Virus Infections In The Eye

我们对病毒和寄生虫在眼睛微环境中复制时发生的各种病毒学和免疫病理过程的研究包括四个方面：(1)病毒诱导的视网膜退化过程；(2)病毒在人类疾病中的可能作用；(3)人类巨细胞病毒(CMV)感染的分子诊断和发病机制；(4)眼睛的疱疹病毒感染。我们已经建立了一个研究视网膜退行性疾病的模型系统-实验性冠状病毒视网膜病变(EcoR)。该病毒能够在存在轻度视网膜血管炎症的情况下引发急性感染。最初的视网膜损伤之后是感染性病毒的清除和进行性的视网膜变性。这是第一个显示病毒引起的变性、病毒持久性、对病毒引起的组织损伤的遗传易感性和病毒引发的自身免疫反应的视网膜模型。我们的目标是确定视网膜退行性疾病的病理生理机制，并确定与之相关的基因。在过去的一年里，我们取得了以下重要发现。我们评估了非常早期的细胞因子和趋化因子谱，作为衡量冠状病毒感染的小鼠免疫反应强度的指标，这些小鼠具有视网膜变性易感性和视网膜变性抵抗背景。这些研究发现，这两个小鼠品系产生的早期免疫反应有明显的差异。这些差异体现在干扰素-伽马和由干扰素-伽马引发的两种趋化因子CXCL9和CXCL10的产生上。在第2天和第3天，BALB/c小鼠的血清中有较高水平的干扰素-γ、CXCL9和CXCL10。与此同时，在CD-1小鼠的血清中检测到的这些分子的水平要低得多。此外，实时荧光定量聚合酶链式反应分析证实，CXCL9和CXCL10基因在Babl/c小鼠的视网膜中的表达显著高于CD-1小鼠。这些研究确实令人兴奋，并确定了可能的机制，使BALB/c小鼠具有强大的免疫反应，可能触发自身免疫成分。CXCL9和CXCL10是与存在于活化的T细胞和NK细胞上的CXCR3相互作用的有效趋化因子。它们指导这些细胞的迁移并刺激它们的黏附。这些T细胞参与针对视网膜内感染和自身靶点的免疫反应。 角膜疱疹感染引起强烈的炎症反应，最终导致威胁视力的间质角膜炎，这是潜伏病毒频繁重新激活的后遗症。疱疹间质角膜炎(HSK)的病因学被认为是一种异常的TH1细胞因子介导的免疫病理。HSK在缺乏高水平感染性HSV的情况下进展，与血管生成和炎症介质有关。在HSK患者中，HSV在重新激活期间的传播受到中和抗体的限制。因此，HSV的成分，如HSV-DNA和中和的HSV-Ig G复合体(IC)可能通过触发血管生成和炎症介质而参与HSK的发生。体外模型系统被用来评估病毒成分诱导炎症和血管生成的机制。我们发现，当HSV-DNA或中和HSV-IC处理时，常驻角膜细胞(成纤维细胞和上皮细胞)和巨噬细胞被激活。这些激活的细胞增加了血管内皮生长因子和基质金属蛋白酶-9的基因表达和蛋白产生。此外，这些细胞还会释放促炎症细胞因子，从而进一步诱导血管生成因子。进一步的分析表明，参与这些过程的机制包括Fc受体和TLR的相互作用。因此，HSV-DNA和HSV-Ig G IC在眼部微环境中的持续存在可能有助于HSK的血管生成和炎症反应，细胞因子和TLR可能是干预的潜在靶点。

Our studies of various virologic and immunopathologic processes that occur when viruses and parasites replicate in the ocular microenvironment comprise four areas: (1) virus induced retinal degenerative processes; (2) the possible roles of viruses in human diseases; (3) molecular diagnosis and pathogenesis of cytomegalovirus (CMV) infections in man; and (4) herpesvirus infections of the eye. We have established a model system for studying retinal degenerative diseases, experimental coronavirus retinopathy (ECOR). The virus is capable of inducing an acute infection in the presence of mild retinal vascular inflammation. Initial retinal damage is followed by clearance of infectious virus and progressive retinal degeneration. This is the first retinal model to demonstrate a virus induced degeneration, viral persistence, a genetic predisposition to virus induced tissue damage and a virus triggered autoimmune response. Our goal is to determine the pathophysiological mechanisms and to identify genes involved in the retinal degenerative disease. During the past year we have made the following key findings. We evaluated very early cytokine and chemokine profiles as a measure of intensity of immune reactivity in coronavirus infected mice with a retinal degeneration susceptible and a retinal degeneration resistant background. These studies identified a distinct difference in the early immune response that is generated by the two mouse strains. These differences are noted in the production of IFN-gamma and the two chemokines triggered by IFN-gamma, CXCL9 and CXCL10. At day 2 and 3 PI, BALB/c mice have high levels of IFN-gamma, CXCL9 and CXCL10 in their sera. At the same time, significantly lower levels of these molecules are detected in sera from CD-1 mice. Moreover, real time PCR analysis of retinas identified that CXCL9 and CXCL10 gene expression is significantly greater in retinas from BABL/c mice in comparison to CD-1 mice. These studies are truly exciting and identify possible mechanisms that allow the BALB/c mouse to have a robust immune response that could trigger an autoimmune component. CXCL9 and CXCL10 are potent chemokines that interact with CXCR3 present on activated T cells and NK cells. They direct the migration and stimulate the adhesion of these cells. These T cells participate in immune reactivity against infected and self targets within the retina. Corneal herpes infection elicits a robust inflammatory response and eventually leads to a vision-threatening stromal keratitis as a sequela of frequent reactivation of latent virus. The aetiology of herpetic stromal keratitis (HSK) is thought to be an aberrant TH1 cytokine mediated immunopathology. HSK progresses in the absence of high levels of infectious HSV and is associated with angiogenesis and inflammatory mediators. In HSK patients, spread of HSV during reactivation is limited by neutralizing antibody. Therefore, components of HSV, such as HSV-DNA and neutralized HSV-IgG complexes (IC) may contribute to HSK by triggering mediators of angiogenesis and inflammation. An in vitro model system was employed to evaluate mechanisms by which viral components could induce inflammation and angiogenesis. We found that resident corneal cells (fibroblasts and epithelial) and macrophages are activated when treated with HSV-DNA or neutralized HSV-IC. These activated cells have augmented gene expression and protein production of VEGF and MMP-9. Moreover, these cells release proinflammatory cytkines that can further induce angiogenic factors. Further analysis revealed that the mechanisms involved in these processes include the Fc receptors and TLR interactions. Therefore, the continued presence of HSV-DNA and HSV-IgG IC within the ocular microenvironment may contribute to angiogenesis and inflammation observed in HSK, and cytokines and TLRs may be potential targets for intervention.