Virologic and Immunologic Studies of Murine CMV Retinitis

描述（由申请人提供）：随着高效抗逆转录病毒治疗（HAART）的广泛使用，人类免疫缺陷病毒（HIV）感染患者巨细胞病毒视网膜炎的发病率已经下降。然而，尽管这种减少，仍然有艾滋病毒感染的患者仍然有患巨细胞病毒性视网膜炎的风险，因为他们没有/不能遵循抗艾滋病毒药物的规定治疗方案。因此，对巨细胞病毒发病机制的进一步了解有可能转化为对仍有发生巨细胞病毒视网膜炎风险的患者进行预防或改善治疗。在这个应用中，结合体外和体内研究，使用小鼠，小鼠组织，和培养的细胞被提出来解释小鼠巨细胞病毒（MCMV）视网膜炎的几个特征的机制，包括从外部视网膜（通过睫上途径接种病毒，使病毒进入视网膜下空间）到内部视网膜的顺序感染过程，以及从小鼠和人类研究中观察到的巨细胞病毒感染的细胞通常不会发生凋亡。在本应用程序中提出的研究将利用最近在理解细胞凋亡途径和巨细胞病毒如何与宿主相互作用以诱导视网膜损伤方面的进展。第一个具体目标是确定RPE细胞的感染如何诱导视网膜上的细胞凋亡。这一目的将验证MCMV视网膜炎期间，受感染的RPE细胞产生iNOS和NO开始视网膜感染和损伤的级联反应的假设。这一目标也将使我们能够确定TNF-1在RPE初始感染中的作用。第二个特异性目的将确定TNF-1和NO分别通过caspase依赖性和caspase非依赖性途径导致未感染视网膜细胞凋亡的程度。本研究旨在验证caspase依赖性和caspase非依赖性细胞凋亡在MCMV视网膜病变中的作用。第三个特异性目的将确定为什么MCMV感染的视网膜细胞不发生凋亡。本研究旨在验证MCMV即时早期蛋白3 （IE-3）通过上调细胞flice抑制蛋白（c-FLIP）在保护感染细胞免于凋亡中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，可能适用于新的治疗方法来预防视网膜巨细胞病毒感染或减少人类患者的视网膜损伤。公共卫生相关性：随着高活性抗逆转录病毒治疗（HAART）的广泛使用，人类免疫缺陷病毒（HIV）感染患者巨细胞病毒视网膜炎的发病率有所下降。然而，尽管这一数字有所下降，但仍有感染艾滋病毒的患者仍有患巨细胞病毒视网膜炎的风险，因为他们没有/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耐药性。对巨细胞病毒感染发病机制的进一步了解，有可能转化为对仍有巨细胞病毒视网膜炎发展风险的患者进行预防或改善治疗。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，这可能适用于新的治疗方法，以预防视网膜感染和/或减少视网膜损伤。

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.