VARICELLA ZOSTER VIRUS--T CELL/SKIN TROPISMS AND IMMUNIT

水痘带状疱疹病毒（VZV）引起水痘和带状疱疹。我们的目标是提高对这种常见病原体如何引起疾病以及自然免疫和疫苗诱导免疫所提供的保护的认识。糖蛋白可能是T细胞和皮肤的宿主范围决定因素，而T细胞和皮肤是VZV感染期间的关键靶细胞。我们的重点是糖蛋白，gI （ORF67）和gE （ORF68）。在体外实验中，将确定在宇宙中产生的gI或gE突变对VZV复制的影响。人类CD4+和CD8+ T细胞或皮肤的传染性将在SCIDhu小鼠VZV发病机制模型中进行体内评估，这揭示了VZV蛋白在组织培养中完全不可缺少的关键作用。使用绿色荧光蛋白（gfp）标记的VZV，还将在胸腺器官培养和II23细胞（一种CD4+ T细胞杂交瘤）中研究T细胞的趋向性。将在MDCK细胞中评估VZV、gI和gE对上皮细胞的影响。疫苗株V-Oka将与其亲本P-Oka进行比较，以确定gE或gI突变是否解释了V-Oka的衰减。在VZV特异性免疫被诱导之前，VZV会感染原生宿主的T细胞并扩散。我们发现VZV干扰细胞表面MHC I类和II类的表达。我们的目标是鉴定允许VZV逃避免疫监视的病毒免疫调节蛋白，并确定皮肤归巢受体是否促进被感染的T细胞运输到皮肤。在自然感染期间，这些机制是否在皮肤部位起作用，将在急性水痘病变的活检中确定。快速获得VZV特异性T细胞反应与轻度水痘和维持潜伏期相关。我们提出用新的方法来测量CD4+和CD8+ T细胞对优势病毒蛋白gE和即时早期被覆/反激活蛋白IE62的应答频率，以解决有关VZV适应性免疫的重要问题。我们将研究自然免疫和疫苗诱导免疫的保护差异，成人水痘疫苗免疫原性的降低，以及随着年龄增长VZV - T细胞反应的下降。细胞内细胞因子测定将定量比较gE和IE62蛋白和肽对CD4+和CD8+的识别。将鉴定适合合成MHC I类和II类四聚体的肽，并用于枚举CD4+和CD8+亚群中的VZV特异性应答T细胞。这些对水痘病毒发病机制和免疫的平行研究与改进水痘减毒活疫苗的实际意义直接相关。

Varicella-zoster virus (VZV) causes varicella and herpes zoster. Our goal is to improve knowledge about how this common pathogen causes disease and about protection provided by natural and vaccine-induced immunity. Glycoproteins are likely to be host range determinants for T cells and skin, which are critical target cells during VZV infection. Our focus is on glycoproteins, gI (ORF67) and gE (ORF68). The effect of gI or gE mutations made in cosmids, on VZV replication will be determined in vitro. Infectivity for human CD4+ and CD8+ T cells or skin will be assessed in vivo in the SCIDhu mouse model of VZV pathogenesis, which reveals critical roles for VZV proteins that are completely dispensable in tissue culture. T cell tropism will also be investigated in thymic organ cultures and II23 cells, a CD4+ T cell hybridoma, using green fluorescent protein (gfp)-labeled VZV. VZV gI and gE effects on epithelial cells will be evaluated in MDCK cells. The vaccine strain, V-Oka, will be compared with its parent, P-Oka, to determine whether gE or gI mutations explain V-Oka attenuation. VZV infects T cells in the naive host and spreads before VZV specific immunity is induced. We have found that VZV interferes with cell surface expression of major histocompatibility (MHC) class I and class II. Our goals are to identify viral immunomodulatory proteins that allow VZV to escape from immune surveillance and to determine whether skin homing receptors facilitate transport of infected T cells to skin. Whether these mechanisms function at skin sites during natural infection will be determined in biopsies from acute varicella lesions. Rapid acquisition of VZV specific T cell responses correlates with mild varicella and maintenance of latency. We propose to address important questions about adaptive VZV immunity with new methods to measure CD4+ and CD8+ T cell responder frequencies against dominant viral proteins, gE and the immediate early tegument/transactivating protein, IE62. We will examine differences in protection afforded by natural and vaccine-induced immunity, diminished immunogenicity of varicella vaccine in adults, and declining VZV T cell responses with aging. Quantitative comparisons of CD4+ and CD8+ recognition of gE and IE62 protein and peptides will be made using intracellular cytokine assays. Peptides appropriate for synthesis as MHC class I and class II tetramers will be identified and used to enumerate VZV specific responder T cells in CD4+ and CD8+ subsets. These parallel investigations of VZV pathogenesis and immunity are directly linked by their practical relevance for improving live attenuated varicella vaccines.