The role of gK in HSV-1 induced corneal scarring

描述(由申请人提供)：众所周知，HSV-1诱导的角膜瘢痕形成(CS)，从而导致HSV-1诱导的角膜失明，是病毒引发的免疫反应的结果。免疫反应的确切特征，包括表达导致CS的经典TCR12抗原受体的潜在有害T细胞效应器的精细特异性，仍然是一个激烈争议的领域。绝大多数非眼部HSV感染要么完全没有症状，要么症状非常轻微，几乎完全无法识别。相比之下，HSV-1重新激活后的症状性眼病是导致角膜疾病和失明的主要原因。在反复感染期间表现出的预先存在的免疫反应的破坏性影响是对临床观察的一个合理解释，即人类CS更有可能发生在反复HSV感染之后，而不是在初次感染上。我们先前(附录1)和初步结果表明，感染HSV-1的小鼠角膜中没有CD8+T细胞。感染的小鼠的眼睛中没有CD8+T细胞，这与感染所用的病毒株或感染的小鼠的株无关。然而，值得注意的是，我们发现，感染了HSV-1糖蛋白之一GK的小鼠，既加剧了CS，又导致角膜中存在CD8+CD25+细胞。在免疫的小鼠中，CS的恶化与GK抗体的存在有关，并且在GK免疫的小鼠的角膜中可以检测到CD8+细胞，但用活病毒疫苗或任何其他已知的HSV-1糖蛋白免疫的小鼠不能检测到CD8+细胞。这种由小鼠现有的免疫反应引起的疾病恶化尤其令人感兴趣，因为它似乎模拟了临床疾病的过程。根据我们现在产生的初步数据，我们假设，在未免疫的小鼠中，病毒gk作用于阻断感染小鼠角膜中CD8+CD25+T细胞的诱导。在免疫的小鼠中，gk特异性抗体结合病毒gk并掩盖其对CD8+T细胞的抑制作用所需的位点。因此，在GK免疫的小鼠中，“抗原掩蔽”不依赖于MHC I类或任何其他病毒基因(S)，而导致感染小鼠角膜中CD8+CD25+T细胞的刺激。随后角膜中CD8+CD25+T细胞数量的增加导致CS的加重。我们建议通过以下特定目的来验证这一假设：目的1：证实抗GK抗体在GK免疫小鼠角膜中诱导CD8+CD25+T细胞起主要作用的假设。目的：证实在gk免疫的小鼠中，角膜中CD8+CD25+T细胞的存在与CS对HSV-1感染的加重有关。目的3：绘制参与CD8+CD25+T细胞刺激和CS1加重的gk区域。HSV-1感染是美国最常见的严重病毒性眼部感染之一，也是病毒致盲的主要原因。HSV-1诱导的CS，也被广泛地称为疱疹间质角膜炎(HSK)，可导致失明；因此，HSV-1是发达国家由于感染性因素导致的角膜失明的主要原因。除了坏死性间质角膜炎外，眼部感染HSV-1还可导致严重的眼部疾病，从睑缘炎、结膜炎、树突性角膜炎到盘状间质水肿。在美国，每年约有50万人患有眼部HSV反复发作，需要看医生、服用药物，严重时还需要角膜移植。我们首次表明，HSV-1的80多个基因中的一个会加剧眼睛感染的小鼠的眼部疾病。我们现在计划确定GK诱导的角膜瘢痕形成小鼠的机制。这些研究应该揭示导致眼病的免疫反应的类型或类型的组合。

DESCRIPTION (provided by applicant): It is well established that HSV-1-induced corneal scarring (CS), and thus HSV-1-induced corneal blindness, are the result of immune responses triggered by the virus. The exact identity of the immune responses, including the fine specificity of the potentially harmful T-cell effectors expressing classic TCR12 antigen receptors that lead to CS remains an area of intense controversy. The vast majority of non-ocular HSV infections are either entirely asymptomatic or so mildly symptomatic as to go almost completely unrecognized. In contrast, symptomatic eye disease after HSV-1 reactivation is a major cause of corneal disease and blindness. The damaging effects of pre-existing immune responses that are manifested during recurrent infections is one rational explanation for the clinical observation that CS in humans is much more likely to occur following recurrent HSV infections rather than on primary infections. Our previous (Appendix 1) and preliminary results demonstrate an absence of CD8+ T cells in the cornea of mice infected with HSV-1. This absence of CD8+ T cells in the eyes of infected mice was observed irrespective of the strain of virus used for infection or the strain of mouse infected. Notably, however, we have found that infection of mice that have been immunized with one of the HSV-1 glycoproteins, gK, both exacerbates CS and results in the presence of CD8+CD25+ cells in the cornea. The exacerbation of CS in the immunized mice is associated with the presence of gK antibody and that the CD8+ cells are detectable in the corneas of mice immunized with gK, but not with live virus vaccine or any of the other known HSV-1 glycoproteins. This exacerbation of disease by an existing immune response in mice is of particular interest as it would appear to mimic the clinical disease process. Based on the preliminary data we have now generated, we hypothesize that, in unimmunized mice, viral gK acts to block induction of CD8+CD25+ T cells in the cornea of infected mice. In immunized mice, the gK-specific antibody binds the viral gK and masks the sites required for its inhibitory effects on the CD8+ T cells. Thus, "antigenic masking", independent of MHC class I or any other viral gene(s) in gK-immunized mice, leads to the stimulation of CD8+CD25+ T cells in the cornea of infected mice. The subsequent increase in the population of CD8+CD25+ T cells in the cornea leads to exacerbation of CS. We propose to test this hypothesis through the following Specific Aims: Aim 1: Confirm the hypothesis that anti-gK antibody play a major role in the induction of CD8+CD25+ T cells in the cornea of gK-immunized mice. Aim 2: Confirm the hypothesis that, in gK-immunized mice, the presence of CD8+CD25+ T cells in the cornea is responsible for exacerbation of CS on HSV-1 infection. Aim 3: Map the gK region that is involved in stimulation of CD8+CD25+ T cells and exacerbation of CS.HSV-1 infections are among the most frequent serious viral eye infections in the U.S. and are a major cause of viral-induced blindness. HSV-1-induced CS, also broadly referred to as herpes stromal keratitis (HSK), can lead to blindness; thus, HSV-1 is the leading cause of corneal blindness due to an infectious agent in developed countries. In addition to necrotizing stromal keratitis, ocular infection with HSV-1 can cause eye disease ranging in severity from blepharitis, conjunctivitis, and dendritic keratitis, to disciform stromal edema. In the U.S., approximately 500,000 people suffer recurrent ocular HSV episodes annually, requiring doctor visits, medication, and, in severe cases, corneal transplants. We have shown for the first time that one of more than 80 genes of HSV-1 exacerbates eye disease in ocularly infected mice. We now plan to determine the mechanism of gK induced corneal scarring mice. These studies should reveal the type or combination of types of immune response that induces eye disease.