM。使用三个模型系统，(I)人类SS患者；(Ii)模拟SS临床特征的自发性自身免疫性外分泌病和角膜病变的有效小鼠模型；以及(Iii)体外培养的角膜上皮细胞研究，我们先前的工作证明了自身反应性CD4+T细胞及其与促炎细胞因子IL-1的相互作用在SQM的发病机制中的重要作用。然而，CD4+T细胞和IL-1促进自身免疫性干眼症的SQM的具体机制仍不清楚。为了加深我们对免疫介导的SQM的分子和细胞致病机制的理解，并确定可能的治疗靶点，我们试图(I)确定CD4+细胞如何与IL-1一起工作来激发SQM；以及(Ii)IL-1如何促进眼表上皮向粘蛋白耗竭和病理性角化的转分化。我们假设自身抗原激活的CD4+T细胞与浸润性单核细胞协同工作，促进局部促炎介质(如IL-1)的释放，从而在眼表建立并维持慢性炎症状态。一旦确定，局部炎症反应通过破坏眼粘膜表型的主要调节因子PAX6，改变眼粘蛋白的糖基化和表达，导致眼表损伤并促进SQM。为了验证这一假设，我们将(目标1)探索渗透的单个核细胞作为潜在的细胞中间体，将抗原启动的CD4+T细胞与局部产生的IL-1联系起来；(目标2)通过研究IL-1对控制角膜表型的主要基因Pax6的影响，确定IL-1如何指导病理性角化的早期阶段；以及(目标3)通过识别调节糖结合物生物合成的关键基因来表征IL-1改变眼粘蛋白的机制。这项工作的总体影响将包括更好地了解自身免疫性疾病中SQM的免疫病理学，这可能会发现有助于其早期诊断和/或治疗的关键事件。

DESCRIPTION (provided by applicant): In systemic autoimmune diseases like Sj"gren's syndrome (SS), exocrinopathy of the lacrimal gland leads to an aqueous-deficient dry eye that is among the most common and debilitating clinical manifestations. As dry eye disease progresses, it initiates a multi-step, immune-mediated process that includes two pathological characteristics (1) transdifferentiation a nonkeratinized, mucosal ocular surface to a keratinized, "skin-like" epithelium; and (2) disruption of ocular surface and goblet cell mucins. This process, known as squamous metaplasia (SQM), is a devastating, end-stage consequence of dry eye disease that can cause considerable morbidity as advanced keratinzation couples with subepithelial fibrosis to cause corneal opacification and blindness. While immune-mediated inflammation has been implicated in the pathogenesis of SQM, little is known about the precise immunopathogenic mechanisms, leaving us with treatment strategies that are limited, costly and largely palliative. The goal of our research is to decipher how autoimmune-mediated inflammation provokes vision-threatening SQM. Using three model systems, (i) human patients with SS; (ii) a validated mouse model of spontaneous autoimmune exocrinopathy and keratopathy that mimics the clinical characteristics of SS; and (iii) in vitro studies of cultured corneal epithelial cells, our previous work demonstrated an essential role for autoreactive CD4+ T cells and their interplay with the proinflammatory cytokine IL-1 in the pathogenesis of SQM. Yet, the specific mechanism whereby CD4+ T cells and IL-1 promote SQM in autoimmune dry eye remains unknown. To deepen our understanding of the molecular and cellular pathogenic mechanism of immune-mediated SQM, and to identify possible targets for treatment, we seek to (i) determine how CD4+ cells work together with IL-1 to provoke SQM; and (ii) how IL-1 promotes transdifferentiation of the ocular surface epithelium to one that is mucin-depleted and pathologically keratinized. We hypothesize that autoantigen-primed CD4+ T cells work in collaboration with infiltrating monocytes to promote the local release of proinflammatory mediators (e.g., IL-1) that establish and sustain a chronic inflammatory state at the ocular surface. Once established, the local inflammatory response causes ocular surface damage and promotes SQM by disrupting, PAX6, the master regulator of ocular mucosal phenotype and altering the glycosylation and expression of ocular mucins. To test this hypothesis we will (Aim 1) explore infiltrating mononuclear cells as potential cellular intermediate linking antigen-primed CD4+ T cells to the local production of IL-1; (Aim 2) define how IL-1 directs the early stages of pathological keratinization, by studying its effects on the master gene controlling corneal phenotype, Pax6; and (Aim 3) characterize the mechanisms whereby IL-1 alters ocular mucins by identifying key genes regulating glycoconjugate biosynthesis. The overall impact of this work will include a better understanding of the immunopathology of SQM in autoimmune disease that is likely to uncover key events that aid in its early diagnosis and/or treatment.