The goal of the project is to understand the role of cytokines in the cellular and molecular mechanisms responsible for human liver allograft rejection. The specific objectives are: 1) determine temporal relationship of the local cytokine profile and functional effector cells in human liver allografts; 2) characterize the immunogenicity of specific hepatic elements; 3) define specific cytokine regulatory pathways that lead to alloreactive effector cell development. The sequential cytokine profile in liver allografts will be studied using a molecular-approach. RNA obtained from allograft biopsies will be reverse transcribed, amplified and analyzed for cytokine gene expression using semi-quantitative polymerase chain reaction technology. Immunohistochemistry and in situ hybridization techniques will provide information on functional effector cells within the allograft. Using this methodology we will obtain a longitudinal picture of the soluble mediators and the diversity of regulatory and effector cells present within the allograft in association with defined clinical states. The identification of specific cytokines within the allograft during rejection may 1) aide in differentiation of rejection from other clinical conditions 2) provide a marker for identifying a """"""""pre-rejection"""""""" state 3) facilitate the development of immunosuppression which target specific mediators of rejection and 4) lead to the definition of an immunological environment that promotes longterm graft survival. Studies in vitro will determine the requirements for sensitization by hepatic elements and the cytokines produced in response to hepatic alloantigen. The presence of a specific cytokine in vivo and its production in response to hepatic alloantigen in vitro will be used as supportive evidence for its role in allograft rejection. The effects of immunosuppressive agents on IL-2 and IL-5 gene expression will be examined as well as the role of IL-5 in human CTL generation. In combination, these studies will contribute to our understanding of cytokine pathways in liver allografts and will increase our knowledge concerning the immunological basis of allograft rejection.
