The goal of this work is to better define T-lymphocyte function in autoimmune illness and following the transplantation of allogeneic organs, so as to support the design of novel therapies to regulate the immune response. In particular, we have focused on:(1) The biochemical signaling mechanisms utilized by various T cell costimulatory molecules.(2) The role of cytokines and other immunomodulating molecules during autoimmune illnesses and the immune response following the transplant of an allograft.In most circumstances, the adaptive immune response has developed to appropriately respond to pathogens while ignoring autologous tissues. Two overlapping models are thought to explain the checks-and-balances that regulate the immune response: (1) central tolerance, whereby T cells undergo an education process within the thymus that results in a repertoire that can distinguish between self and non-self and (2) peripheral tolerance, which regulates self-reactive T cells that escape the thymic education process. Peripheral tolerance, in turn, is thought to be mediated via processes called deletion, anergy, ignorance, or suppression. A failure in either the central or the peripheral tolerance mechanisms can result in autoimmune disorders. T lymphocytes regulate immune responses by signaling other cells through direct contact via surface receptors or by the production of small pleotrophic proteins termed cytokines. The two-signal model of T cell activation proposes that activation requires signaling through two different kinds of receptors. """"""""Signal one""""""""; resulting from antigen recognition via the antigen specific T cell receptor (TCR), and """"""""signal two"""""""" resulting from a second class of receptors called costimulatory receptors. It is now known that costimulatory receptor signaling is the result of highly specific interactions between co-receptors on effector cells and ligands expressed by target cells. Some of these events result in activation while others lead to T cell inhibition. Aside from the TCR and costimulatory receptors, other receptors involved in the immune response increase the binding avidity of antigen presenting cell (APC) and T cells, and thereby enhance signals generated through the TCR. They also can provide synergistic signals that modulate T cell expansion and effector function. Often the functional outcome depends on a balance between opposing effects mediated through these receptors. For instance, two T cell costimulatory receptors (CD28 and CD152) both interact with the same APC counter-receptors and yet mediate opposite responses. The signal transduction pathways initiated by these important costimulatory receptors (CD28 and CD152) are not well defined. Co-ligation of CD3 (to deliver signal one) and CD28 results in T cell activation and the induction and stabilization of cytokine and cell-survival proteins. In contrast, co-ligation of CD3 and CD152 inhibits T cell responses by blocking IL-2 production and cell-cycle advancement. Recently another T cell costimulatory molecule, CD154, has been shown to play an integral role in immune regulation. CD154, a molecule with a wide but tightly controlled distribution, impacts T and B cell function, antigen presentation, and chemotaxis. Agents that modulate the CD28 signaling pathway prolong allograft survival and regulate autoimmunity. However, these treatments do not induce complete tolerance. Studies demonstrating that agents modulating CD154 signaling pathways influence both APCs and T-cells (activation and effector functions) have suggested that this pathway could be manipulated to suppress potentially undesirable immune responses. We have utilized strategies to modulate costimulatory signals to prevent rejection of primate renal allografts using CD152-Ig and an anti-CD154 monoclonal antibody. Further, whereas CD28 modifying reagents alone were unsuccessful, anti-CD154-treatment resulted in sustained graft survival. Further, the addition of additional immunosuppressive agents abrogated the anti-CD154 affect. This fiscal year, we performed experiments to better define CD154 signaling using isolated human T cells. We demonstrated that costimulating human CD4+ T cells with anti-CD3 plus anti-CD154 (in the absence of CD28 signaling) resulted in transient proliferation and activation. This CD154-mediated T cell activation was characterized by the increased expression of activation and adhesion molecules and by the enhanced production of immunomodulatory cytokines. Most strikingly however, many of these anti-CD3/CD154 stimulated T cells died by apoptosis within three days. The paradox of first allowing and then failing to sustain T cell proliferation may suggest a critical role for CD154 in driving short term T cell effector responses, while not leading to immunological memory development. We also participated in studies to better understand the molecular signaling mechanisms for the T cell co-receptors CD152 and ICOS (inducible costimulatory molecule). Cytokines are potent immunomodulatory molecules that act as mediators of inflammation and the immune response. Cytokine production is under genetic control. This is evidenced by the identification of polymorphisms in cytokine gene regulatory regions that correlate with intra-individual variations in actual cytokine production. As these polymorphisms segregate independently, each person can be a mosaic of high-, intermediate-, and low-producing phenotypes. Cytokine polymorphisms have been implicated in a number of diseases including type-1 diabetes. Also, heritable cytokine gene allelic differences have also been shown to account for differences in allograft survival. A number of techniques have been utilized to analyze the level of gene transcript message or protein in a transplanted graft. These techniques have been limited by the small amount of tissue available for analysis and uncertain assay sensitivity. We therefore initiated a comprehensive analysis of cellular targets thought to play a role in renal allograft survival. We analyzed cytokine gene polymorphisms, their cytokine transcripts, and the expression of a wide range of targets relevant to cell survival, transcription regulation, tissue inflammatory, and vasoactivation. This allowed us to determine the expression and distribution of genetic polymorphisms in various cytokine genes from DNA extracted from renal allograft transplant patients and controls. Further, we were able to rapidly and accurately quantitate transplantation relevant transcripts in biopsies taken at distinct time points following allograft transplantation. We identified a correlation between ethnicity and polymorphisms in several cytokine genes. In addition, we found that patients requiring renal transplantation differ from the general population with regard to certain cytokine gene polymorphisms. In a similar vein, we hypothesized that an imbalance in cytokine expression may induce an inflammatory response resulting in pancreatic beta cell destruction. Thus, we are also studying the inheritance patterns of cytokine polymorphisms in patients with type-1-diabetes mellitus (T1DM). Finally, genetic variations in CD152 have been reported to confer T1DM susceptibility. We have studied the inheritance of three separate polymorphisms in the CD152 gene in an ethnically diverse population of individuals with and without T1DM. We are currently analyzing the functional effects of these polymorphisms. The identification and characterization of genes that are linked to T1DM may provide an explanation into the mechanisms by which individuals are predisposed to T1DM.
| Ferrant, Janine L; Benjamin, Christopher D; Cutler, Anne H et al. (2004) The contribution of Fc effector mechanisms in the efficacy of anti-CD154 immunotherapy depends on the nature of the immune challenge. Int Immunol 16:1583-94 |
| Hoffmann, S C; Stanley, E M; Darrin Cox, E et al. (2001) Association of cytokine polymorphic inheritance and in vitro cytokine production in anti-CD3/CD28-stimulated peripheral blood lymphocytes. Transplantation 72:1444-50 |
