Molecular Mechanisms of Squamous Metaplasia in Dry Eye

描述（由申请人提供）：在系统性自身免疫性疾病中，如Sj"gren's综合征（SS），泪腺外源性病变导致缺水性干眼，这是最常见和最虚弱的临床表现之一。随着干眼病的进展，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)转分化，从非角化的眼粘膜表面到角化的“皮肤样”上皮；(2)眼表和杯状细胞粘蛋白的破坏。这一过程被称为鳞状皮化生（SQM），是干眼病的一种毁灭性的终末期后果，可引起相当大的发病率，因为晚期角化加上上皮下纤维化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，但对精确的免疫致病机制知之甚少，这使得我们的治疗策略有限，成本高，而且很大程度上是姑息性的。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。使用三种模型系统，(i)人类SS患者；（ii）模拟SS临床特征的自发性自身免疫性外源性病变和角膜病变小鼠模型；（iii）在体外培养角膜上皮细胞的研究中，我们之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的发病机制中起重要作用。然而，CD4+ T细胞和IL-1促进自身免疫性干眼症SQM的具体机制尚不清楚。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，我们寻求(i)确定CD4+细胞如何与IL-1共同作用以引发SQM；以及（ii） IL-1如何促进眼表上皮的转分化，使其成为粘液蛋白耗竭和病理性角化的细胞。我们假设自体抗原引发的CD4+ T细胞与浸润的单核细胞协同作用，促进促炎介质（如IL-1）的局部释放，从而在眼表建立并维持慢性炎症状态。一旦建立，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（目的1）探索浸润单核细胞作为潜在的细胞中介，将抗原引发的CD4+ T细胞与IL-1的局部产生联系起来；（Aim 2）通过研究IL-1对控制角膜表型的主基因Pax6的影响，确定IL-1如何指导病理性角化的早期阶段；（Aim 3）通过鉴定调节糖缀合物生物合成的关键基因，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理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. PUBLIC HEALTH RELEVANCE: Autoimmune diseases like Sj"gren's syndrome cause a severe dry eye that is highly recalcitrant to treatment. Despite powerful immunosuppressive and immunomodulatory therapy, autoimmune dry eye can progress to complete corneal opacification and blindness through a process known as squamous metaplasia. Little is known about the pathogenesis of squamous metaplasia and there is no cure. A better understanding of the underlying disease process would open the possibility of developing novel treatments to prevent corneal blindness.