Following infection with human immunodeficiency virus type 1 (HIV-1), a quasi-steady state is achieved where viral replication appears to be balanced by the immune response and by death (and replacement) of target cell populations. Highly active antiretroviral therapy (HAART) effectively perturbs the steady state by reducing viral loads and unmasks the dynamic characteristics of the CD4+ T cell population; the effects of HAART on HIV-specific CD8+ T cell responses have not been characterized. Direct staining techniques, using soluble tetramers of human leukocyte antigen (HLA) class I complexes with peptides from HIV, allow rapid enumeration of HIV-specific CD8+ T cells and qualitative analyses of many of their phenotypic and functional properties. Preliminary studies of a small number of patients (prior to HAART) reveal frequencies of antigen specific cells often exceeding 1.5% of CD8+ cells, yet in most cases, these cells do not appear to be activated, an unexpected finding given the chronic exposure to antigen. In several cases, antigen-specific cells appear to die following in vitro stimulation with antigenic peptides. The investigator will extend these studies to monitor frequencies and phenotypes of HIV-specific CD8+ T cells as a function of time before and following the initiation of HAART, and to include greater numbers of patients. The prevalence and cause of resting phenotypes will be investigated through phenotypic analysis of cell surface markers, staining of molecules directly responsible for effector function, and analysis of potential negative regulation due to changes in signaling molecule expression or T cell antagonism. Antigen availability will be analyzed by sequencing viral epitopes to monitor CTL escape. The functional potential of the antigen-specific populations will be determined by staining of cultures stimulated with antigenic peptides in vitro for short periods of time. These experiments will reveal the fraction of the populations that proliferate, develop effector function, or are primed to die upon contact with antigen. Finally, the repertoire of the antigen-specific populations as a function of time following viral load reduction will be monitored over time by T cell receptor spectratyping analysis of magnetically sorted antigen specific cells.
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