Research Project
2065151
The goal of this proposal is to define the time course of development and the targets of the human immune response to Herpes Simplex Virus (HSV) acquired following genital infection and to determine if these responses vary with clinical disease expression. Due to the predominant role of cellular immunity and, in particular, cytotoxic response in the resolution of herpes infection in both animals and humans, it is essential to define these specific responses to aid in the development of strategies to limit or control disease. The initial emphasis is to characterize the HSV- specific cytotoxic T lymphocyte (CTL) responses in patients with genital HSV with respect to 1) MHC class restriction and phenotype 2) primary HSV target antigens eliciting CTL, and 3) frequency of these CTL to specific antigens. First, autologous EBV-lymphoblastoid B cell lines of fibroblasts infected with HSV-2 will be employed as cytolytic targets to include all HSV gene products in the survey. Second, drug blocks will be used during infection to limit the repertoire of viral gene expression in target cells to select transcriptional classes. Third, specific HSV-2 genes will be expressed in target cells including IE gene products ICPO, ICP4 and ICP27, virion glycoproteins gB2, gD2 and gG2 and an abundant virion tegument protein and transcriptional activator, VP16. The demonstration of HSV- specific, MHC-restricted cytolytic activity in vitro requires antigen prestimulation of the effector cell population. CTL clones will be established to optimize variables that may effect the efficiency of detection and isolation of such effector cells from peripheral blood lymphocytes including 1) choice of antigen presenting cell (APC) for the expansion phase 2) the mode of introduction of HSV antigen into the APC 3) choice of target cell 4) mode of antigen introduction into the target cells 5) level of HSV antigen expression required for effective stimulation and target recognition. After this method development, the frequency of specific CTL precursors in the bulk PBL culture will be assayed by limiting dilution. Antibody responses will also be determined for specific antigens that elicit an abundant CTL response. The ultimate goal is to determine if there are differences in the target specificity and/or the quantity of cytotoxic lymphocytes in individuals with symptomatic vs asymptomatic HSV or variations in the time to develop HSV-CTL post primary infection that correlate with subsequent asymptomatic viral shedding or recurrent disease episodes.
