The long term goal of this project is to understand the role of cytokines in the cellular and molecular mechanisms of liver allograft rejection. During the previous funding period the in situ cytokine profile during human liver allograft rejection was characterized for the first time. The findings indicate that human allograft rejection does not conform to the Th1/Th2 paradigm. Elucidation of the intragraft cytokine profile led to the identification of cellular effector pathways that are operative in human allografts. Furthermore, a novel T cell subset that expresses CD30 and is the primary source of IL-5 and IFN-gamma produced in response to alloantigen has been identified. The hypothesis to be tested is that CD30 expression identifies a population of alloreactive, cytokine-producing T cells that promote graft rejection. In the first specific aim the development, regulation, and function of alloreactive CD30+ T cells will be characterized. Alloactivated CD30+ T cells obtained by fluorescence activated cell sorting and single cell cloning will be utilized to define the cytokine profile of CD4+CD30+ and CD8+CD30+ cells. CD30+ T cells will be added to mixed leukocyte cultures to determine their ability to regulate allospecific proliferative responses, cytotoxic T cell development, and antibody production. Antibody blocking and transfection strategies will define the contribution of the co- stimulatory molecules B7-1 and B7-2 to the development of CD30 cells. The influence of immunosuppression and the form of alloantigen on the development of CD30 T cells will be determined.
Aim 2 will characterize the function of the CD30 molecule. The hypothesis to be tested is that CD30 ligation engages the cytokine gene expression program in alloactivated T cells. CD30+ T cells will be stimulated with agonistic anti-CD30 mAbs and the effect on cytokine gene and protein expression will be determined. Transfection experiments will examine the link between CD30 signaling and cytokine gene promoter activity. Truncation mutants of the CD30 molecule will identify the intracellular regions necessary for induction of cytokine gene expression.
Aim 3 will define the role of CD30+ T cells in allograft rejection utilizing a rat orthotopic liver transplant model. The development and function of alloreactive CD30 cells during graft rejection will be assessed by immunohistochemical staining and by isolation and characterization of CD30+ T cells from rejecting allografts. CD30+ T cells will be depleted by antibody-based approaches, including immunotoxins, and the impact on the cellular and molecular pathways of graft rejection will be assessed. The contribution of CD30+ T cells to graft rejection will be directly determined in an adoptive transfer model. We anticipate elucidation of the function of CD30+ cells and their role in liver allograft rejection. Identification and characterization of an alloreactive immunoregulatory subset with discrete phenotypic markers would enhance the development of therapeutic strategies to achieve specific immunosuppression in organ allograft recipients.
| Chan, Keith W; Hopke, Corwyn D; Krams, Sheri M et al. (2002) CD30 expression identifies the predominant proliferating T lymphocyte population in human alloimmune responses. J Immunol 169:1784-91 |
