Virologic and Immunologic Studies of Murine CMV Retinitis

描述(申请人提供)：随着高效抗逆转录病毒疗法(HAART)的广泛使用，人类免疫缺陷病毒(HIV)感染患者的巨细胞病毒视网膜炎的发病率已经下降。然而，尽管艾滋病毒感染者的人数有所下降，但他们仍然有患巨细胞病毒视网膜炎的风险，因为他们不/不能遵循抗艾滋病毒药物的规定治疗方案。因此，对CMV发病机制的深入了解有可能转化为仍有发生CMV视网膜炎风险的患者的预防性或改善性治疗。在这一应用中，利用小鼠、小鼠组织和培养细胞的体外和体内研究相结合的方法来破译小鼠巨细胞病毒(MCMV)视网膜炎的几个特征的机制，包括从视网膜外部(通过睫状上途径接种病毒后，病毒可以进入视网膜下腔)到视网膜内的顺序感染过程，以及来自小鼠和人类的研究结果，即巨细胞病毒感染的细胞通常不会发生凋亡。这项申请中提出的研究将利用最近在了解细胞凋亡途径和巨细胞病毒如何与宿主相互作用导致视网膜损伤方面的进展。第一个特定的目标将确定RPE细胞感染如何诱导覆盖在上面的视网膜中的细胞凋亡。这一目标将检验一种假设，即在MCMV视网膜炎期间，受感染的RPE细胞产生iNOS和NO开始级联视网膜感染和损害。这一目标也将使我们能够确定肿瘤坏死因子-1在RPE初始感染过程中的作用。第二个特定目标将确定肿瘤坏死因子-1和一氧化氮分别通过caspase依赖和非caspase途径在多大程度上导致未感染的视网膜细胞凋亡。这一目标将检验这一假说，即在MCMV视网膜炎期间，caspase依赖和caspase非依赖的细胞凋亡都有助于视网膜病理。第三个特定目标将确定为什么感染MCMV的视网膜细胞不会发生凋亡。这一目标将验证MCMV立即早期蛋白3(IE-3)通过上调细胞内FLICE抑制蛋白(c-FLIP)在保护感染细胞免受凋亡方面发挥重要作用的假设。这些研究结果将为MCMV感染视网膜的机制提供新的信息，为预防巨细胞病毒感染或减轻人类视网膜损伤提供新的治疗方法。公共卫生相关性：随着高效抗逆转录病毒疗法(HAART)的广泛使用，人类免疫缺陷病毒(HIV)感染患者中巨细胞病毒视网膜炎的发病率有所下降。然而，尽管艾滋病毒感染者的人数有所下降，但他们仍然有患巨细胞病毒视网膜炎的风险，因为他们不/不能遵循抗艾滋病毒药物的规定治疗方案，或者他们的病毒具有抗药性。对巨细胞病毒感染发病机制的了解增加，有可能转化为预防或改善仍有患巨细胞病毒视网膜炎风险的患者的治疗。这些研究结果将为巨细胞病毒感染视网膜的机制提供新的信息，为预防视网膜感染和/或减少视网膜损伤提供新的治疗方法。

DESCRIPTION (provided by applicant): The incidence of cytomegalovirus retinitis has decreased in human immunodeficiency virus (HIV)-infected patients concomitant with the widespread use of highly actively antiretroviral therapy (HAART). However despite this decrease, there are HIV infected patients who are still at risk for CMV retinitis because they do not/cannot follow the prescribed therapeutic regimen of anti-HIV drugs. Therefore, an increased understanding of the mechanisms underlying the pathogenesis of CMV has the potential for translation into preventative or ameliorative treatments in patients who are at still at risk for developing CMV retinitis. In this application, a combination of in vitro and in vivo studies using mice, mouse tissues, and cultured cells are proposed to decipher the mechanism of several features of murine cytomegalovirus (MCMV) retinitis including the process of sequential infection from the outer retina (following inoculation of virus via the supraciliary route which allows the virus to gain access to the subretinal space) to the inner retina and the observation from both mouse and from human studies that cytomegalovirus infected cells usually do not undergo apoptosis. The studies proposed in this application will take advantage of recent advances in understanding apoptotic pathways and in how cytomegalovirus may interact with the host to induce retinal damage. The first specific aim will determine how infection of RPE cells induces apoptosis in the overlying retina. This aim will test the hypothesis that production of iNOS and NO by infected RPE cells begins the cascade of retinal infection and damage during MCMV retinitis. This aim will also allow us to determine the contribution of TNF-1 during initial infection of the RPE. The second Specific Aim will determine to what extent TNF-1 and NO cause apoptosis of uninfected retinal cells via caspase-dependent and caspase-independent pathways, respectively. This aim will test the hypothesis that caspase-dependent as well as caspase-independent apoptosis contributes to retinal pathology during MCMV retinitis. The third Specific Aim will determine why MCMV infected retinal cells do not undergo apoptosis. This aim will test the hypothesis that MCMV immediate early protein 3 (IE-3) plays an important role in protecting infected cells from apoptosis by upregulating the cellular FLICE-inhibitory protein (c-FLIP). The results of these studies will provide new information about the mechanisms of MCMV infection of the retina which may be applicable to new therapeutic approaches to prevent cytomegalovirus infection of the retina or to reduce retinal damage in human patients. PUBLIC HEALTH RELEVANCE: The incidence of cytomegalovirus retinitis has decreased in human immunodeficiency virus (HIV)-infected patients concomitant with the widespread use of highly actively antiretroviral therapy (HAART). Despite this decrease however, there are HIV infected patients who are still at risk for cytomegalovirus retinitis because they do not/cannot follow the prescribed therapeutic regimen of anti-HIV drugs or their virus becomes drug resistant. Increased understanding of the mechanisms underlying the pathogenesis of cytomegalovirus infections has the potential for translation into preventative or ameliorative treatments in patients who are at still at risk for development of CMV retinitis. Results of the proposed studies will provide new information about the mechanisms of cytomegalovirus infection of the retina which may be applicable to new therapeutic approaches to prevent retinal infection and/or to reduce retinal damage.