Identification of immune mechanisms that potentiate the body's own immune function while decreasing viral replication remains a major goal of AIDS therapy. Consistent with this objective, the ling range goal of this proposal is to define how IL-13, a unique cytokine that remains largely unstudied, might act to restore immune responsiveness while decreasing viral load in HIV-1 patients. Our preliminary data indicate the IL-13 can restore IL-12 secretion in HIV-1-infected cells, act in combination with TNF-alpha to protect macrophages from HIV-1 infection and induce the expansion of T-cells secreting IFN-gamma able to protect against de novo infection via putative inhibitory factors. Based on those data, we hypothesize that macrophage-dependent immunoregulation by IL-13 results in both direct macrophage and indirect T-cell antiviral mechanisms while supporting type-1 T-cell expansion. To test this hypothesis, we will examine the mechanism by which IL-13 can maintain virostasis and ultimately a protective or possible virocidal effect in combination with TNF-alpha and the mechanism leading to IL-13's indirect induction of a type-1 T cell expansion able to inhibit HIV-1. We will study the mechanisms responsible for these effects using in vitro culture systems of either primary isolated macrophages or our unique peripheral blood mononuclear cell (PBMC model system cultured with IL-2 sensitive to endogenous T-cell activation pathways. To define the mechanism of action by which combined TNF-alpha and IL-13 can protect macrophages from HIV-1 infection, suggesting a novel mechanism to abolish both the enhanced macrophage infection mediated by IL-13 and the enhanced viral gene expression ultimately with TNF-alpha, we will focus on early events in macrophage infection. Early infection events studied include reverse transcription kinetics, state of integration and initial intracellular location of reverse transcripts to test candidate sit of viralstasis observed in IL-13-treated macrophages and a potential viral clearance when combined with TNF-alpha. T-cell oriented studies in HIV-1-infected PBMC cultured with IL-2 will focus on the mechanism by which IL-13 induces type-1 T-cell expansion and putative inhibitory factors by analyzing: (1) the dependence of immune activating mechanisms mediating IFN-gamma induction (IL-12, CD28/B7), (2) patterns of cytokine secretions in the face of HIV-1 infection, (3) the potential of inducing this effect in the absence of productive infection, (4) viral products as a result of viral gene expression (p24 antigen, reverse transcriptase activity), and (5) the contribution of chemokine (RANTES, MIP-1alpha, MIP-1beta) or IFN- gamma and IL-10 in mediating inhibitory effects. Expanded T-cells within PBMC cultures will be further analyzed for their ability to protect PM1 cells, which are infectable by macrophagetropic or lymphotropic HIV-1 isolates, from infection by our distinctive HIV-1 89.6 green fluorescent protein reporter virus or by HIV-1 IIIB. Our studies will broaden the currently limited understanding of this powerful cytokine displaying various properties of interest for AIDS immunotherapy.
