Cellular and molecular mechanisms involved in T cell-mediated autoimmunity against immunologically privileged retinal antigens are being studied. The questions are aimed at elucidating the natural development and maintenance of self-tolerance to retinal antigens, and defining the processes that lead to their pathological breakdown. The goal is to use this knowledge for designing novel and rational strategies for immunotherapy. The experimental approaches utilize the model of experimental autoimmune uveoretinitis (EAU), which resembles immune-mediated uveitic diseases in humans that can lead to blindness. EAU is induced in mice and rats by immunization with retinal antigens such as IRBP, Arrestin, or their component peptide epitopes, or by infusion of cultured lymphocytes that recognize these antigens. The mechanisms controlling disease susceptibility and pathogenesis are being defined at the genetic, developmental, and immunological levels. Some recent findings are described ahead.The nature and extent of tolerance to immunologically privileged self antigens is poorly understood. To investigate whether taking a retinal antigen out of its immunologically privileged status will affect the ability of the host to develop EAU, we prepared mice that express part of the IRBP gene, under control of the MHC class II promoter, in most tissues of the body. Using this model, we found that extraocular expression of a privileged retinal antigen enhances tolerance and abolishes the ability to develop EAU in response to immunization with an epitope contained in the transgene. However, cultured T cells from wild type mice were still able to induce disease, implying that the level of antigen expression sufficient to tolerize the naive endogenous T cell repertoire is insufficient to tolerize primed uveitogenic effector T cells in the periphery. By using transplantation of bone marrow and thymus cells between the IRBP-transgenic, wild type, and IRBP knockout animals (which are also available) it will be possible to create mice expressing IRBP selectively in the thymus or in the peripheral tissues. This will permit to dissect the relative contributions of central and peripheral tolerance mechanisms to susceptibility, and to define the T cell repertoires involved. EAU, similarly to uveitis in humans, is genetically controlled and is dependent on MHC as well as on non-MHC genes. We study the genetic control of EAU using two approaches: defining genetic markers associated with susceptibility in genetically defined rodents and correlating them with known immunologically relevant genes, and using HLA-transgenic mice, in which murine MHC genes have been replaced by human MHC genes, to create a humanized model of EAU. The first approach identified 3 chromosomal regions affecting EAU susceptibility in rats, which co-localize with a number of immunologically relevant loci. Some of these represent loci that produce or regulate cytokines and responses to cytokines. The second approach demonstrated that presentation of retinal antigens by MHC molecules can lead to retinal disease, and is being used to define the specific epitopes that being recognized. This might help to define antigenic specificities involved in human uveitis. Cytokine response patterns play an important role in determining susceptibility. The susceptible, but not the resistant, individual has a cytokine response pattern polarized towards cellular immunity (Th1). The ability to produce interleukin 12 (IL-12), an innate cytokine that drives Th1 and inflammatory responses, is required to develop EAU. Mice deficient in IL-12, or treated with neutralizing antibodies to IL-12, are unable to develop EAU by immunization with a retinal antigen because they cannot generate the Th1-like pathogenic effector T cell. Paradoxically, an excess of IL-12 aborts disease development by eliciting a negative feedback loop involving massive production of interferon-gamma and nitric oxide, that culminates in apoptosis of the disease-inducing Th1 lymphocytes during their initial exposureto antigen.Thismechanism may serve to prevent recruitment of additional lymphocytes into the effector pool when a strong cellular response, signified by high levels of inflammatory cytokines such as IL-12:00 AM and interferon-gamma, is already present. As part of our effort to develop new approaches to immunotherapy, we designed a strategy for retroviral gene therapy of EAU using the combinedprinciplesof immunoglobulins as tolerogenic carriers and B cells asantigen presenting cells. Activated B cells or bone marrow cells, transduced with a retroviral vector containing a major uveitogenic epitope in frame with mouse IgG1 heavy chain and infused into syngeneic recipients, protected from clinical disease induced by a uveitogenic challenge. Protected mice had reduced antigen-specific responses, but showed no evidence for a classical Th1/Th2 shift or for generalized anergy. Importantly, in mice primed 7 days earlier with a uveitogenic challenge, three infusions of transduced cells reversed the disease process. This epitope-specific protection, effective in naive and in already primed recipients, may provide a protocol for treatment of established autoimmunity.
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